United States

Department of

Agriculture

Forest Service

Southern Forest

Experiment Station

New Orleans,

Louisiana

General Technical Report

SO-106

September 1994

Tree Seed Technology

Training Course

E 7: Bonner, J. A. Vozzo, W. W. Elam, and S. B. Land, Jr.
SUMMARY

This manual is intended primarily to train seed collectors, seed-plant managers, seed

analysts, and nursery managers, but it can serve as a resource for any training course in forest

regeneration. It includes both temperate and tropical tree species of all intended uses. The

manual covers the following topics: seed biology, seed collection, seed handling, seed-quality

evaluation, seed protection, seed basics for nurseries, and seed programs. It also includes a

course evaluation questionnaire and practical exercises.

All parts of the course will not be suitable for all groups. The instructor can choose the

pertinent material for presentation. The scope of material is wide enough to serve many

purposes.
Introduction ..................................................................

Official Welcome .....................................................

Introduction of Instructors and Students ..............................

GoalsoftheCourse ..................................................

Scope ...............................................................

Sources of Information and Technical Help. ............................

Group Assignments ..................................................

Presentations ........................................................

Administrative Matters. ..............................................

Biology ......................................................................

I . Flowering, Pollination, and Seed Maturation. ...........................

II. SeedDormancy ......................................................

III. Germination ........................................................

Collection ....................................................................

I . Genetics and Seed Source. ............................................

II. Production ..........................................................

III. Collection Operations. ................................................

IV Maturity ............................................................

. Postharvest Care ....................................................

Handling .....................................................................

I . Drying and Extracting ...............................................

II. Cleaning and Upgrading. .............................................

III. Storage Principles ...................................................

IV Storage Applications .................................................

Evaluating Quality ............................................................

I. Sampling ...........................................................

II. MoistureContent ....................................................

III. Purity and Weight ...................................................

IV Germination lbsts ...................................................

. Rapid Tests: Cutting, Vital Stains,

Excised Embryo, and Hydrogen Peroxide. ............................

VI. Rapid Tests: X Rays and Leachate Conductivity. ........................

VII. Vigor Tests ..........................................................

Protection ....................................................................

I. Insects ..............................................................

II. Pathogens ...........................................................

Basics for Nurseries ...........................................................

I. ProductionSystems ..................................................

Seed Programs ...............................................................

I. National Programs. ..................................................

II. SeedCenters ........................................................

III. Labeling and Certification ............................................

IV Germplasm Conservation. ............................................

V Applied Research ....................................................

Conclusion ...................................................................

Laboratory Exercises ..........................................................

Literature Cited ..............................................................

Appendix .....................................................................

1

2

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2

2

4

4

4

5

6

15

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23

24

27

29

37

44

49

50

52

55

59

65

66

68

70

72

81

87

91

97

98

100

103

104

107

108

113

116

120

122

124

125

139

145

The Seed Technology Thining Course

Instructor’s Manual

F. T. Banner, J. A. Vozzo, W. W. Elam, and S. B. Land, Jr,

F.T. Bonner and J.A. Vozzo are supervisory plant physiologist and research plant physiologist, respectively, US. Department of Agriculture,

Forest Service, Southern Forest Experiment Station, Starkville, MS 39759; W.W. Elam and S.B. Land, Jr., are professors of forestry, Mississippi

State University, Mississippi State, MS 39762. In cooperation with: Forest Tree Seed Program, Department of Forestry, Mississippi Agricultural

and Forest Experiment Station, and Mississippi State University.

.. . 111


OFFICXAL WELCOME

Introductory remarks and welcoming speeches from

the host institution and government representatives

come first. Take care of all administrative matters and

housekeeping items at this time. Be sure to include an

orientation to the site and facilities. A map handout is

desirable.

INTRODUCTION OF INSTRUCTORS

AND STUDENTS

First instructors, then students, stand and tell the

group:

a. Name

b. Affiliation and title (if any).

c. Job assignment at home (e.g., agroforestry or traditional forestry).

d. What they hope to learn in this course (skills,

species information, personal contacts, etc.).

GOALS OF Tm COURSE

The objective of this course is to provide a basic

understanding of the following topics:

A.

B.

C.

D.

E.

F.

G.

H.

I.

J.

K.

L.

M.

N.

0.

P.

QR.

Reproductive cycles, from flowering through seed

germination

Seed origin

Seed collection

Seed maturity

Collection and postharvest care

Seed extracting, cleaning, and conditioning

Insect and disease problems

Seed storage

Seedlot sampling

Tests for moisture, purity, weight, germination,

and vigor

Rapid viability estimates

Seed test results

Seed handling in nurseries

Seed programs

Seed labeling and certification

Germplasm conservation

Seed center design and staffing

Applied seed research (fig. 1) [no equivalent figure in Student Outline1

I

HIGH-QUALITY

SEEDS I

Figure 1. -The relationship of seed technology, basic seed biology,

and high-quality seeds [no equivalent figure in Student

Outline],

All figures and tables in the Instructor’s Manual are

not presented in the Student Outline. In order for the

instructor to reference all figures and tables in the

student book, these differences are noted throughout

the manual.

SCOPE

The emphasis in this course will be on indigenous,

multipurpose tree species suited for forestry and rural

agroforestry. However, exotics are included, because

fast-growing exotics have a definite place in forestry

programs.

SOURCES OF INFORMATION

AND TECHNICAL HELP

Sources of information and technical help include

journals, reference books, international organizations,

regional organizations, and research institutions.

Journals

Seed Science and Technology

International Seed Testing Association (ISTA)

Beckenholz, P.O. Box 412, CH-8046

Zurich

Switzerland

Journal of Seed Technology

Association of Offkial Seed Analysts (AOSA)

Seed Abstracts

CAB International Information Services

Wallington, Oxon OX 10 8 DE

U K

Agroforestry Abstracts

(same as above)

New Forests

Dr. Mary Duryea, Editor-in-Chief

Department of Forestry

118 N-Z Hall

University of Florida

Gainesville, FL 32611

U.S.A.

Indian Forester

(includes seed technology articles)

Indian Journal of Forestry

(includes seed technology articles)

Journal of Tropical Forest Science

Business Manager

Forest Research Institute of Malaysia

P.O. Box 201

Kepong, 52109 Kuala Lumpur

Malaysia

2
Commonwealth Forestry Review

Commonwealth Forestry Institute

11 Keble Road

Oxford

U K

Pakistan Journal of Forestry

Editor

P.O. Pakistan Forestry Institute

Peshawar, N.W.F.P.

Pakistan

Canadian Journal of Forestry Research

Editor

Forestry Canada

P.O. Box 490

Sault Ste. Marie, Ontario

P6A 5M7

Canada

Forest Science

Society of American Foresters

5400 Grosevenor Lane

Bethesda, MD 20814-2198

U.S.A.

Reference Books

The following reference books provide information and

technical assistance:

Bewley and Black 1982

Chin and Roberts 1980

Murray 1984a, b

Schopmeyer 1974

von Carlowitz 1986

Willan 1985

International Organizations

Food and Agriculture Organization (FAO) of the

United Nations

Forest Resources Development Branch

Forest Resources Division

Via delle Terme di Caracalla

I-00100 Rome

Italy

International Union of Forestry Research Organizations (IUFRO)

Secretariat

Schonbrunn

A-1131 Vienna

Austria

IUFRO Seed Problems Project Group

Current Chair: Dr. D.G. Edwards

Forestry Canada

Pacific Forestry Centre

506 West Burnside Road

Victoria, BC

V8Z lM5 Canada

International Seed Testing Association (ISTA)

ISTA Secretariat

Reckenholz, P.O. Box 412

CH-8046 Zurich

Switzerland

Regional Organizations

F/FRED Coordinating Unit

c/o Kasetsart University

Faculty of Forestry

P.O. Box 1038, Kasetsart Post Office

Bangkok 10903

Thailand

Organization for Economic Cooperation and

Development (OECD)

Directorate for Agriculture and Food

Paris

France

Research Institutes

ASEAN-Canada Forest Tree Seed Centre

Mauk Lek, Saraburi

Thailand

Nitrogen Fixing Lee Association

P.O. Box 680

Waimanalo, HI 96734

U.S.A.

International Council for Research in Agroforestry WRAF)

P.O. Box 30677

Nairobi

Kenya

DANIDA Forest Seed Centre

Krogerupvej 3A

DK 3050, Humlebaek

Denmark

Forest Research Centre

P.O. Box HG 595

Highlands, Harare

Zimbabwe

Centre National de Semences Forestieres

PB 2682, Ouagadougo

Burkina Faso

CSIRO Division of Forest Research

P.O. Box 4008

Queen Victoria Terrace

ACT 2600, Canberra

Australia

Commonwealth Forestry Institute (CFI)

University of Oxford

Department of Forestry

Oxford

U K

USDA Forest Service

Tree Seed Research Unit

Forestry Sciences Laboratory

P.O. Box 906

Starkville, MS 39759

U.S.A.

3
USDA Forest Service

National Tree Seed Laboratory

Rt. 1, Box 182-B

Dry Branch, GA 31020

U.S.A.

USDA Forest Service

Institute of Tropical Forestry

University of Puerto Rico, Agricultural Experiment

Station

P.O. Box 25000

Rio Piedras, PR 00928-2500

U.S.A.

Centro Agronomieo ‘Ikopical de Investigation y

Ensenanza (CATIE)

Turrialba

Costa Rica

Petawawa National Forestry Institute

Box 2000

Chalk River, Ontario

KOJ 1JO Canada

GROUP ASSIGNMENTS

If library and study facilities are available to the

trainees, a short report should be assigned. A wide

range of topics is possible, but those of a general nature

are best. A lot may depend on the participants’ level of

education and training and the amount of free time

available to them.

Suggested Topics

Species reports on how to collect, clean, store, and

test seeds of a designated species. The information can

be developed during the course and may encourage

classroom participation by the trainees. This topic is

appropriate for technicians.

Country or province status reports on the current

status of tree seed needs and the biggest obstacles to

filling them in the trainee’s country, province, state,

village, and so forth. This topic is appropriate for

administrators.

Collection plans for a species or a group of species to

supply seeds for a country or province planting program of a designated size.

PRESENTATIONS

Near the end of the course, reports should be presented to the class, four or five per day or all in a special

period. Oral presentations (10 minutes maximum)

should be emphasized, but written reports should be

required with at least an outline of the material.

ADMINISTRATIVE MAWERS

Take time to explain administrative matters, such as

currency exchange, fee payment, housing, meals, transportation, coffee/tea breaks, class schedules, and so on.

The local administrator of the host agency or institution should convey this information. During this

period, also preview any scheduled field trips, tours, or

shopping trips. Tell when, where, what to wear, and so

on. Finally, allow a short time for questions from the

group on any topic related to the course. Give everyone

a chance to understand how the course will be conducted.

4

I. Flowering, Pollination, and Seed

Maturation

A. Introduction

Knowledge of the seed biology of a tree species

is essential to successful seed production and

handling. The sexual life cycle must be known

to plan for genetic improvement, production,

collection, conditioning, storage, and planting of

the seeds.

B. Objectives

1 . Define common terms used to describe life

cycles of plants.

2. Describe the general sexual cycle, flower

structure, seed structure, and origin of the

fruit of gymnosperms.

3. Describe the general sexual cycle, flower

structure, seed structure, and origin of the

fruit of angiosperms.

4. Identify primary differences between

angiosperm and gymnosperm sexual cycles.

5. Describe the general development of fruits

and seeds.

C. Key Points

The following points are essential for understanding flowering, pollination, and seed maturation:

1 . A plant’s life cycle is the time required to

grow from zygote to seed production; there

are two developmental cycles- a sexual

cycle and an asexual cycle.

2. Knowledge of the sexual cycle is required

for:

a. tree-breeding programs

b. seed orchard management

c. seed collection

d. seed conditioning and storage

e. nursery management

3. The gymnosperm life cycle follows this

order:

a. naked seed

b. seedling

c. mature sporophyte

d. strobili (cones)

e. microspore and megaspore mother cells

f. meiosis

g. microspores and megaspores

h. male and female gametophytes

i. pollination

j. single fertilization

k. zygote and gametophytic tissue

1. embryo

m. naked seed on ovulate cone scale

4. The angiosperm life cycle differs from the

gymnosperm life cycle in having:

a. seeds enclosed in fruit (ripened ovary)

b. true flowers rather than strobili

c. double fertilization

d. triploid endosperm tissue rather than

haploid female gametophytic tissue in

the seed

D. Definition of Terms

1 . Life cycle-the time required to progress

from zygote to seed production. There are

two possible developmental cycles-the sexual cycle involving seeds (used in regeneration of “high forest” systems) and the

asexual cycle involving vegetative propagules of coppice forestry (used in

regeneration of “low forest” systems).

2. Genotype- the genetic makeup of a cell

nucleus or an individual.

3. Phenotype-the external appearance of

an organism; it is an expression of the genotype interacting with the environment.

4. Mitosis-nuclear (and usually cellular) cell

division in which the chromosomes duplicate and divide to produce two nuclei that

are identical to the original nucleus.

5. Meiosis-two successive nuclear divisions

in which the chromosome number is halved

and genetic segregation occurs; it occurs in

reproductive cells during sexual reproduction.

6. Pollination-transfer of pollen grains

from the anther or microsporophyll to the

stigma or ovule.

7. Fertilization-fusion of sperm and egg

(and also sperm with two polar nuclei to

form endosperm in angiosperms).

8. Diploid (2N) -two sets of chromosomes in

a cell nucleus. Somatic cells have two sets

and are diploid (except for polyploid plants).

9. Haploid (1N) - one set of chromosomes in

a cell nucleus. Germ cells (egg and sperm

cells) from a diploid plant are haploid.

10. Fruit-a ripened ovary, sometimes including accessory flower parts, that surrounds the seed in angiosperms. The term

“fruit” is used to refer to any seed-bearing

structure; thus, it can include the ovulate

cone of fleshy aril of the conifers.

11. Seed- a ripened ovule that consists of an

embryo, its stored food supply, and protective coverings. The term “seed” is also

often used to include the fruit wall of oneseeded, dry, indehiscent fruits, such as

achenes and nuts. The “seed of commerce”

is the reproductive structure as bought and

sold. It may be multiseeded in some species.

Examples include Tectona and Liriodendron.
12. Mature seed-a seed that can be removed

from the tree without impairing the seed’s

germination.

E. Life Cycles

An understanding of life cycles is needed

because:

1. Sexual and asexual systems reproduce

genetically different populations.

2. Knowledge of the asexual cycle is needed

before vegetative propagation can be used

for coppice systems, cloning of selected genotpes, conservation of germplasm, direct

planting of propagules, or enhancement of

flowering in clonai seed orchards.

3. Knowledge of the sexual cycle is needed for

success in:

a. tree breeding

b. seed production

c. seed orchard management

d. seed harvesting and conditioning

e. nursery management

f. planting and subsequent management

of forest stands

F. Angiosperm and Gymnosperm Sexual Cycles

1 . All tree species are seed-producing plants

(division, Spermatophyta) and belong to

either the class Gymnospermae or

Angiospermae.

a. The true flowering plants of angiosperms have seeds enclosed in carpels.

b. The coniferous gymnosperms have

seeds borne naked on scales arranged

spirally on a central axis to form a cone.

c. Seeds of nonconiferous gymnosperms

are borne singly, each enclosed in a

fleshy, arillike covering.

2. Gymnosperm life cycle (fig. 2) [figure 1

in Student Outline]

WINTER

Figure 2. -Life cycle of a gymnosperm (Pinus spp.) (Banner 1991b) [figure 1 in Student Outline].

7
a. Sporophyte - The diploid plant (tree)

that arises from the zygote.

b. Strobilus or cone- Using a pine from

the Conifer-ales as an example:

(1) Reproductive short shoot (not a

“flower’‘-a term that should be

limited to angiosperms).

(2) Staminate cone (male) -axis with

spirally arranged microsporophylls

and two microsporangia on the bottom side of each sporophyll.

(3) Ovulate cone (female) -axis with

spirally arranged bracts and

ovuliferous scales, Bvo ovules are

located on the upper side near the

base of each scale.

(4) Gymnosperms may be either monoecious (female and male strobili on

same tree) or dioecious (tree has

only one sex).

c. Meiosis and gametophytes

(1) Diploid microspore mother cell in

microsporangium undergoes meiosis to produce four haploid microspores (pollen grains). After

pollination, the pollen grain germinates to produce a six-nucleate male

gametophyte, two sperm nuclei, two

generative nuclei, tube nucleus, and

stalk nucleus).

(2) Diploid megaspore mother cell in

the megasporangia of the ovule

undergoes meiosis to produce four

haploid megaspores, of which three

deteriorate. The remaining megaspore undergoes many mitotic

nuclear divisions, has a free nuclei

stage, and finally forms cell walls to

produce a haploid female gametophyte (archegonia with egg and

gametophytic tissue).

d. Fertilization-A sperm cell from the

male gametophyte unites with the egg

ceI1 to form the diploid zygote (single

fertilization).

e. Seed (fig. 3) [figure 2 in Student Outlinel

(1) Develops from the fertilized ovule.

(2) Contains an embryo (cotyledons,

hypocotyl, radicle) (2N from sexual

recombination), a seedcoat (2N of

mother plant, from integuments),

storage tissue (1N from female

gametophyte), and sometimes a

seed wing (2N from mother plant,

from upper surface of ovuliferous

scale>. - -

SEEDCOAT

STORAGE TISSUE

(FEMALE GAMETOPHYTE)

COTYLEDONS

HYPOCOTYL

RADICLE

Figure 3. -Cross section of a typical mature gymnosperm seed (Pinus ponderosa) (adapted from Krugman and Jenkinson 1974)

[figure 2 in Student Outline].

8
f. Fruit

(1) Gymnosperms do not have true

“fruits” (matured ovaries).

(2) The types of structures that enclose

gymnosperm seeds are:

(a) dry ovulate cones (e.g., Abies,

Araucaria, Cupressus, Pinus,

and Tsuga)

(b) fleshy, arillike structures

enclosing single seeds (e.g.,

Ginkgo, Taxus, and Torreya)

(c) berrylike ovulate cones (e.g.,

Juniperus -one to two seeds

per cone)

3. Angiosperm life cycle

a. Sporophyte - the diploid plant (tree)

that arises from the zygote (same as

gymnosperm).

b. Flower-a short shoot with sterile and

reproductive leaves and a receptacle.

(1) Sterile leaves include:

(a) sepals (outer whorl), collectively,

corolla

(b) petals (inner whorl), collectively,

corolla

(c) perianth, a collective term for

sepals and petals

(2) Reproductive leaves (sporophylls)

include:

(a) stamen (male), composed of

anther (a pollen sac containing

microsporogenous tissue) and

filament

(b) carpel (female), composed of

stigma, style, and ovary. The

ovary is the basal portion of a

pistil that bears the ovules and

its surrounding carpel wall. The

ovule is composed of megasporogenous tissue surrounded

by the nucellus, two integuments, and the micropyle.

(c) pistil, a collective term that

describes visible female structures, whether a single carpel or

a fusion of many carpels.

(3) Receptacle, a stem axis of the flower.

(4) Flowers may have one or both sexes.

(a) Perfect flowers contain both stamens and pistils.

(b) Imperfect flowers contain only

one sex, and the tree may

be either monoecious or

dioecious.

(c) Polygamous includes some perfect and imperfect flowers on

the same tree.

c. Meiosis and gametophytes

(1) Diploid microspore mother cell in

microsporogenous tissue of anthers

undergoes meiosis to produce four

haploid microspores (pollen grains).

After pollination, the pollen grain

germinates to produce a sixnucleate male gametdphyte (same

as for gymnosperms).

(2) Diploid megaspore mother cell in

megasporogenous tissue of ovule

undergoes meiosis to produce four

haploid megaspores, of which three

deteriorate. The remaining megaspore undergoes three mitotic divisions and forms cell walls to produce

a haploid female gametophyte

(eight-nucleate, seven-celled embryo

sac containing three antipodal cells,

two polar nuclei, two synergid cells,

and the egg).

d. Fertilization

(1) A sperm cell unites with the egg cell

to form the diploid zygote (one fertilization).

(2) The second sperm cell unites with

the two polar nuclei (triple fusion)

to ,form a triploid (3N) endosperm

nucleus (second fertilization).

e. Seeds (figs. 4, 5) [no equivalent figures

in Student Outline]

(1) Seeds develop from the doublefertilized ovule.

(2) They contain an embryo (2N from

sexual recombination), seed coat, storage tissue (may be 2N cotyledons,

hypocotyl from embryo, 3N

endosperm from triple fusion, or 2N

perisperm from nucellus of mother

tree), and sometimes other seed coverings (2N of mother tree from remains

of nucellus, 3N from remains of

endosperm, or 2N of mother tree

from attached parts of fruit).

(3) They may be classified as endospermic (embryo is reduced in

size compared with the rest of the

seed) or nonendospermic (embryo is

dominant).

f. Fruits

(1) Fruits develop from the matured

ovary (and sometimes from the

receptacle or perianth); thus, the

ovary has the mother tree’s diploid

genotype.

(2) They enclose the seed (matured

ovule).
COTYLEDON

STROPHIOLE-I \-HILUM

L E

L

Figure 4. -Cross section of a mature seed of Centrolobium paraense (adapted from Tkiviti and others 1990) [no equivalent jigure in

Student Outline].

(3) It is not always possible to separate

the fruit and the seed when the

seedcoat and fruit are joined as a

single unit. In this case, the fruit

itself is referred to as the “seed.”

4. Sexual cycles -The gymnosperm and

angiosperm sexual cycles differ in four

ways:

a. In gymnosperms, seeds are not enclosed

in the ovary, and flowers are unisexual;

in angiosperms, seeds are borne in a

closed ovary, and flowers are perfect or

imperfect.

b. Flowers are true flowers in angiosperms

but are strobili (cones) in gymnosperms.

c. Double fertilization takes place in

angiosperms; single fertilization takes

place in gymnosperms.

d. In gymnosperms, the developing embryo

is nourished by the haploid female

gametophyte; in angiosperms, it is nourished from either diploid cotyledons,

hypocotyl of the embryo, triploid

endosperm, or diploid nucdlar material.

G. Seed and Fruit Development

1. Physical development

a. Angiosperms

(1) Pollination and fertilization trigger:

(a) Formation of embryo and

endosperm within ovules.

(b) Cell divisions and enlargements

in ovary and peripheral tissues

leading to production of fruit.

Most angiosperms flower and

ripen in one growing season

(except for the red oak subgenus

of Quercus).

(2) Legumes have:

(a) simple pistil (one) with a superior ovary having one cavity

(locule). When the pistil

matures, a fruit (pod) is

produced-a single, dry, dehiscent fruit. Seeds develop along

the side of the pod where the

carpel margins join. The other

side is the midrib, where the

pod splits open. A typical seed is

dicotyledonous and lacks endosperm. Each seed is attached to

the pod by the seedstalk

(funiculus). As the seeds leave

the pod, the seedstalk breaks

off, leaving a scar (hilum).

Above the hilum is the raphe;

below is the micropyle (fig. 6)

[figure 3 in Student Outline].

(b) Seedcoats are composed of a histologically dense cuticle, radial

columnar cells, sclerenchy10
- HILU

‘.A)! RADI

M

CLE

SEEDCOAT

. I

. I COTYLEDON

-PERICARP

UNCLE

Figure 5. -Cross section of a mature seed of Cordia alliodora (adapted from Zkivirio and others

1990) [no equivalent figure in Student Outline].

matous cells, lignin, and

osteosclereid cells. These seedcoats are highly impervious to

water and gases. Malpighian

cells internal to the palisade

layer are characterized as sclerenchyma without intercellular

spaces -very dense cells, protecting the seed and denying

water uptake. The “light line”

in the palisade cells is caused by

wax globules, also a water barrier (See fig. 7) [figure 4 in Student Outline].

(3) Definition of Terms

(a) cuticle- waxy layer on outer

walls of epidermal cells

(b) lignin - organic component of

cells associated with cellulose

(c) light line -continuous thin

layer of wax globules

(d) osteosclereid- bone-shaped

sclerenchyma

(e) palisade cells - elongated cells

perpendicular to the coat surface

(f) parenchyma- undifferentiated, live cells

1 1
MICROPYLE

MICROPYLE

I

RAPHE

tilLUM

cm

Figure 6. -External morphology of a typical legume seed of Schizolobium parahybium (adapted from Divirio and others 1990)

[figure 3 in Student Outlinel.

(g) sclerenchyma - thick, lign&d cells

b. Gymnosperms - Fertilization stimulates growth of conelets and development

of the embryo; female gametophytic

tissue is already present when fertilization occurs. Many conifers flower and

ripen seeds in one growing season, but

some require two seasons, and a few

require three seasons (pines require

2.25 years from formation of flower primordia to seed dispersal) (fig. 2) [figure

1 in Student Outline].

2. Physiological development

a. Moisture content increases rapidly after

fertilization and decreases at maturity.

There is evidence that a desiccation

period is required before all germination

and growth enzymes can be synthesized

in orthodox seeds (fig. 8) [no equivalent

figure in Student Outline]. In recalcitrant seeds there is only slight

desiccation before maturity (fig. 9) [no

equivalent figure in Student Outline].

b. Hormone contents are higher where

meristematic activity is greater, that is

in immature (maturing) seeds.

c. Metabolic changes are many; simple

sugars, fatty acids, and amino acids are

converted to proteins, oils, and lipids

(fig. 10) [no equivalent figure in Student

Outline].

3. Classification of mature fruits (see

table 1) [table 1 in Student Outline]

H. Sources

For additional information, see Dogra 1983;

Hardin 1960; Hartmann and others 1983, chap.

3, p. 59-65; Krugman and others 1974; Willan

1985, p. 7-10,13-E.

12
CUTICLE LAYER

EPIDERMIS

PALISADE LAYER

I

“LIGHT LINE”-

INTERCELLULAR SPACE

OSTEOSCLEREID’

SCLERENCHYMA

PARENCHYMA

Figure ‘7. -Partial section through the seedcoat of a hard seed (legume) [figure 4 in Student Outline].

13
- FRESH WEIGHT

--- DRY WEIGHT

---- MOISTURE

I :

JUN. JUL. AUG. SEPT. OCT.

Figure 8. -Seasonal changes in fresh weight, dry weight, and moisture content during maturation of samaras of Fraxinus

pennsylvanica (adapted from Bonner 1973) [no equivalent

figure in Student Outline].

- FRESH WEIGHT

--- DRY WEIGHT

---- MOISTURE

\

-__----- \

\

\

“:xye

/

/

‘.

-.

,

/

JUN. JUL. AUG. SEPT. OCT. NOV

Quercus o/ha

2

2

5 40

Q

3 0

2 0

IO

0

-INSOL. CARBOHYDRATE

--- CRUDE LIPID

Q. nigra

/

0

b e ----- 0

1

. . . . . .

JUN. JUL. AUG. SEPT. OCT

Figure 10. -Changes in insoluble carbohydrate and crude lipid fractions in maturing acorns of Quercus alba and Q. nigra

(adapted from Bonner 1974b, 1976) [no equivalent figure

in Student Outline].

Figure 9.-Seasonal changes in fresh weight, dry weight, and moisture content during maturation of Quercus shumardii

acorns (adapted from Bonner 1976) [no equivalent figure

in Student Outline].

1 4
Table l.-Commonfruit types for woody trees (adapted from Hardin 1960) [table 1 in Student

Outline]

Description Type Example

Simple Fruit (product of single pistil)

Dehiscent walls (splitting naturally)

Product of one carpel

Dehiscing by one suture

Dehiscing by two sutures

Product of two or more carpels

Walls indehiscent (not splitting naturally)

Exocarp fleshy or leathery

Pericarp fleshy throughout

Pericarp heterogeneous

Exocarp leathery rind

Exocarp fleshy

Endocarp a “stone”

Endocarp cartilaginous

Exocarp dry (papery, woody, or fibrous)

Fruit winged

Fruit without wings

One-loculed ovary; thin wall; small seed

Several-loculed ovary; thick wall; large seed

Compound Fruit (product of multiple pistils)

Pistils of a single flower Aggregate Magnolia

Pistils from different flowers (inflorescence) Multiple P&anus

Follicle Zanthoxylum

Legume Acacia, Prosopiq Robinia

Capsule Eucalyptus, Populus

Berry

Hesperidium

Drupe

Pome

Samara

Achene

Nut

Vaccinium, Diospyros

Citrus

Prunus, Vitez, Tectona

Malus, Crataegus

!!kiplochiton, 7krminalia, Acer

P&anus, Cordia

Quercus

II. Seed Dormancy

A.

B.

c.

Introduction

Once seeds have matured, survival of the species requires that they germinate at a time and

place favorable for growth and survival of the

seedlings. The mechanism that prevents germination at undesirable times is called dormancy. The mechanics of seed dormancy must

be known before nursery practices for overcoming dormancy can be developed to ensure timely

germination and uniform growth of the seedlings.

Objectives

1. Describe the different types of seed dormancy.

2. Discuss methods for overcoming seed dormancy, both for germination testing and for

nursery operations.

Key Points

The following points are essential to understanding seed dormancy:

1. To a large degree, dormancy is under

genetic control.

2. Environmental conditions during seed

maturation can influence the degree of dormancy.

3. Seeds can have more than one type of dormancy mechanism.

4.

5.

6.

Postharvest environment can create secondary dormancy.

The distinction between “dormancy” and

“delayed germination” is not always clear.

The least severe treatment to overcome dormancy should be tested first to avoid

damage to the seeds; then increasingly

severe treatments can be tested as needed.

D. Definition of Terms (Bonner 1984a)

1 .

2.

3.

4.

5.

6.

Afterripening - physiological process in

seeds after harvest or abscission that

occurs before, and is often necessary for,

germination or resumption of growth under

favorable environmental conditions.

Dormancy- a physiological state in which

a seed disposed to germinate does not, even

in the presence of favorable environmental

conditions.

Chilling- subjection of seeds to cold and

moisture to induce afterripening.

Prechilling- cold, moist treatment

applied to seeds to hasten afterripening or

to overcome dormancy before sowing in soil

or germinating in the laboratory.

Pretreatment -any kind of treatment

applied to seeds to overcome dormancy and

hasten germination.

Scarification-weakening of seedcoats,

usually by mechanical abrasion or by brief

15
soaks in strong acids, to increase their permeability to water and gases or to lower

their mechanical resistance to swelling

embryos.

7. Stratification-placing seeds in a moist

medium, often in alternate layers, to hasten

afterripening or to overcome dormancy.

Commonly applied to any technique that

keeps seeds in a cold, moist environment, it

is sometimes used to describe warm stratification (warm incubation).

8. Delayed germination- a general term

applied to seeds that do not germinate

immediately but are not slow enough to be

described as dormant.

E. !l)pes of Dormancy

Many different classifications of dormancy have

been used by seed scientists. Because there is no

universal agreement on the subject, anyone can

devise a system. However, most workers accept

the following classifications:

1. Seedcoat (or external) dormancy

a. Impermeability to moisture or gases;

e.g., Acacia, Prosopis, Robinia, and

other legumes

b. Mechanical resistance to swelling

embryo; e.g., Pinus and Quercus.

(Mechanical resistance frequently contributes to seedcoat dormancy but is seldom the primary factor.)

2. Embryo (or internal) dormancy

a. Inhibiting substances usually within

the embryo or surrounding tissues; e.g.,

Fraxinus, Ilex, and Magnolia.

b. Physiological immaturity - Some

enzyme system or crucial metabolite

may not be “in place”; e.g., Juniperus

virginiana. (Supporting data are weak

for this type of dormancy.)

3. Morphological dormancy results from

the embryo not being completely developed

when the seeds are disseminated; additional

growth is required; e.g., Ilex opaca and

some Fraxinus spp. and Pinus spp. in

northern latitudes or high elevations. Morphological dormancy is similar to physiological immaturity.

4. Secondary dormancy results from some

action, treatment, or injury to seeds during

collecting, handling, or sowing; e.g.; Pinus

taeda being exposed to high temperatures

and moisture during storage.

5. Combined dormancy results from two or

more primary factors, such as seedcoat dormancy and embryo dormancy; e.g., Tilia,

which has a very hard seedcoat, plus

embryo dormancy that requires stratification.

6. Double dormancy results from embryo

dormancy in both the radicle and epicotyl.

Double dormancy is difficult to demonstrate

but apparently common in Prunus. In some

Quercus species, radicles are nondormant,

while epicotyls have a dormancy.

F. Overcoming Dormancy

1 . Seedcoat dormancy - Treatment must

increase moisture uptake and gas exchange

and ease radicle emergence. Try the gentlest method first; then increase the severity of the method until success is achieved.

a . Cold water soak-Soak seeds in water at

room temperature for 24 to 48 hours.

This method softens seedcoats and may

remove inhibiting substances in the

outer coverings. Change the soak water

every 12 to 24 hours if inhibitors are

present.

b. Hot water soak-Bring water to a boil,

put seeds in, remove from heat, and

allow to stand until water cools. Hot

water softens the seedcoat, and imbibition occurs as the water cools. Some

workers prescribe a specific period for

the hot water soak, but seed variation

prohibits such specific recommendations

for most species.

c. Hot wire-Use a heated needle or electric woodburner to burn a small hole

through the seedcoats. Hot wire is a

promising new method. Once “burned,”

seeds can be shipped or even stored

without damage; the length of storage is

still being tested, but at least several

months are possible. The method works

on all hard-seeded species, not just

legumes.

d. Acid treatment -Pour a strong mineral

acid over the seeds and mix. (Sulfuric

acid is preferred.) Remove the seeds

after a time determined by trials with

samples, usually 15 to 60 minutes, and

wash thoroughly to remove acid from

the surface. Seeds scarified by water

soaks or acid cannot be returned to storage. Acid treatment has three disadvantages:

(1) Excessive treatment can harm

seeds.

(2) Acid is expensive.

(3) Unskilled workers may be injured.

e. Physical scarification - Crack or break

the hard seedcoats.

(1) Use hand methods (nicking) with
2 .

files, knives, clippers, sandpaper, (4) Inhibitor/promoter balances

etc. change.

(2) Use mechanical methods for large

quantities including motor-driven

scarifiers, cement mixers with

rocks, or the impact machine

recently developed by DANIDA

(Stubsgaard 1986) for Prosopis and

Acacia.

Embryo dormancy- Treatment must

overcome physiological barriers within the

seeds.

a. Stratification (chilling and

prechilling) -Refrigerate fully imbibed

seeds at 1 to 5 “C for 1 to 6 months.

Primarily for Temperate Zone species,

this treatment simulates natural winter

conditions after seeds fall. Many nondormant seeds respond to short stratification periods (1 to 2 weeks) with faster

and often more complete germination.

One benefit that nurseries often overlook is the delay of early germination by

low temperature, which produces a big

flush of germination when high temperature is encountered (table 2) [table 2

in Student Outline]. During stratification:

(1) Imbibition is completed.

(2) Enzyme systems are activated.

(3) Storage foods change to soluble

forms.

b. Incubation/stratification - Some species

respond to a short, warm incubation (15

to 20 “0, followed by cold stratification.

c. Chemical treatment-Some species

respond to chemical stimuli well enough

to justify treatment.

(1) Hydrogen peroxide- Soak for 48

hours in a l-percent hydrogen peroxide solution (Pseudotsuga menziesii).

(2) Citric acid-Soak for 48 hours in

l-percent citric acid solution, followed by go-day stratification

(Juniperus, Tmodium distichum).

(3) Gibberellins -Many species respond

in the laboratory, but field results

are lacking or rare.

(4) Ethylene -Treat seeds for 48 hours

with 0.021 M Ethrel solution

(reported to help germination for

Ricirwdendron rautentenii in South

Africa).

d. Light-Provide light treatments (red/

far-red mechanism) (Betulu).

G. Significance

1 . Survival strategy- Dormancy allows germination during favorable environmental

conditions. It also permits seed survival in

litter or soil for several years, even in the

Tropics.

Table 2. -Recommended prechill periods for nursery sowing of some pines of the Southern United States (Bonner

1991b) [lkble 2 in Student Outline]

Pine

species

Normal sowing*

Fresh seed Stored seed

Early

sowing+

Seed conditions

Deer, dormancy* Low vigor

Pinus strobus

I! taeda

P palustris

l? rigida

I? serotina

P. clausa

var. immuginata

var. clausa

P echinata

I? elliottii

var. elliottii

var. densa

l? glabra

P oirginiana

30-60 6 0

30-60 30-60

0 0

0 O-30

0 O-30

o-15 o-21

0 0

O-15 O-30

0 030

30 O-30

30 3 0

O-30 3 0

&&ill(Days) _____--__-______-_______________________--------

60-90 60-90 30

60 60-90 20-30

. ..+ o-15 0

. . . . . . . . .

. . . . . . 15-30 30-60 0

1530 0

... ... ...

3 0 ... ...

‘Spring sowing when mean minimum soil temperature at seed depth is at least 10 “C

+Early sowing when soil temperatures at seed depth may be below 10 “C.

*Dormancy demonstrated by paired tests or past performance of the seedlot.

Konditions not encountered with this species.

17
2. Genetic factor-Dormancy in many seeds

is under genetic control, particularly those

with seedcoat dormancy because seedcoats

are maternal tissue. Dormancy has been

“bred out” of most agricultural crops.

3. Multiple causes- Many species have

probably evolved with more than one dormancy mechanism; that is, if one fails,

another is in place. Quercus nigra is a good

example, with probably three separate

mechanisms.

4. Environmental influence -Hot, dry

weather during maturation may increase

dormancy, particularly that associated with

seedcoats. Research is just beginning to

show this effect in trees.

H. Sources

For additional information, see Khan 1984;

Krugman and others 1974; Murray 1984b;

Nikolaeva 1967; Willan 1985, p. 17-19, chap. 8.

III. Germination

A. Introduction

The goals of seed technology are successful germination and seedling establishment. The two

major considerations are the physiology of the

seed and the condition of the environment. In

the two preceding sections, seed maturation and

dormancy were considered. In this section,

environmental factors and how they control germination through their interactions with seed

biology will be examined.

B. Objectives

1 . Describe the two types of germination and

their importance in woody plants.

2. Review environmental requirements for germination.

3. Review physiological changes within seeds

that lead to germination.

4. Discuss how seed physiology and environmental factors interact in germination.

C. Key Points

The following points are essential to understanding germination:

1 . The two types of germination are epigeous

and hypogeous.

2. Moisture availability is the primary factor

controlling germination.

3. The effects of temperature and light on germination are strongly related.

4. Constant and alternating temperature

regimes may lead to similar total germination, but germination is usually faster

under alternating regimes.

5. As germination begins, the key to internal

processes is the change from insoluble to

soluble metabolites. Details of such metabolism are beyond the scope of this course.

D. Types of Germination

1. Epigeous (epigeal) germination occurs

when cotyledons are forced above the

ground. by elongation of the hypocotyl (fig.

11) [figure 5 in Student Outline]; e.g.,

Pinus, Acacia, Fraxinus, and Populus.

2. Hypogeous (hypogeal) germination

occurs when the cotyledons remain below

ground while the epicotyl elongates (fig. 12)

[figure 6 in Student Outline]; e.g., JugZans,

Quercus, and Shorea.

3 . In Prunus, both types of germination may

be found (fig. 13) [no equivalent figure in

Student Outline].

E . Environmental Requirements for Germination

The four primary environmental requirements

for germination are moisture, temperature,

light, and gases.

1. Moisture

a. Imbibition is usually considered the

first step in germination; thus, availability of moisture is the first requirement

for germination.

b. Uptake typically occurs in the following

three phases:

(1) A rapid, mainly physical, initial

phase that occurs in dead seeds as

well as live ones.

(2) An extremely slow second phase, the

“flat” portion of the uptake curve.

The greater the dormancy, the

longer this phase takes.

(3) A rapid third phase that occurs as

metabolism becomes very active.

Some evidence suggests that splitting of seedcoats by emerging radicles is required for the most rapid

examples of this phase.

c. The first phase is imbibitional; it

releases energy in the form of heat, displaces gases, and creates great imbibitional pressure via colloidal swelling

(proteins).

d. A minimum state of hydration is needed

within a seed for mobilization of food

reserves and activation of enzyme systems.

e. Minimal requirements for germination

are frequently studied with osmotic

solutions of mannitol or polyethylene

gl ycol .

(1) The best germination may occur at

slight moisture stress (0.005 to

18
HYPOCOTYL

Figure 11. - Epigeal germination sequence of Fraxinus spp. (adapted from Bonner 1974a) [figure 5 in Student Outline].

0.500 bars); zero stress may form a

film of water around the seed, inhibiting oxygen uptake.

(2) Even slightly lowered water potentials will slow, but not stop, germination.

(3) Critical levels of water potential

vary by species (table 3) [no equivalent table in Student Outline]. Germination stops in 12 European

conifers at - 3 to - 12 bars.

2. Temperature

a. It is difficult to separate the effects of

temperature from those of light and

moisture.

b. Woody species usually germinate over a

wide range of temperatures; the germination rate is strongly temperature

dependent. Some Temperate Zone species will eventually germinate in stratification.

c. The upper limit of temperature is

around 45 “C. Many species begin germinating above 40 “C, but seedlings are

abnormal.

d. The lower limit is around 3 to 5 “C

because germination processes will

occur near freezing with emergence of

radicle and plumule; however, few species can produce normal seedlings under

these conditions. Picea mariana (black

spruce) is one that can.

e. Optimum temperatures are as follows:

(1) For Temperate Zone species, alternating regimes of 20 “C! (night) and

30 “C (day) have proved best for

many species; similar results can be

obtained at constant temperatures

of about 25 “C, but germination

rates are almost always greater with

alternating regimes. Amplitude (10

to 12 “0, not cardinal points, seems

to be most important (within

limits>.

(2) For tropical species, although few

critical studies are available, constant temperatures may be best,

although optimums are not necessarily higher than those of temperate species. Examples include:

Azadirachta indica, 25 “C; Bombax

ceiba, 25 “C; Eucalyptus camaldulensis, 30 “C; Leucaena leucocephala, 30 “C; Prosopis cineraria,

19
EPICOT YLS

I’- ti- -

3 /

.L

. .. ;- ._ ‘-2.

c

I cm

0

/

COTYLEDON

J.---:3

\-

c

c

Figure 12. - Hypogeal germination sequence of Quercus spp. (adapted from Olson 1974) [figure 6 in Student

Outline].

20
COTYLEDONS

HYPOGEAL EPIGEAL

Figure 13. -Both epigeal and hypogeal germination as they occur in Prunus spp. (adapted from Grisez 1974) [na equivalent figure in Student

Outline].

J’&le 3. -Critical levels of water potential for germination [no equivalent table in

Student Outline1

Species

Pinus ponderosa

I? eldarica

Populus ciliata

Quercus palustris

Germination

Significantly Effectively

decreased stopped

_____________ Bars’ ___~~~~-~__---

- 4 - 8

- 6 - 12

- 1 - 3

- 5 -20

Reference

Djavanshir and Reid 1975

Djavanshir and Reid 1975

Singh and Singh 1983

Bonner 1968

‘1 bar equals 0.1 MPa.

21
3 .

30 “C; and Tectona grandis, 30 “C.

Other species do as well or even better under alternating temperatures;

e.g., Acacia spp., Cedrela spp., and

tropical Pinus species.

Light

a. Light stimulates germination of many

tree seeds but is necessary for few, if

any. Stratification or high temperature

can sometimes overcome dark inhibition, probably through the phytochrome

system.

b. Phytochrome is a pigment involved in

the photocontrol of germination. It

exists in two reversible forms. One form,

Pr, has a maximum absorption at 6,600

A, while the other, Pfr, has a maximum

absorption at 7,300 A. Less than a second of exposure to red light can supply

the stimulation, which increases as seed

moisture increases. Among trees, it has

been shown to occur in p taeda and

Betula pubescens. The exact mechanisms of this action are still unknown.

The general reaction is as follows:

red light _ ,dormancy removal

Pr< Pfr

far red light

c. In germination testing, minimal light

levels should be 75 to 125 fc (750 to

1,250 lux). Light is usually supplied during testing of all species, even to those

that do not require it or to those that

have been stratified.

4. Gases

a. Respiration requires a certain supply of

oxygen, and the carbon dioxide produced

must be removed. High levels of carbon

dioxide can inhibit germination of most

species, while lowering oxygen can do

the same. Excessive moisture on germination test blotters will retard germination of many species.

b. Some species germinate well in anaerobic conditions, even under water.

c. Oxygen uptake patterns in seeds are

similar to those of moisture.

d. Many aspects of the influences of gases

on germination need to be studied.

F. Internal Physiological Changes

1 . Structural changes - Imbibition is a precursor to necessary metabolism through its

role in restoring structural integrity in

membranes and organelles.

2 . Enzymes - Some systems are present in dry

seeds; others are synthesized as imbibition

proceeds. In some species, a desiccation

period at the end of maturity is necessary

before rehydration to produce all the needed

enzymes. No research on this subject has

been done on tree seeds, but this finding can

probably be assumed for orthodox species.

3. Reserve food mobilization - Generally,

insoluble forms are converted to soluble

forms (in some ways a reverse of maturation trends).

a. Carbohydrates. Amylases are the primary enzyme systems to change starch

to soluble sugars.

b. Lipids. Lipase enzymes are important

for these compounds; they separate fats

into fatty acids and glycerol. Fatty acids

then undergo beta-oxidation to acetyl

coenzyme A, which enters the glyoxylate

cycle, eventually showing up as carbohydrates.

c. Proteins. Some proteins are important

storage foods, but most tree seeds

depend on carbohydrates and lipids.

4. Nucleic acids-These compounds are

closely allied to the new enzymes formed

during germination.

5. ‘lkauslocation- The movement of materials within the embryo is crucial. In some

plants, a stimulus originates from the radicle tip, which controls amylase activity.

There are also other stimuli (possibly hormones) in cotyledons that are necessary for

translocation in some species. These stimuli

have been studied very little in tree seeds.

G. Sources

For additional information, see Bonner 1972,

Mayer and Poljakoff-Mayber 1975, Murray

1984b, Stanwood and McDonald 1989, Willan

1985.

22
23
I. Genetics and Seed Source

A. Introduction

Seed quality involves both the genetic and the

physiological quality of seeds. In this section,

the general principles and methods for selection

of seed source and improvement of seed quality

through genetic selection are presented. Genetic

improvement of seed quality is based on the

seeds’ ability to produce trees that are genetically well suited to the sites where planted and

for the products desired. In later sections, the

physiological quality of seeds will be considered.

Good seeds are those that have both high physiological quality and genetic suitability.

B. Objectives

1. Recognize the importance of seed origin

(provenance) and recommend general rules

for seed movement.

2. Review the advantages and disadvantages

of exotic tree species and interspecific

hybrids for tree improvement.

3: Define factors that must be considered

when a tree improvement program is

initiated.

4. Identify the conditions required for genetic

improvement of tree seeds (genetic gain

concept).

5. Distinguish between a minimum initial

strategy of genetic improvement and a maximum long-term strategy.

6. Identify some terms and concepts of new

biotechnology for genetic improvement.

C. Key Points

The following points are essential to understanding seed source and genetic improvement:

1. A successful tree improvement program

should not be tried in another country or

region without considering desired products

and available sites.

2. Knowledge of phenotype and genotype is

necessary to understand genetic improvement of trees.

3. The genetic gain equation explains the

advantages of one improvement method

over another.

4. Genetic gains can be obtained from selections among species, provenances within

species, and/or trees within provenances.

5. The primary risk of using exotics or nonlocal provenances is planting on unsuitable

sites.

6. Test plantings are the only sure method to

determine genetic quality of seeds.

7. Without results of test plantings, the safest

rule is to use seeds from phenotypically

selected stands or trees in the local provenance for native species or land race for

exotic species.

8. The seed orchard concept has two partsthe breeding program and the production

program.

9. Seed orchard breeding programs involve

progeny tests and selection for the next

advanced generation of genetic improvement.

10. Seed orchard production programs are

managed to maximiie seed production

through protection and cultural treatments.

D. Tree Improvement

1. ‘bee improvement begins with the decision to use artificial rather than natural

regeneration. Tree improvement is the

development and application of genetically

improved trees and intensive cultural practices to increase forest productivity through

artifical regeneration.

2. Wee improvement programs are plans

of action to bring about desired objectives.

The following factors should be considered

when a tree improvement program is initiated:

a. The products desired determine what

species can be used and what traits

should be emphasized when breeding for

genetic improvement.

b. The geographic location and physical

and climatic characteristics of the sites

to be regenerated determine what species and seed sources can be used.

c. Only species and provenances that are

adapted to the planting sites should be

used to avoid failure or substandard performance.

d. Conservation of forest gene resources

should be planned from the beginning to

maintain genetic diversity.

E. Strategies for Genetic Improvement

1. Genetic gain

a. Genetic improvement (genetic gain) is

accomplished by:

(1) Having a population of trees with

genetic differences

(2) Selecting the genetically desirable

trees to serve as parents for production of seeds

b. The amount of genetic gain (R) to he

captured from phenotypic selection of

parent trees for a particular trait can be

expressed as:

R = iVph2

where i = the intensity of selection

h2 = the heritability of the trait

2 4
V, = the amount of phenotypic

variation.

c. Gain can be obtained from selection

among species (R,), selection among

provenances within species {BP), or

selection among individual trees within

provenances (RI). The total gain CRT) is

the sum:

R, = R, + R, + R,

2. Species selection

a. Species-site studies provide information

about the relative performance of different species when planted together on

various sites.

b. Exotic tree species should be used only

when the desired product cannot be

obtained with native species at a comparable cost.

c. Interspecific hybridization has been

used to obtain unique combinations of

valuable traits (combinational hybridization) and to obtain hybrid vigor (e.g.,

Populus and Pinus).

3. Seed source

a. Provenance refers to where the mother

trees were growing and the seeds were

collected. The stand may be indigenous

or nonindigenous. Seed source is the

same as provenance. Origin refers to

where the original progenitors of nonindigenous stands were growing in natural forests and where their genetic

characteristics developed through natural selection. For an indigenous stand of

trees, origin, provenance, and seed

source are the same. Some foresters in

the United States use the term “provenance” in place of “origin” when referring to nonindigenous stands.

b. The largest, cheapest, and fastest gains

in most tree improvement programs can

be made by ensuring the use of adapted,

productive provenances of the desired

species. “Local” sources should be used

until provenance test results are available.

c. Provenance tests should be conducted

during the early stages of a tree

improvement program so that geographic collection and planting zones

can be delineated. Results of provenance

tests include:

(1) Mapping patterns of geographic

genetic variation

(2) Delineating provenance boundaries

(3) Determining provenances best

adapted and most productive for

specific geographic areas.

d. The general results of provenance testing are:

(1) Wide seed transfer is safer near the

center of a species’ range than near

its edge.

(2) Where environmental gradients are

steep, movement of material must

be restricted.

(3) Provenances from harsh climates

(cold or dry) are slower growing but

more hardy because of the stress

factor than provenances from

milder climates.

4. Improvement strategies

a . The initial strategies for a new program

are:

(1) Collect available information.

(2) Select among indigenous tree species.

(3) Select seed production areas within

the “local” seed sources near the

planting site.

(4) Remove phenotypically inferior

trees from seed production areas.

b. The long-term strategies for maximum

and continued gains are:

(1) Collect all existing information.

(2) Select several species for the program, and begin supplemental tests

(species-site, exotic, or interspecific

hybridization) to obtain missing

information on these and other

promising species.

(3) Conduct provenance tests or use

existing provenance tests in the area

to delineate optimal provenances for

the planting sites.

(4) Select the phenotypically “best”

trees from the best provenances.

(5) Establish a first-generation seed

orchard from the phenotypically

selected trees.

(6) Test the progeny of the selected

trees.

(7) Remove genetically poor trees from

the orchard.

(8) Select the best individuals from the

best families in the progeny tests

and place these in a second-generation seed orchard.

(9) Test the progeny of these secondgeneration selections and repeat

steps (61, (7), and (8) above for subsequent generations. Search con25
tinually for new first-generation

selections or for selected clones from

other programs to incorporate into

the program (enrichment).

c. New strategies for genetic improvement

are:

(1) Gene transfer, using recombinant

DNA technology, to insert a desired

gene into a species where it does not

occur.

(2) Cell selection in a cell-suspension

culture to screen for resistant cells

to a pathogen, toxin, or herbicide,

and developing these cells into

mature plants by tissue culture

techniques.

(3) Fusion of protoplasts ‘(without cell

walls) to create a new hybrid cell

with two sets of chromosomes.

(4) Somaclonal variation, which is

genetic variation among individual

propagules regenerated from cell

and tissue culture of the clone. It

provides a new source of genetic

variation for selection.

F. The Seed Production Program

The seed production program may be combined

with the breeding program or may be kept separate. The objective of a seed production program is to produce sufficient quantities of

genetically high-quality seeds to meet seed

needs.

1. Seed Production Areas

a. Seed production areas (SPA’s), or seed

stands, are existing stands of natural or

plantation origin that are selected for

phenotypic superiority and managed for

production of seeds.

b. SPA’s are used on an interim basis until

seed orchards come into production.

c. SPA’s can use genetic improvement from

superior provenances.

d. SPA’s can provide seeds for minor species having small planting programs.

e. Practices to improve genetic quality of

seeds include:

(1) Removing phenotypically undesirable trees

(2) Establishing a pollen dilution zone

around the SPA

f. Practices to increase seed production

include:

(1) Thinning the stand

(2) Fertilizing

(3) Establishing access roads (for protection and for seed collection)

2 . Seed orchards-A seed orchard is a collection of selected trees established and

grown together under intensive management for production of genetically improved

seeds.

a. There are two types of orchards: seedling seed orchards and clonal seed

orchards.

(1) Seedling seed orchards involve progeny tests that are rogued (poor

families and individuals removed) so

that the remaining trees can crosspollinate and produce seeds.

(2) Clonal seed orchards are collections

of vegetative propagules (usually

grafts) of select trees. The propagules are establised together, progeny tested when they flower, and

then rogued based on progeny test

results.

b. The genetic quality of seeds is increased

by:

(1) Reducing inbreeding through nonrandom assignment of clones or

familes to the orchard.

(2) Establishing a pollen dilution zone

around the orchard.

(3) Separating provenances into different orchards.

c. Seed production from orchards is increased by:

(1) Establishing the orchard where soil

and climatic conditions are most

favorable for seed production.

(2) Spacing wide enough for maintenance of a full crown but not so wide

that cross-pollination is reduced.

(3) Applying fertilizers to increase the

number of flowers and the portion

of the crown bearing flowers.

(4) Irrigating all year during dry

periods to stimulate flower production. (Irrigation has not provided

consistent results.)

(5) Subsoiling to improve health and

wind firmness of orchard trees and

to stimulate flower production.

(6) Protecting flowers, fruits, cones,

and seeds from insects by periodically applying insecticides.

(7) Protecting flowers from late spring

freezes through cold water irrigation.

(8) Ensuring supplemental mass pollination.

26
G. Sources

For additional information, see Burley and

Styles 1976, Khosla 1982, Nienstadt and

Snyder 1974, Rudolf and others 1974, Wright

1976, Zobel and Talbert 1984, Zobel and others

1987.

II. Production

A. Introduction

Most tree-planting programs are begun by collecting seeds from in-country sources, both natural stands and plantations. To plan these

collections effectively, seed managers should

understand the factors that affect tree seed

crops and generally know what seed yields may

be expected. With this basic information, opportunities may arise to stimulate seed production

in key areas, such as seed orchards or managed

seed stands.

B. Objectives

1 . Recognize the problem of periodicity of seed

production in trees.

2. Learn how environmental factors affect

seed production.

3. Learn how seed production can be stimulated in trees.

C. Key Points

The following points are essential to understanding seed production:

1 . Many tree species bear good crops in cycles.

2. Production is less frequent in high latitudes

and high altitudes and among heavy predator populations.

3. Environmental factors influence flower production, pollination, and seed maturation.

4. Several options are available to stimulate

seed production.

5. Except for seed orchards of a few species,

production data are extremely variable.

D. Periodicity of Seed Crops

1. Temperate secies

a. Many conifers bear in cycles, producing

good crops every 3 to 4 years.

b. Some trees, mainly angiosperms, produce good seed crops every year (e.g.,

Acer, Betula, and Fraxinus).

c. As latitude or altitude increases, the

interval between good crops and the frequency of crop failure increase.

2. ‘Ilropical species

a. Periodicity may depend on wet/dry

cycles. In Nigeria, good seed years occur

after very dry weather in August.

b. Some species (e.g., Tectona grandis)

usually flower each year, with bumper

crops every 3 to 4 years. Other species

(e.g., Pinus kesiya, Cassia siamea,

Cupressus lusitanica, and Delonix

regia) produce good crops most years.

Bawa and Webb (1984) found 93 percent

flower abortion in seven Costa Rican

tropical hardwoods within 2 days, but

the reasons for the abortion are largely

unknown.

c. Dipterocarps in Malaysia bear irregular

heavy seed crops at l- to 6-year intervals.

d. Some Eucalyptus species have large

crops more regularly when grown in

plantations perhaps because of the concentration of pollen in a pure stand. In

the United States, the same finding may

be true for many genera, especially

Liriodendron and P&anus.

3. Genetics- Fecundity is an inherited trait.

4. Documentation-There have been many

observations but few detailed physiological

studies on periodicity of seed crops.

E. Effects of Environment During Flowering

1. Temperature

a. During hot summers, trees usually produce many floral buds and thus large

crops of seeds the next year, but the

reason is not known. It may be related

to partitioning of carbon within the

tree.

b. Late freezes can destroy flowers; sprinkler irrigation can provide some protection.

c. The combination of hot summers and

late freezes suggests that orchards

should be moved to warmer climates

(also to escape insects). However, Pinus

taeda orchards were once moved to

south Texas, but without success.

.2. Light -The effect of light has not been

studied extensively. In the Northern Temperate Zone, southern and western sides of

crowns have the largest flower and fruit

crops. The reason could be light, temperature, or pollen supply. A study of Acerpennsylvanicum found an increase in female-tomale flower ratio as crowns closed in the

stand. The reason could be environment or

internal physiology.

3. Photoperiod does not appear to have a

direct effect on trees.

4. Moisture

a. Drought, coupled with high temperatures, seems to stimulate flower production the following year.

b. Excessive rain during pollination leads

27
to low seed yields in wind-pollinated species.

5. Mineral nutrients - Some general observations and weak study results suggest that

trees on fertile sites flower and produce

more fruit than trees on infertile sites.

Nitrogen and phosphorous are most important for flowering, but the reasons are still

unknown.

6. Biotic agents - Insects, birds, mammals,

and micro-organisms can destroy flowers,

especially in the following tropical species:

a. Triplochiton scleroxylon; attacked by

Apion ghanaense (weevil)

b. Tectona grandis; attacked by Pagyda

salvaris larvae

c. Pinus mrkusii; attacked by Dioryctria

spp. (cone worms)

F. Pollination Agents

1. Wind pollination occurs among all conifers and most Temperate Zone hardwoods

of commercial value.

a. Wind pollination requires:

(1) Lots of pollen

(2) Pollen shed coinciding with receptivity

(3) Relatively close spacing of plants

(4) Good weather-low rainfall, low

humidity, and good winds

b. Supplemental mass pollination (SMP)

has been used in United States southern

pine orchards.

c. Contamination in orchards is a concern.

The degree of contamination can be

determined by isoenzyme analyses.

2. Animal pollination

a. Insects pollinate many temperate hardwoods. (e.g., Liriodendron and Prunus).

b. Bats and birds pollinate many tropical

hardwoods, such as honeyeaters in

Acacia.

c. Animal pollination is usually common in

tropical forests with:

(1) High species diversity and wide

spacing

(2) Abundant foliage to filter out pollen

(3) High humidity and frequent rainfall

(4) Absence of strong stimuli to coordinate flowering (day-length and temperature changes)

(5) Abundant animal vectors

G. Stimulation of Flowering

Flowering can be stimulated in seed production

areas and seed orchards by fertilizing, girdling

and other wounding, thinning, growth regulator treatment, and supplemental mass pollination.

1. Fertilizing

a. Use primarily nitrogen and phosphorous, but frequently potassium.

Spring and summer application in pines

of the Southern United States usually

increases flowering and cone production. Zobel and Talbert (1984) recommend the following annual fertilizer

levels for loblolly pine orchards: 400

kg/ha nitrogen, 80 kg/ha potassium, 40

kg/ha phosphorous, and 50 kg/ha magnesium.

b. Irrigation at the same time as fertilizing

may also help; however, success has not

been universal. Many orchards use dripirrigation techniques.

c. Hardwoods may react favorably; e.g.,

Acer, Fagus, and Juglans. In Hungary,

200 kg/ha nitrogen and 240 kg/ha phosphoric acid more than doubled nut production and tripled the number of sound

seeds.

2. Girdling and other wounding

a . Girdling and other wounding are used to

produce the so-called “stress crops.”

b. Girdling is supposed to inhibit downward translocation of carbohydrates,

thus improving the carbon/nitrogen

ratio. The optimum time to girdle

Douglas-fir is 1 month before vegetative

bud burst.

c. Girdling has increased production somewhat in hardwoods; e.g., Castanea and

Fraxinus.

3. Thinning-In pine orchards, the benefits

of thinning are apparent 3 to 4 years after

thinning. The goal is full crown development but not so much open space as to

reduce cross-pollination.

4. Growth regulator treatment -Great

strides have been made with gibberellins

(GA) applied to conifers by Ross and Pharis

(1976) in Canada.

a. Water-based foliar spray is best.

b. A GA 4/7 mixture is most effective.

c. Both pollen and seed cones are induced.

d. Apply at bud determination.

e. Mode of action is still not known.

f Treatments are most successful when

applied with girdling, root pruning, or

moisture stress.

5. Supplemental mass pollination (SMP)

has been used in pine orchards in the

Southern United States.

H. Postfertilization Problems

1 . Insect damage to cones-A major cause

of losses in conifers.

2 8
2. Drought-Extremely dry weather during

seed development and maturation may

cause cone drop (e.g., Pinus monticola) and

losses in seed weight. Russian data show

lower seed weight in Robinia; the same

reaction occurs with Pinus and Quercus.

3. Cone drop-Postfertilization cone drop in

Pinus is not common except for insect loss.

4. High winds- Late summer storms can

cause great losses of large cones (e.g., r!

palustris and P. elliottii in the Southern

United States).

I. Production Data

Production data include the following published

yield figures:

1. Pinus seed orchard production- In 15

to 20-year-old orchards in the Southern

United States, Pinus produced these crops:

I? taeda, 98 kg/ha; I? elliottii, 86 kg/ha; and

P. strobus, 24 kg/ha. These values should

increase as the orchards get older. If 1 kg of

seeds produces 17,000 plantable seedlings,

1 ha of orchards yielding 50 kg should produce enough seeds annually to plant 600 ha.

2. Pinus elliottii in Brazil produced up to

94 kg/ha of seeds with 500 trees per hectre

in a natural stand, not an orchard.

3. Hardwoods

a. Cecropia obtusifolia, a Mexican

dioecious pioneer, produced 2 to 3 kg/ha

of seeds in a mature forest. The average

was 900,000 plus seeds per tree per

fruiting (Estrada and others 1984).

b. Quercus spp., in the North Carolina

mountains, produced 16,500 to 236,500

seeds per hectre and 3,700 kg/ha (5

species).

c. Liquidambar styraciflua, in the Mississippi River floodplain, had twice the

seeds per fruit head as did trees in

Coastal Plain sites; the former were

almost twice as “fruitful” also.

d. Acacia albida, in Sudan, produced 0.5

million seeds from mature trees (Doran

and others 1983); and in South Africa,

large trees produced several million

seeds. In the Sahel, 125 to 135 kg of

pods per tree, 400 to 600 kg of pods/

hectre and of: 20 seeds per pod were produced, but 95 percent may have been

lost to insects.

e. Tectona grandis, in Thailand, produced

1 kg per tree at age 10 with 8- by 8-m

spacing (1,000 seeds per kilogram).

4. Trade-off-In Temperate Zone species,

radial growth could be decreased 30 to 40

percent in good seed years because of carbon allocation. This means that the current

year’s carbohydrates are used in cone

growth. For Picea abies in Poland, growth

was less in good seed years. The researchers

concluded that selection for fast growth is

also a positive selection for fecundity. It

may be more important in other species;

foresters must decide whether to select for

growth or seeds. The agroforestry aspect

must also be considered.

J. Sources

For additional information, see Franklin 1982;

Owens and Blake 1985; Rudolf and others 1974;

Sedgley and Griffin 1989; Whitehead 1983;

Willan 1985, chap. 3; Zobel and Talbert 1984.

III. Collection Operations

A. Introduction

Successful collection of tree seeds is usually the

result of detailed early planning. Ample time

must be allowed to plan an efficient and practical collection strategy and to assemble the

resources necessary for its implementation. Key

elements include a good estimate of crop size,

proper equipment, and a well-trained crew.

Comprehensive collections for research certainly require more detailed planning than mutine bulk collections and may require a lead

time of 1 to several years depending on the

circumstances.

B. Objectives

1. Identify simple techniques for seed crop

estimation.

2. Determine the factors that should be considered when collections are planned.

3. Understand the importance of documentation.

C. Key Points

The following points are essential for planning

collection operations:

1. The best seed sources available must be

selected.

2. Good planning requires advance estimates

of the seed crop and, at a later date, estimates of seed yield per fruit.

3. Large collection planning must include

choice of personnel, training, transportation, collection equipment, safety of

workers, labeling of seedlots, description of

sites and stands, etc.

D. Seed Source

Seed source is extremely vital for all seed supplies; this point must never be underestimated.

(See “Genetics and Seed Source.“) Terms used

in genetics are:

29
1 . Origin-the natural stand location of the

origina.l mother tree.

2. Provenance- the place where mother

trees that produced the seeds are growing.

Seed source and provenance are the same.

(Origin and provenance are usually

reversed in the United States.)

3. Laud race-exotics that adapt to develop

improved sources.

4. Seed zone maps -necessary for a good

program to eventually develop seed zone

maps (figs. 14 through 16) [no equivalent

figures in Student Outline].

E. Estimating Seed Crops

If seeds are in short supply, all possible seeds

should be collected, stretching crews and equipment. If the location of large crops is known,

more seeds can be collected for a given amount

of cost and effort. Seed crops can be estimated

by any of the following five methods:

1 . Flower counts- only feasible with large,

showy flowers. Pinus spp. strobili are about

the minimum size.

2. Immature fruit and seed countsuseful in the collection zone before seed

maturity because many flowers may abort.

3. Fruit counts on standing trees- work

best when seeds are nearly mature.

a. Total counts are feasible only for light

seed crops.

b. Crown sampling is most common and

uses portions of the crown (10 or 25

percent). [Do the exercise in exercise 2 if

there is time.] A 5-percent sample of

orchard trees in midsummer gave a good

Figure 14.-Recommended seed collection and planting zones for

Pinus palustris in the United States (Lank and Kraus

1987) [no equivalent figure in Student Outline].

estimate in I? taeda in Virginia. Some

people use a two-person system (one on

each side of the tree).

4. Rating systems-Examples of rating systems are the Tanzania rating system (table

4) [no equivalent table in Student Outline]

and a North American rating system for

conifers (table 5) [no equivalent table in

Student Outline].

5. Cross-section seed counts-Cones are

cut lengthwise (or perpendicular to the axis

for round fruits), and exposed filled seeds

are counted (table 6) [table 3 in Student

Outline]. This number can be related to

total good seeds per fruit. Enough seeds

must be counted to calculate a simple

regression. [Do the exercise in exercise 2 if

fruits are available.]

F. Planning Considerations

The steps of planning a collection are:

1. Define the objectives

a. For effective planning, the coordinator

of collecting operations needs a clear

statement of objectives. Prior knowledge

of species and provenance priorities,

sampling strategy, the standard of documentation, and amount of seeds required is important to assemble the

resources needed for the type of collection proposed.

b. Flexibility should be built into the planning to allow collectors to make decisions about unforeseen circumstances.

For example, it would be highly wasteful

for collectors to return empty handed

from remote areas because of restricted

or narrowly defined aims and procedures when an alternative and potentially valuable seed harvest was available for collection.

Table 4.-Tanzania rating system for seed crops (adapted from

Willan 1985) [no equivalent table in Student Outline]

Crop rating

Numerical Wording Criteria

0 None Trees without flowers and fruits

1 Weak Flowering and medium-sized seed

crops on free-growing trees and on

trees on free borders of stands

2 Medium Flowering and very good seed crops on

free-growing trees and on free borders

of stands; trees within stands bearing

seed crops at top of crowns

3 Very good Flowering and very good seed crops on

most trees

30
n

3

3 2

/ L$-.-8 - -.- - -._, - - -._
phitsonulok I

Phi& Phetch%un

Zone II

J

,60

\

Nokhonsowon

\

419 P : T Ratio

Chiengroi 7 3 (r795.0 / 24.6,

Moi Hongsarn 49 (1253.7 / 25.4,

Chiengmai 5 1 (1270.9/25.01

Nan 50 (1298.0 /26.0)

Lampong 43 (rlO%O /26.0)

Phroe 41 (1114.8 /270)

Moe Sarieng 47 (1223.4 /26.2)

Uttoradit 5 2 (1452.2/277)

Phitsonulok 50 (1358.0 1222)

Tok 38 (1054.0 /225)

Moe Sod 58 (1525.0 126.5)

Nokhonsowon 41 (1173.7 /28.5)

Ton

I

J

4.5

99O TOO0 E

Thailand teak seed collection zones :

Zone I = dry-humid zone Zone III ‘= moist zone

Zone II = medium-humid zone Zone IV = wet zone

J

Figure 16. -A proposed seed zone map for teak (Tectona grandis) in Thailand based on the annual precipitation/mean

annual temperature ratio moisture index value for 1951-1975 (Kaosa-ard 1983) [no equivalent figure in

Student Outline].

3 2
Table 5. -North American rating system for conifer seed crops

(adapted from Willan 1985) [no equivalent table in Student Outline]

Crop rating*

Numerical Condition Criteria

1 Failure No cones to a few scattered on a few

trees

2 Very light Some cones on some trees

3 Light Good to fair cone crop on 50 percent

of exposed crown of 50 percent of

trees

4 Medium Good to medium cone crop on 75

percent of exposed crown on most

trees

5 Heavy Good cone crop on all exposed crowns

of most trees.

‘A numerical rating of 4 or 5 is a good prospect for all collectors. A

rating of 3 has possibilities for more experienced collectors. A rating

of 1 or 2 is a poor prospect for all collectors.

Table 6.-Sound seed yield per cone for four Pinus species as estimated from the number of sound

seeds exposed when cones are bisected longitudinally (Derr and Mann 1971) [Table 3 in

Student Outline]

Sound

seeds

exposed

2

4

6

8

10

12

14

I? elliottii

I? palustris P taeda rl elliottii (Georgia- I? echinata

(Louisiana) (Louisiana) (Louisiana) Florida) (Virginia)

_____-________-_____------------------ Sound seeds per cow _---__---------_-__--------------------

2 3 31 2 0 31 12

3 5 4 4 35 50 22

4 7 57 50 69 31

5 9 70 65 a7 41

71 a 3 80 106 51

83 9 6 95 124 6 0

95 109 110 143 70

2 . Gather background data

a. A literature search may provide information on the natural distribution of a

species -its habitat, ecology, and genetic

variability - and on fruiting and flowering times in different parts of its range.

A visit to regional herbaria to examine

relevant herbarium specimens may add

significantly to the published information on species distribution, variability,

and reproductive phenology. Personal

contact with botanists, field foresters,

rural people, and others who study or

use a species can also help.

b. If the collections are wide ranging and

cross political boundaries, early official

contacts with the forest services in the

States or countries concerned are essential to develop a good working relationship of mutual benefit to all parties.

c. All available information should be collected and summarized. Data on natural

occurrence are best plotted on maps

detailed enough to show the main transportation systems (e.g., roads, rivers,

raiIways, and airstrips), topography, and

other information that may assist in

selecting collection sites. Seed-zone overlays for these maps are very useful.

d. Field reconnaissance is essential, preferably including adequate seed crop estimates and samples of fruits and seeds.

e. The coordinator of collecting operations

must determine the number of person33
3.

nel needed for the collecting team and

must verify that they have the skills to

meet the stated objectives. Key personnel should be selected early so that they

can familiarize themselves with the project and species and, if necessary, be

trained in the techniques they will be

using in the field. Collection team

leaders are responsible for the discipline,

morale, and safety of personnel and the

success of the operation.

Collect field data

The field data provide the information for

relocating the site in future years for further work and provide the background

essential for interpreting the results of

the experiments. The data to be gathered

at each collection site must be specified,

and systems must be developed to make

their collection as reliable and easy as

possible for the worker in the field. Specially prepared field data sheets are recommended to ensure that uniform records

are obtained from all sites. The standardized “Seed Collection Report” sheet

adopted for the Fuelwood Project of the

FAO and the International Board of Plant

Genetic Resources and the “Seed Data

Sheet” of CSIRO are good examples (figs.

17, 18) [no equivalent figures in Student

Outline]. Significant features of these

documents are:

a. Locality as indicated by latitude and

altitude is essential to define the provenance area. They should be stated accurately and concisely so that future

collectors can return with certainty to

the same collection site. Distances from

the nearest town, village, or geographical feature (river or mountain), the forest district, and any other facts that will

assist future field parties should be

recorded. Maps (hand drawn if others

are not available) and aerial photographs showing the stands and the position of seed trees are useful and should

be filed with the data sheets for ease of

reference.

b. Aspect, slope, climate, soils, and associated species help to build a picture of the

environment and ecology in which the

trees grow and may help in interpreting

experimental results. The collector

should encourage relevant comments

from local people on the history of the

area and its climate. Recording tolerance

to such features as alkalinity or salinity

4.

in the soil, seasonal inundation, etc. may

be important later.

c. Individual tree descriptions (e.g.,

height, diameter, stem form, and

branching habit) and the number of

trees in a provenance are of obvious

importance to any future tree-breeding

work. Photographs are a useful adjunct

to written descriptions.

d. Collecting herbarium specimens of little

known or variable species allows for

later scrutiny by botanical experts. Care

is needed in handling the specimens,

and it is best, after drying, to dispatch

them to the base at the earliest opportunity to prevent damage.

e. Other data and notes that may be of

great value to future collectors are crop

and seed details, collection methods,

etc.

f It is paramount in provenance work to

adopt foolproof systems for maintaining

the identity and purity of each collection. Systems must prevent any seedlot

from being contaminated with the seeds

of another lot through all steps, from

collection to registration in the seed

store. Careful labeling through each process is essential and can be facilitated by

using preprinted durable labels. A wellproven method of numbering collections

used by CSIRO is based on party

leaders’ assigning numbers to the collections in numerical sequence, prefixed by

their initials. The number assigned to a

particular tree appears on and in the

collection sacks and seed bags, on the

herbarium voucher specimen, and on

the photograph, and it identifies any

other sample or recording of that tree.

The provenance number is not assigned

until the seeds are ready for storage.

Plan the itinerary:

a. Reaching the collection region well in

advance of the proposed date for beginning the collection is important. In

developing an itinerary for the collection

team, team leaders must be aware of

this point.

b. Organizing the sequence of operations

in a particular region may require 2 to 4

weeks. Extra permits may have to be

obtained, labor recruited and trained,

and reliable transportation arranged.

c. Make the schedule flexible. The itinerary is important as a guide, but it must

be flexible to account for the unexpected

3 4
FAO PROJECT ON GEhETIC REI;(XlRCES OF ARID/SWI-ARID ZONE ARBOREAL SPECIES:

SEED COLLECTION RIJ'ORT

Collection No. . . . . . . . . . . . . . . . . . . .

Species: . . . . . . . . . . . . . . . . . . . . . . . . ..*...*.......

Country: . . . . . . . . . . . . . . . . ..r.................*.

Province: . . . . . . . . . . . . . . . . . . . . District: . . . . . . . . . . ë . . . . . . l

tat: 0 I a:

0

. . . . . . . . . . , . . . . . . . . . . EleV: . . . . . . . . . m

T~papptphy: Blat/hilly

Slope: steep/medium/gentle

Soil: Deep/shallow/intermediate

Drainage: . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . ..-....................................

Stoniness: . . . . . . . . . . . ..*........*..................

Texture : .r.........................................

. . . . . . . . . . . . . . . . ..r.......*.........*......

pH: Acid/neutral/alkaline

Rainfall: Xean annual . . . . . . . . mm; Wet months: . . . . . . . . . . . . . .

Temperature: Hean annual: . . . . . OC; Xean ma: .*... OC;

Frost: . . . . . . . . . ..days/year

Stand: Natural: Gmups/open Thin/dense

Dry months: . . . . . . . . . . .

Mean min: . . . ..Oc

Young/middle-aged/old

Plantation: Age . ..-..-.. y-s Height . . . . . . . . m Dism . . . . . . . . .cm

OsQinal 6ource: . . . ...~.............................................

Associated species: . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Form: *lea: Single/multiple Straight/fair/poor

cn3Me : li'lat/narrow/average/uide

Seed Crop: Light/medium/heavy

Seed collection: No, of trees . . . . . . . . . Xi& distance apa.r-ta ..r.r....o~; Egg . . . . . . . . . .

Remarks8 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ..*...*......... ,.....

. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ..*...................................

..........................................................................................

Date of collectiona ....................

.................................................

Officer in Charge

Figure 17. -Seed collection report form used in the FAO Project on Genetic Resources of AridlSemi-arid Species (Willan 1988)

[no equivalent figure in Student Outline].

35
CSIRO Division of Forest Research

PO Box 4008

Canberra ACT 2600

SEED DATA SHEET

Species:

Collection hpcality:

Seedlot No.

‘Latitude: O 'S Longitude: ' 'E Altitude:(m) - -

Aspect Slope - -

Climatic zone: Met. Station:

Association includes:

Geology and Soil: pH:

Collection Dot. Photo. tit dbh

no. spec. no. (4 (cd

Tree description Seed No. of

wt viable seec

(9) /IO 9

Work supervised by: Date:

Figure 18. -Seed data sheet used by CSIRO CWillan 1985) [no equivalent figure in Student Outline].
problems that invariably arise in the

field. Overly restrictive schedules may

lead to loss of accuracy and attention to

detail, and to lower worker morale.

5. Organize equipment permits and

transportation

a. Team leaders must specify at an early

stage what equipment is to be used if

long delays in purchase or delivery are

anticipated.

b. Identify applicable government regulations. Most countries have regulations

governing collection, export, introduction, and, perhaps, movement of seeds.

Official permits may be required for any

of these procedures, and, in some cases,

separate permits are needed for each

collection site and for each individual

seedlot exported or introduced. Customs

officials and plant. health authorities

may seriously delay the operation or

even destroy seeds if the full entry procedures are not followed. For some

seeds, extra delays of even a few days in

transmission can be fatal.

c. Us& care between the collection of the

seeds and their arrival in the seed laboratories. This period is critical to their

viability and vigor. Transportation must

be where it is wanted when it is needed;

thus, prior organization is essential to

minimize transit time.

G. Collection Equipment -A Comprehensive List

The following items are necessary for collection

operations:

1. Administrative items:

a. Movement approvals

b. Collection authorities

c. Radio transmission permits

d. Drivers’ licenses

e. Firearm permits

f. Facilities for purchasing stores; e.g.,

gasoline (petrol) and oil

2. Literature:

a. Road, topographic, and soil maps to

cover the collection route itinerary

b. Literature on the genera and species to

be collected

3. Collection equipment:

a. Notebooks, recording forms, pens, and

pencils

b. Binoculars

c. Markers; e.g., colored plastic ribbon

d. Camera and accessories

e. Tree-measuring instruments; e.g., diameter tape, height-measuring instrument, and length tape

f. Soil sampler, pH testing kit, and soil

charts

g. Compass

h. Altimeter

i. Hand lens

j. Large collecting sheets; e.g., 4 by 4 m of

heavy duty plastic or canvas

k. Small collecting sheets; e.g., 2 by 2 m of

calico or other finely woven cloth

1. Seed bags made of finely woven cloth of

various sizes; e.g., 100 by 100 cm to 10

by 20 cm for small seed samples, all with

ties

m. Large grain bags for dispatching seeds

n. Cutting equipment; e.g., secateurs,

long-handle pruning saws, shears, ladders, chain saws with fuel and accessories, bowsaws, flexible saws, throwing

ropes, axes, rifles with ammunition,

and rakes

o. Safety gear; e.g., steel-capped boots,

leather gloves, safety helmets, and

safety belts

p. Weatherproof tags for labeling each

seedlot, to prevent the markings from

becoming illegible when wet or abraded

q. Tags for botanical specimens; e.g., white

cardboard “jewelers” tags

r. Plant presses for botanical specimens

s. Papers to dry specimens in the plant

presses

t. Plastic bags

u. Specimen bottles with preservative fluid

v. Containers for soil samples

w. String

H. Sources

For additional information, see Barner and

Olesen 1984; Bramlett and others 1977; Doran

and others 1983; Ontario Ministry of Natural

Resources 1983; Willan 1985, chaps. 3, 4, 5 +

appendices 1,5,6.

Maturity

A. Introduction

Choosing good stands and trees for seed collection means nothing if fruit or seed maturity

cannot be easily identified on the trees by

unskilled workers. If seeds are disseminated

immediately at maturity, workers must know

how much in advance of maturity seeds can be

collected without collecting seeds that will not

germinate. If predators inflict large losses on

mature seed crops, a similar problem exists.

Good maturity indices are often the keys to

successful collection.

37
B. Objectives

1. Learn the common indices of maturity

employed in tree seed collections.

2. Recognize how these techniques can be

adapted for new species.

C. Key Points

The following points are essential to recognizing seed maturity:

1. Seed moisture content is very important,

but direct measurement in the field is

impractical; indirect estimates may be substituted.

2. Color changes are the most common

indices.

3. Chemical indices are possible but impractiCd.

4. Artificial maturation of immature seeds is

an option for some species.

D. Successful Collection

The following points are essential to successful

collection:

1. Biological ideal-to collect seeds at the

peak of their physiological maturity; however, this ideal is not possible for large-scale

collections. First, physiological maturity,

maximum food reserves, minimum moisture content, and ideal regulator balance

must be defined. For example, ideal regulator balance is expected at abscission/

dispersal, but it may not occur then.

2 . Practical collection - In most collection

operations, fruits and seeds are:

a. Collected from the ground after dissemination (good for large seeds only).

b. Collected from logging operations.

c. Collected from standing trees as close to

full maturity as possible before dissemination.

d. Collected from standing trees well in

advance of maturity and ripen artificially.

e. Both c and d require climbing and/or the

use of special equipment.

E. Collection after Dissemination

Seeds that can be collected after dissemination

are primarily large, single-seeded fruits; e.g.,

those of Quercus, Carya, Juglans, and Aesculus

(temperate), and dipterocarps, and other large

seeds (tropical). However, the first seeds to fall

are usually bad because of insect damage or an

immature embryo. Workers must quickly collect

the seeds before animals destroy them,

especially in the Tropics.

F. Other Collection Strategies

Other collection strategies involve collection of

seeds before dissemination. These collections

would usually be from standing trees, although

collection from logging slash would fall into this

category also. These strategies depend greatly

on the ability to judge seed maturity.

G. Maturity Indices

Maturity indices include physical and chemical

characteristics. They are needed for all collection strategies.

1. Physical characteristics

a. Color changes from green to yellow to

brown or black (e.g., Pterocarpus); from

green to red to purple or black (e.g.,

Prunus); from green to yellow; from

green to yellow to purple (e.g., Gleditsia

triacanthos); and from green to brown

(e.g., conifers)

b. Moisture content

(1) There are three moisture trends

during ripening:

(a) In dry, orthodox seeds and

fruits, moisture decreases

slowly as seeds mature (fig. 19)

[no equivalent figure in Student

Outline].

(b) In pulpy, orthodox fruits, moisture decreases at first, then

increases mainly in the pulp

(fig. 20) [no equivalent figure in

Student Outline].

(c) In recalcitrant seeds, moisture

increases early, then slightly

decreases (fig. 21) [no equivalent figure in Student Outline].

(2) Moisture content is related to protein synthesis (see Rosenberg and

Rinne 119861). Seed moisture must

drop below 60 percent to trigger

protein synthesis. Without this happening, seedling growth is arrested.

This may be true for all orthodox

seeds. Ellis and others (1987) found

a critical level of 45 to 50 percent

moisture for six grain legumes.

(3) Moisture content can be measured

directly by oven methods; that is,

cut cones, large fruits, or seeds;

weigh; dry for 17 hours at 103 “C!;

and then weigh again.

(4) Specific gravity is usually discussed

separately, but it really is just an

estimate of moisture content. Specific gravity has been measured in:

(a) Conifers-See table 7 [table 4 in

Student Outline] and fig. 22

[figure 7 in Student Outline].

(b) Quercus data from Russian

sources found significant

changes in specific gravity in

38
I”------.--/--+-

/-

/’

- FRESH WEIGHT

---- MOISTURE

--- DRY WEfGHT

JUN. JUL. AUG. SEPT. OCT.

6r

5 -.

4-

I -

/

Figure 19. -Seasonal changes in fresh weight, dry weight, and moisture content during maturation of fruits of Platanus

occidentalis (adapted from Bonner 1972a) [no equivalent

figure in Student Outline].

7

- FRESH WEIGHT

6

---- MOISTURE

--- DRY WEIGHT

5

Y

2

4

:

.3

4

2

I

0

_---

-

_---

-4---

-_--

.

/

-2

- C

JUN. JUL. AUG. SE PT. OCT. NOV.

Figure 21.-Seasonal changes in fresh weight, dry weight, and moisture content during maturation of

Quercus alba (adapted from Banner 1976) [no equivalent figure in Student Outline].

- FRESH WEIGHT

---- MOISTURE

--- DRY WEIGHT

\

\

\

\

\

MAY JUNE JULY AUGUST

Figure 20. -Seasonal changes in fresh weight, dry weight, and maisture content during maturation of Prunus serotina

(adapted from Banner 1975) [no equivalent figure in

Student Outline].

39
Table 7. -Cone specific grauity values that indicate seed maturity in

some conifers [!&ble 4 in Student Outline]

Species

Specific

gravity Reference

Abies grandis

Cunninghamia kmceolata

Pinus elliottii

P merkusii

P palustris

I? strobus

I? taeda

I? virginiana

0.90 Pfister 1967

0.95 Jian and Peipei 1988

0.95 Barnett 1976

1.00 Daryono and others 1979

0.90 Barnett 1976

0.90 Bonner 1986a

0.90 Barnett 1976

1.00 Fenton and Sucoff 1965

MOISTURE CONTENT = 102.84 - 44.79 (

r : 0.92

I (Et) Pinus strobus

60.

(A) Pinus foeda

IO- '

I I I I I

0.95 0.85 0.75 0.65 0.55

CONE SPECIFIC GRAVITY

‘0

1

Figure 22. -The relationship of moisture content to specific gravity for cones of Pinus

taeda and P. strobus (Bonner 1991b) [figure 7 in Student Outline].

40
acorns of some Quercus species

as they matured, but results

with North American species

have been inconsistent.

(c) Other angiosperms-there is

usually a flat line over maturation period (little success).

(d) Barnett (1979) developed a simple procedure for determining

specific gravity using water

flotation that is easy to perform

(fig. 23) [figure 8 in Student

Outline].

c. Other physical indices:

(1) Acorn cup release in Quercus is

good; easy release signifies maturity.

(2) If bending Pinus strobus cones

causes scales to “open,” they are

mature.

(3) When the white, brittle embryo in

Fraxinus (and some other genera) is

excised and bent, mature embryos

will break.

(4) A minimum percentage of the

embryo cavity must be filled; 75 percent is a general guideline for Fraxinus excelsior and Pinus sylvestris

in northern Europe.

750 ml 775 ml

600 ml

CONE WEIGHT: (6) 750 - (A)600: 150 (IramS

CONE VOLUME: (Cl 775 - (A) 600 = 175 gro”s

SPEClFlC GRAVITY: (WEIGHT) 150 g i (VOLUME) 1759 = 0.66 ‘2

Figure 23. -A simple technique for determining specific gravity of

pine cones in the field using a graduated cylinder.

(A) Fill the cylinder with water to the 600 mL mark. (B)

Float the cone in the water and record the water level. (C)

Using a pin or needle, submerge the cone enough to

completely cover the cone with water, but no more. Record

the new water level (adapted from Burnett 1979) [figure

8 in Student Outline].

2. Chemical characteristics

Chemical indices are biologically sound but

are not practical, even in sophisticated collection situations.

a. Accumulation of storage foods can indicate full maturity.

(1) Angiosperms of the Southern

United States: Liquidambar styraciflua, 25 percent crude fat; Fraxinus pennsylvanica, 10 percent fat

and 14 percent insoluble carbohydrates; Quercus spp. (red oaks), 20

percent fat and 25 percent insoluble

carbohydrates; Quercus alba (white

oak), 40 percent insoluble carbohydrates (figs. 24 through 27) [no

equivalent figures in Student Outline].

(2) Western conifers: Abies procera, 25

percent crude fat; Pseudotsuga

menziesii, 23 percent crude fat or

1.3 percent reducing sugars.

b. Elemental analyses of calcium, magnesium, and phosphorus have shown some

good relationships in angiosperms of the

Southern United States. Relative

amounts will decrease in acorns and

increase in small seeds; absolute

amounts increase in all seeds as they

mature (fig. 28) [no equivalent figure in

Student Outline].

c. Growth substances. In Poland, auxins

decrease in maturing Quercus robur

according to Michalski (1969).

(1) Predicting Picea abies germination

by level of indoleacetic acid (IAA)

has even been proposed by several

European workers.

(2) Gibberellin changes have been followed, and levels usually increase

with maturation.

H. Artificial Maturation

Artificial maturation is an option for some situations. Immature seeds can be artificially

ripened by storing them in cool, moist conditions. Krugman (1966) (Western United States)

and Lantz (1979) (Southern United States) successfully used artificial maturation with Pinus

spp. Bonner (1972c) (Southern United States)

was successful with Liquidambar styraciflua

and Liriodendron tulipifera but not with species of Fraxinus or Quercus.

1. Single-seed versus multiple-seed

fruits

Single-seed fruits are cut off from all nutrition when picked, but multiple-seed fruits

4 1
CRUDE FAT CONTENT -

GERMINATIVE CAPACITY

0

2

6-24 7- 8 7-22 8-5

b

E-19 S-I S-17 I O - 6 IO-21

COLLECTION DATE

Figure 24. -The relationship of crude fat content and germination in maturing seeds of Liquidambar

styraciflua (Banner 1972a) [no equivalent figure in Student Outline].

2

’ 6C

8

\h

F” 40

2 0

0

----- PHOSPHORUS (PERCENT)

--- CARBOHYDRATE (mg/g)

- CRUDE FAT (mg/g)

---- PROTEIN -N (mg/g)

JUN. JUL. AUG. SEPT. OCT. NOV.

Figure 25. -Seasonal changes in phosphorus, soluble carbohydrates,

protein-nitrogen, and crude fat in samaras of Fraxinus

pennsylvanica (adapted from Bonner 1973) [no equiw

alent figure in Student Outline].

42
---- 0. shumordii

I -P Q. nigra

I -

0

JUN. JUL. AUG. SEPT. OCT.

Figure 26. -Seasonal changes in crude fat content of fruits of Quercus nigra and Q. shumardii

(adapted from Bonner 19743, 3976) [no equivalent figure in Student Outline].

---- INSOLUBLE

SOLUBLE

400 -

/

/’

/

/

cc-

/

/

300 - /

/

/

/’

//

200 - /’

.’

0-N /

.

0

/

0

d

JUN. JUL. AUG. SEPT. OCT. NOV.

Figure 27.-Seasonal changes in insoluble and soluble carbohydrates in fruits of Quercus alba

(Bonner 1976) [no equivalent figure in Student Outline].

4 3
- PERCENT Ca

- ---- PERCENT Mg

JUN. JUL. AUG. SEPT. OCT. NOV.

Figure 28. -Seasonal changes in calcium and magnesium in samaras of Fraxinus pennsylvanica

(adapted from Bonner 1973) [rw equivalent figure in Student Outline].

can obtain some nutrition or growth factors

from cone or fruit tissues to complete maturation.

2. Avoiding dormancy

a. Tilia americana-This species can be

picked before full maturity when the

seedcoat is not completely dried.

b. Acacia spp. -Early collections were

reported by Doran and others (19831.

c. Gleditsia triacanthos -Early collection

also works with this legume; the pod is

picked while it still shows some yellow.

Hard-seeded dormancy is not so strong,

but resistance to disease is low.

3. Usefulness-Artificial maturation is useful when collecting on remote or expensive

sites. However, seed yield and quality usually suffer. In one case, Liquidambar

styra.cifZua yield was 20 percent lower than

with natural maturation; germination was

equal to that of seeds from later collections.

I. Delayed Collections

For certain species, there is no rush to collect

the fruits; they do not disperse their seeds right

away (orthodox seeds only>.

1 . Serotinous cones (Pinus and Picea) -

Resins prevent cones from opening until

they melt in wildfires. In cool climates,

seeds can remain viable for several years in

the cones.

2. Delay in abscission- Platanus spp. are

good examples of a fruit that stays intact on

the tree for several months past maturity

without decreasing seed quality.

J. Sources

For additional information, see Bonner 1972a,

1972c, 1976; Nautiyal and Purohit 1985;

Rediske 1961; Willan 1985, p. 33-38.

V. Postharvest Care

A. Introduction

The time between collection and extraction is

often overlooked as a crucial segment of seed

acquisition. Fruits and seeds, often high in

moisture content, must be stored and/or transported for extraction and cleaning. Special care

must be taken during this period to avoid loss of

seed quality, especially in tropical and subtropical areas where transportation systems do not

allow immediate delivery to extraction centers.

B. Objectives

1. Recognize the crucial times when seed

quality may be lost.

2 . Plan storage and transportation systems to

minimize the danger to seed quality.

C. Key Points

The following points are essential to postharvest care:

4 4
1 . High moisture contents and high temperatures are dangerous for orthodox species.

2. High moisture levels must be maintained in

recalcitrant seeds, but excessive heat is a

problem with these seeds.

3. Fruit storage can be advantageous for some

species because of the afterripening processes that occur in the seeds.

D. Storage Before Extraction

1. Operation schedules-Time does not

permit seeds from all trees or families to be

collected at peak maturity; therefore, some

must be picked and stored. Extractories

could not handle them all at once, anyway,

at least for pines in the United States.

2. Predrying- When artificial heat can be

used, drying during storage can remove

much of the moisture and thus lower drying costs. In pines of the Southern United

States, two-thirds of the moisture removal

required for cone opening can be done in

predrying storage. Moisture should decrease from about 70 to 30 percent.

3. Completion of maturation- Completion

of maturation is an often overlooked

benefit, but it is just as important as

predrying.

a. Abies is a well-known example; 5 or 6

months in cool, moist conditions complete maturation. In natural habitats,

seeds mature as cones disintegrate.

Picea glauca in Canada needs 6 weeks of

storage in the cone to achieve the best

germination.

b. New data show similar benefits to seed

quality from storage before extraction

for some pines of the Southern United

States.

c. Premature collection is suitable for some

multiseed angiosperms; e.g., Liquidambar styraciflua, Liriodendron tulipifera,

and P&anus occidentalis. This is done

to lengthen collection season or, in

northern climates (Scandinavia and

Russia), to complete ripening in conifers

in short growing seasons. It is usually

not recommended in the United States

because yield per cone and seed quality

generally suffer.

E. Southern Pines

1. Storage is usually related to operation

schedules.

2. Outdoor storage is better than indoor

storage because:

a. Drying is faster outdoors.

b. Cones open better when wet/dry/wet/dry

cycles occur.

3. Containers

a . Containers must provide air circulation.

b. Loose-weave burlap bags (about onethird hectoliter) and 7-hectoliter,

wooden crates are effective.

(1) Burlap bags of Pinus taeda cones

left in the seed orchard in the shade

did well in one study, providing an

extra 5 weeks of collection time.

Shaded concentration points were

no better than sunny ones.

(2) Temperature profiles of the 7-hectoliter crate show no overheating in

the middle.

c. Plastic bags or sacks should not be used.

d. Paper sacks are effective for small lots if

their tops are left open.

4. Time

a. Cone storage can improve germination

rate.

b. Maximum length of storage depends on

the species:

(1) 4 weeks for Pinus elliottii

(2) 5 to 7 weeks for l? taeda

(3) 3 weeks for I? palustris

(4) 5 to 7 days for I? strobus in the

Southern United States

(5) 7 to 9 weeks for P strobus in

Ontario

5. Other factors

a. Original maturity-More mature cones

may have shorter optimum storage

periods (very important), although this

cannot be quantified as yet.

b. Local conditions (weather, equipment,

etc.) are also important; warm, rainy

conditions increase the risk of cone

molds.

6. Immaturity/Dormancy

a. Seed ripening in cones increases germination rate and thus decreases dormancy.

b. Past a certain point, cone storage will

decrease germination - first, germination rate, then total germination. This

point cannot be accurately defined as

yet.

7. Heat and Molds

a. Green cones can generate enough heat

to char cones black.

b. External molds are common in 7-hectoliter crates, but studies have found no

evidence of damage to seed quality.

Pinus echinata data from Pollock, Louisiana, and Abies alba from Yugoslavia

demonstrated better germination in

seeds from moldy cones.

45
c. Good aeration is essential to prevent

mold growth on cones during drying.

F. Serotinous Cones

1. Storage is not a major problem for Pinus

glauca, P contorta, or I? patula.

2. Maturity- Some pine seeds need to stay in

the cone because 3 years may be needed to

reach maturity (Pinus torreyana in the

Western United States).

3. Viability-Seeds can retain viability for

several years in cones, but seed quality may

be reduced. However, Canadian workers

found that seeds from lo-year-old 19 contorta cones in Canada can be very good.

G. Other Conifers

1 . Abzks (true firs) -Seeds of true firs must

complete ripening in the cones. They should

be stored in burlap bags in sheds with

plenty of aeration around the bags for

weeks to 6 months, depending on the species.

2. Piceu (spruce) -Seeds of most spruce species should be extracted as early after collections as possible. Winston and Haddon

(1981) in Canada found that 4 weeks of

cone storage at 5 “C! of I? glauca were

enough, but Wang1 reported that 6 weeks

were needed. These seeds had excellent germination and no dormancy; no stratification was needed. Immature P rubens cones

can be stored for several weeks, then the

seeds are “teased” out with wet/dry cycles.

3. Pseudotsuga - Extended storage of 3 to 4

months is possible in dry, well-ventilated

conditions. Too much moisture leads to

pathogen problems. Some artificial ripening

is possible.

4. Tropical pines

a. Treatment of tropical pines is similar to

that for Southern United States pines,

but the tropical environment is more

stressful. Cones should be stored under

a roof or cover with good ventilation,

and temperature should be kept

between 20 and 35 “C.

b. Rodents and fungi can be big problems,

so special efforts should be made to protect the seeds.

c. In Honduras, Pinus caribaea is precured

until all of the cone changes from green

to brown.

‘Wang, B.S.P. 1985. Personal communication with the author. On

file with: U.S. Department of Agriculture, Forest Service, Southern

Forest Experiment Station, Starkville, MS 39759.

d. In New Zealand, immature cones of P

radiata are stored for 10 weeks at 20 to

24 “C to complete ripening and to fit the

planting schedule (Willan 1985).

e. In Indonesia, green and green/brown

cones of l? merkusii had improved opening, yield, and seed quality when stored

for 2 to 4 weeks before extraction (Arisman and Powell 1986).

H. Hardwoods

1. Artificial ripening-Many hardwoods of

the Southern United States respond to

artificial ripening of immature fruits;

Liguidambar styraciflua can be collected 4

weeks early; Liriodendron tulipifera can be

collected 4 to 6 weeks early. However, seed

yields and quality will suffer. This effort

has not been as successful in pines.

2. Regular storage -The above species

should be stored in loose-weave bags and for

as short a period as possible. They should

be spread one or two fruits deep for drying

and stirred two or three times daily to avoid

overheating. Other orthodox species

include:

a. Eucalyptus -Tight-weave cloth or plastic can be used, but overheating is a danger with plastic; use with caution.

b. Legumes -Storage is not difficult, but

overheating is possible if moisture is

high; they should be spread to dry.

c. Drupes -Short storage to complete

ripening is possible for some species, but

they should be spread in a single layer

and shaded, preferably indoors. As soon

as the color changes, they should be

cleaned. For Prunus species drupes

should be cleaned within 3 days of collection.

I. Summary

Seeds of most species fit into one of three

groups:

1. Harvest dry, keep dry-Usually start

fruits drying immediately and keep dry

after extraction (e.g., Pinus, Liquidambar,

Liriodendron, Acacia, and Eucalyptus).

a. Dry slowly.

b. Provide good aeration; do not use plastic

bags.

c. Use suitable containers, including:

(1) Burlap bags

(2) Racks

(3) Wooden crates

(41 Canvas or plastic sheets

2. Harvest moist, then dry-Keep moist

when collecting and during extraction to

avoid formation of tough, impermeable

46
covers (drupes); extract, then dry seeds for

storage (e.g., Nyssa and Prunus).

a . Spread to avoid heat and (in most cases)

fermentation.

b. Use trays or bags; do not use plastic.

c. Avoid outer coat toughness.

d. Extract, wash, and dry for storage.

3. Moist forever-Recalcitrant seeds are

kept moist forever because drying will

decrease quality; seeds are not stored when

collected, but are extracted immediately

e.g., Quercus, Aesculus, Shorea, and Hopea.

a. Never dry.

b. Keep moisture 230 percent.

c. Refrigerate to a safe temperature:

(1) 1 to 3 “C for temperate recalcitrants

(e.g., Quercus).

(2) 15 to 20 “C for tropical recalcitrants.

d. Use polyethylene-lined containers or

polyethylene bags 4 to 10 mil thick.

J. Sources

For additional information, see Bonner 1987a;

Willan 1985, p. 78-86.

47

49
I. Drying and Extracting

A. Introduction

Like agricultural seeds, many tree fruits dry as

they mature, and the seeds are extracted best at

low moisture contents. Other tree seeds are still

very moist at maturity, and special considerations are needed for extraction. No matter what

type of fruit is involved, however, the objective

of extraction is to obtain the maximum amount

of seeds in the best physiological condition in an

economically efficient operation. During extraction, seed quality can be greatly reduced by

excessively heating the fruit to force opening or

by extracting by hand or machine.

B. Objectives

1. Recognize potential problems of seed

extraction related to the type of fruit.

2. Identify the basic techniques of tree seed

drying and extraction.

C. Key Points

The following points are essential to seed drying and extracting:

1. For species that require drying, excessive

heat in the presence of high moisture content can be deadly.

2. Seed damage can occur during mechanical

separation.

3. Good training of workers is essential.

4. Extraction strategy depends on the type of

fruit involved.

D. Multiseed Fruits

Multiseed fruits include pods; moist, fleshy

fruits; cones and capsules; and other multiple

fruits. The steps in drying and extracting seeds

are:

1. For pods, Leguminosae

a. Dry the pods. Solar drying is fine, but

supplemental heating can be used also.

b. Thresh manually by:

(1) Flailing with poles

(2) Crushing by trampling

(3) Hitting with heavy mallets

c. Thresh mechanically with:

(1) Slow, rotating drums (cement

mixer)

(2) CSIRO flailing thresher (Willan

1985)

(3) Dybvig macerator

(4) Hammer mills

d. Use a series of steps for difficult species

(e.g., Prosopis): break the pods open;

soak in 0.1 N hydrochloric acid for 24

hours, wash and dry, and pound the

dried material with a hammer. For species that have gummy material in the

pods (e.g., P. cinerczria): break the pods

and run them through a coarse meat

grinder to extract the seeds; or break up

the pods, redry, and run them through

the thresher again.

2. For moist, fleshy fruits- In general,

pulp removal will help germination. But, if

fleshy coverings are thin, removal may not

be required (e.g., Vitex parvifloru).

a. Start quickly to avoid fermentation.

(Fermentation may help some species;

e.g., Mach-a pomifera in the United

States.) If this is not possible, spread in

thin layers and stir occasionally.

b. Soak the fruits in water until the pulp is

soft if the fruits cannot be squeezed

away easily with the fingers. The pulp

must be soft for complete removal.

Change the soak water to avoid fermentation.

c. Extract with macerators, mixers, coffee

depulpers (e.g., Gmelina arborea), feed

grinders, hammer mills, etc.; use anything that can tear up the fruits without

hurting the seeds.

d. Run small fruits (e.g., Rubus and

Morus) in blenders at slow speeds with

lots of water to extract the seeds.

e. Use a high-pressure stream of water

against mesh bags of fruit (e.g., Prunus

and Vitis).

f. Float pulp fragments and small underdeveloped seeds away with running

water.

3. For cones, capsules, and other multiple fruits-Drying for extraction assumes

successful predrying.

a. Air-dry on flat surfaces.

(1) Canvas is best for large quantities.

Use a tarpaulin (5 by 7 m or 5 by 10

m) to handle up to 12 to 16 hectoliters of Pinus caribaea cones in a

layer one cone thick. In good drying

weather, the cones will open in 3

days. Pinus caribaea should yield 4

to 6 kg of pine seeds from 16 hectoliters of cones. Agitate the cones every

2 to 3 hours to speed drying. Double

the tarpaulin over to cover the cones

at night. Remove seeds that have

dropped out each morning. There

will be some problems if it rains during drying (common in Central

America).

(2) Use screen trays for smaller lots,

such as single-tree collections. Use

larger trays to handle nearly 1 hec50
toliter at a time. Spread one cone

deep, stir, and protect from rain.

Elevate the trays to speed drying;

catch extracted seeds on a sheet or

cloth.

(3) Plastic sheets are not strong enough

for Pinus cones (1 hL of cones = 35

kg). Plastic also encourages condensation when the cones are covered.

Plastic sheets can be used indoors

for lighter fruits, (e.g., Liquidambar, Alnus, Casuarina, or Populus).

(4) Some common principles: protect

fruits from rain and predators,

spread them thin and stir frequently, and collect seeds on the drying surface as they fall. Protection

from rain is not needed if the fruits

will be artificially dried later.

(5) Dry some species under shade (e.g.,

Hopea spp., Triplochiton schelroxylon, and Pinus oocarpa); dry others

in direct sunlight (e.g., P. caribaea

and II! elliottii).

b. Use solar kilns.

(1) One simple type of solar kiln is clear

polyethylene stretched over the top

of screen trays; this method is common in east Africa.

(2) Solar heat storage units are more

sophisticated. Small buildings can

contain solar panels and heat storage facilities, such as water barrels.

Plans for one that can handle 15 to

18 hectoliters of cones at a time are

available. Such units are good for

easy-to-open cones of Pinus, such as

I? strobus, which open in 1 week

with good sun.

c. Use heated kilns. These kilns are most

efficient for large quantities of cones,

but capital investment and fuel costs

may be too high. There are several types

of heated kilns:

(1) Progressive kilns-Cone containers

move along a gradient of increasing

temperature inside the kiln.

Because they are slow, they are not

used much. They can be vertical or

horizontal. One company in the

United States has a tunnel progressive kiln with multitrayed, wheeled

carts and computerized environmental controls. A few have been

installed in Canada and the Northwest United States.

(2) Large-batch kilns -Large, heated

chambers in which trays hold cones

and rotate positions. Typical drying

capacity is 300 to 350 hectoliters per

day. There is no provision for small

lots, and fuel costs are high. Natural

gas is used in the United States, but

almost all these kilns have been

phased out.

(3) Small-batch kilns-Wire-bottom

drawers hold about one-third hectoliter of cones each. They are inserted

into a chamber in which a fan and

gas-fired heater push heated air

through the drawers. The standard

kiln holds about 16 hectoliters at a

time, but the design is modular for

possible expansion. One big advantage is that small lots can be kept

separate.

(4) Stack-tray system -Developed in

the Southern United States for

Pinus, it is a flexible and portable

system. Wooden trays with perforated sheet-metal bottoms (not wire

screens> fit together in stacks of six;

eight stacks are heated by one heating system. Heated air is blown up

through the trays from a bottom

plenum and recirculated to the

heater. One such unit can hold 100

hectoliters at a time and should process 7,000 hectoliters in a season.

Fuel costs are high, but flexibility is

an advantage.

(5) Tumbler driers-New cylindrical

batch kilns that rotate while drying

to remove seeds as soon as possible.

Humidity sensors and humidifiers

allow moisture and temperature

control. Many sizes are available to

fit individual needs.

(6) Other batch kilns -Many local

designs are available. The Bobbins

kiln, built in Honduras, seems to be

a good, low-cost, forced-draft kiln. It

has a capacity of 32 hectoliters, a

drying period of 12 to 18 hours

under good conditions, and uses old

cones or wood as furnace fuel.

d. Set temperature and humidity parameters. The object is to remove moisture;

high temperatures do this by creating a

greater vapor pressure gradient. For

conifers, 29 to 50 “C may be used, but

initial temperatures should always be on

the lower end of the scale. Pine cones of

the Southern United States are predried

51
to well below 50-percent cone moisture

before going into the kiln. Good predrying decreases cone moisture to 25 percent. Cones of F! contortu should be

predried to 20-percent moisture. Kiln

temperatures should be 43 to 49 “C, for

tough seeds, such as P taeda and P

elliottii, but 30 to 35 “C for “sensitive”

seeds, such as I? strobus and l? palustris. Cones should be dried to lopercent moisture content in the kiln for

best opening.

(1) Serotinous cones (e.g., I? banksiana,

I? pa&la, and P clausa) need 63 to

65 “C or a l&second dip in boiling

water to break the resin bonds

before going into the kiln. Live

steam can also be used to open these

cones. Pinus contorta cones can be

“scorched” by heating 2 minutes at

220 “C to melt resin, then opened by

heating 24 hours at 60 “C.

(2) Eucalyptus saligna seeds, in Brazil,

need 8 hours at 60 “C to dry capsules to below 20-percent moisture

for best extraction.

(3) Populus tremuloides seeds, in Canada, release at catkin moisture content below 70 percent; they are

picked 1 week early and air-dried for

3 to 6 days.

e. Extraction after drying-Once the

cones are open, a simple tumbling action

will extract the seeds.

(1) Tumbler driers were described previously.

(2) Cement mixers are versatile

machines that can also crush fruits

and extract and scarify seeds.

Extraction can be aided by placing a

wire cage inside the drum.

(3) Homemade tumblers are easy to

construct. Wire-screen boxes that

turn on a shaft can be tilted so that

seeds fall out.

E. Single-Seed Fruits

Single-seed fruits include drupes and fleshy

fruits and nuts with husks.

1. Drupes and fleshy fruits (Prunus and

Vitis) -Use macerators, mixers, etc.; use

water to soften the contact. (See “Moist,

fleshy fruits.“)

2. Nuts with husks (Juglam) -Use macerators or hand rubbing.

F. Sources

For additional information, see Willan 1985, p.

87-111.

II. Cleaning and Upgrading

A. Introduction

Cleaning seedlots is a basic step in proper seed

utilization. Cleaning should remove wings or

other seed appendages, empty seeds, damaged

seeds, and nonseed trash. This cleaning should

also provide dramatic decreases in insect and

disease problems. Many seedlots can be

upgraded by removing immature, damaged, and

dead seeds after the initial cleaning. Many people view large mechanical operations as the only

way to clean and upgrade seedlots, but seedlot

quality can be improved with simple equipment

and techniques.

B. Objectives

1. Learn the advantages of cleaned and

upgraded seedlots.

2. Become familiar with the principles of seedcleaning equipment and techniques and

expected results.

3. Apply these principles when seed cleaning

and upgrading are planned.

C. Key Points

The following points are essential for seed

cleaning and upgrading:

1 . Liquid flotation can be an essential aid for

many species, especially recalcitrant ones.

2. Screen cleaning is the basic seed-cleaning

method.

3. Air separation, including winnowing, is a

valuable technique.

4. Cleaning small lots for testing or research

may be very different from cleaning large

lots.

5. Upgrading seedlots offers potential

improvements in eight areas.

6. Seed sizing can be useful for some species

or sources but not for others.

D. Cleaning

1 . Flotation-The simplest technique of all.

Good seeds sink and bad ones float.

a. Initial moisture content is crucial

because it determines whether good

seeds sink or float. Long soaks (up to 24

hours) may be needed to make good

seeds sink if they are extremely dry

when collected.

b. Orthodox seeds are redried after flotation, unless they are sown immediately,

but recalcitrant seeds are not.

c. Flotation

(1) Removes light trash.

(2) Removes many empty, broken, diseased, or insect-damaged seeds.

(3) Is very good for large seeds with

high moisture contents (e.g.,

52
Quercus, Carya, and Juglans); and

for some small seeds (e.g., Liquidambar, Pinus, Juniperus, Robinia,

Gleditsia, and other legumes).

2 . Aspirators-Any machine that uses air to

clean and separate.

a. Large machines are found only in largescale, seed-cleaning plants.

b. Small-lot models are available for testing laboratories, research, or valuable

small collections. Devices include:

(1) General ER - a model with good air

control, but low capacity (75 to 100

mL).

(2) South Dakota-an old standby in

testing laboratories, with enough

capacity for small lots.

(3) Stults-a new blower with less

capacity than the South Dakota, but

much better air control. It has a vacuum gage to help standardization. It

has worked well with pines and

small legumes.

(4) Barnes-a laboratory blower that is

seldom seen anymore. It works well

with small conifers (e.g., spruce and

Douglas-fir) but lacks power to

separate Southern United States

pines.

(5) Other models - (see Willan 1985)

(6) Homemade fan devices -“ winnowing” type cleaning is fine, but

weight separations usually are not

good.

(7) Carter Day Duo Aspirator -a

machine that can handle a wide

range of seedlot sizes in continuous

flow.

3. Screens and sieves

a. Hand screens -a good set of hand

screens is extremely helpful, especially

for testing and research laboratories and

for small seedlots.

b. Mechanical screens -some small “flatscreen” cleaners use screen agitation

only; they are essential for large quantities.

4 . Airscreen cleaners-use both aspiration

and screening. These are the basic seedcleaning machines in most seed plants.

a. Air screens perform the following three

functions:

(1) Scalping - removes large materials

(twigs, leaves, etc.) with the top

screen.

(2) Sizing- drops small particles

through the second screen and

allows the seeds to remain.

(3) Aspiration -removes very light

material, including some empty

seeds.

b. These machines are more efficient for

cleaning, not sizing. Large machines

(for tree-seed plants) have five screens

and two air systems. They can clean 650

to 700 kg of Pinus seeds per hour. Most

plants use one machine to scalp (two to

three screens and one air system) and

another for final cleaning and sizing

(four to five screens and two air systems).

c. Small tabletop cleaners are great for

small lots; they can clean 30 to 40 kg of

small seeds per hour.

5. Electrostatic cleaners-The Helmuth

machine is good for small. seeds (e.g.,

Eucalyptus). It is expensive, however, for

such a limited function.

6. Dewinging- A special type of cleaning

that reduces storage volume, makes

upgrading possible for winged seeds (e.g.,

Pinus and Liriodendron), makes sowing

easier in nurseries, and removes sources of

pathogens. There are two basic methods of

dewinging - wet and dry.

a. The dry method is recommended for

tough seeds because of the damage

potential to thin seed coats. Pinus palustris is an exception.

(1) The popcorn polisher (Crippen

EP-26) is an old method that is

rarely used now because of damage

to seeds.

(2) USDA Forest Service dewinger from

Missoula Equipment Development

Center flails seeds with rubber fingers in a cylinder with soft rubber

on the walls to provide gentle action

and little damage.

(3) Dybvig macerators are good for

Liriodendron and Tilia bra&s.

(4) Electric drum scarifiers can clean

Populus, Salix, or other tiny seeds

with hairy appendages.

(5) New conifer dry dewingers have soft

rubber walls and 90 kg per hour

capacity in continuous flow.

b. The wet method is usually preferred for

pine and spruce seeds. It is quick

enough to avoid much moisture uptake,

but some redrying is usually needed for

most species.

53
(1) Cement mixers are good for medium

lots; seeds are sprayed with a mist

while mixing.

(2) Commercial dewingers are available

with capacities up to 90 kg per hour.

(3) Kitchen blenders can dewing small

lots; water is added to the seeds at

low speed for 10 seconds.

(4) Any cylinder with gentle agitation

will dewing; even rapid stirring by

hand will work for some species.

E . Upgrading

1 . Upgrading is improving the potential performance of a seedlot by removing empty,

damaged, weak, immature, or odd-sized

seeds. Many people clean their seeds but

never upgrade. Upgrading does not make

good seeds from bad but can certainly

improve the seedlot.

2. Upgrading will:

a. Remove weak seeds

b. Remove empty seeds

c. Reduce chances of insect and disease

damage by removing damaged seeds

d. Improve control of density in nursery or

germination beds

e. Reduce planting time in the nursery

f Facilitate nursery operations with uniform seedings

g. Reduce costs and improve uniformity in

container operations

h. Reduce storage space requirements

3. Methods and equipment

a. Specific gravity by flotation removes

empty and some damaged seeds.

(1) Water is used for some Pinus,

Quercus, and other large seeds.

(2) Organic solvents (usually alcohols)

of different densities work on some

small seeds but may damage the

seeds, especially if they are stored

for long periods. Seeds floated in

organic solvents should be sown

soon. The solution must always be

stirred vigorously to give every seed

a chance to float.

b. Air-screen cleaners, the basic cleaning

machines, can also be used to upgrade.

(1) They can separate by three physical

properties: size, shape, and density.

(2) They can upgrade some seedlots

(e.g., Platanus, Liquidambar, and

some Pinus spp.) by sizing or by

removing empties with the air system.

(3) Screen pattern, feed rate, airflow,

screen oscillation (pulleys), and

screen pitch (in some models) can be

regulated.

c. Air separators include large air-column

separators, fractionating aspirators, and

small laboratory blowers.

(1) Large air-column separators are not

common in tree-seed plants.

(2) Fractionating aspirators are now

widely used for Pinus in the United

States.

(3) Small laboratory blowers are suitable for small research or testing

lots.

d. Gravity separators were originally built

to remove ore from clay. Heavy particles

stay in contact with the shaking deck

and “walk up” the high side. Light particles “float” on the air cushion and

come off the low side.

(1) Separators can separate seeds of the

same size and different densities, or

different sizes and the same density;

they cannot separate a mixture of

densities and sizes.

(2) They are widely used to upgrade

conifer seeds in North America.

(3) The operator can regulate feed rate,

air stream through deck, deck pitch

(side and end), and eccentric thrust.

e. Electrostatic separators create a charge

that adheres to the seed surfaces.

(1) The Helmuth cleaner can handle

Eucalyptus, Platanus, and small

conifer seeds. Falling seeds receive

negative charges and are deflected

toward a positively charged plate;

deflection varies according to the

weight of the particle.

(2) Static electricity is used for very

small seeds. The sides of a plastic

beaker are wiped with a nylon cloth,

and the seeds and chaff are poured

into the beaker. The seeds are then

poured into a noncharged glass

beaker, and the chaff clings to the

sides of the first beaker.

f. X-ray radiography is typically used for

valuable research lots only. With Kodak

paper or Polaroid film, production is

good. One United States company has

studied mechanization of x-ray sorting.

g. Color separators remove light-colored

seeds. There is an application for highpriced lots, and one European company

has a color sorter in its production line.

h. The incubation, drying, and separation

(IDS) method is used on some Pinus and

54
Picea species in Sweden (Simak 1984).

The IDS method:

(1) Incubates seeds for 3 days in 100

percent relative humidity, 15 “C, and

light.

(2) Dries them for 12 hours at 15 “C, 35

percent relative humidity, and light.

(3) Separates dead from live seeds by

floating in water; in 5 minutes, viable seeds sink. Aspirators or gravity

tables can also be used to separate

the seeds.

(4) Redries the live seeds to 6- to 7-percent seed moisture.

(5) Can substitute stratification for 30

days at 4 “C for the incubation step

in (1) above for some species.

4. Sizing helps with some species or seedlots,

but not with others (table 8) Cno equivalent

table in Student Outline]; e.g., clonal mixes

vs. single family seedlots.

Table 8. -Correlation of seed size with germination rate and seedling

size (adapted from Bonner 1987b) [no equivalent table in

Student Outline]

F. Summary-See figure 29 Cno equivalent figure

in Student Outline] for a flow chart for extracting and conditioning Pinus seeds.

G. Sources

For additional information, see Bonner 198713;

Doran and others 1983, chap. 5; Willan 1985, p.

87-128.

III. Storage Principles

Species

Germination Seedling

rate size

_--_- _-- Correlation ---------

Acacia albida N o Yes

A. nilotica N o Yes

Acer saccharum Yes *

.

Araucaria angustifolia Yes Yes

Azadirachta indica N o Yes

Carya illinoensis . . . Yes

Picea abiest N o N o

l? abiest N o Yes

H glauca ..I Yes

Pinus elliottii N o Yes

l? koraiensis . . . Yes

I? roxburghii . . . Yes

R sylvestrist Yes N o

I? sylvestrist N o Yes

I? taedat Yes Yes

I? taedat . . . N o

Quercus alba . . . Yes

Q. ilex N o .

Q. petraea Yes

Q. prinus . . . Yes

Q. robur Yes

Q. rubra . . . Yes

Q. velutina Yes

Shorea contorta Yes Yes

Tsuga heterophylla . . N o

Theobroma cacao N o Yes

*Data not available.

tMore than one experiment reported for these species.

D.

E.

A. Introduction

The primary purpose of storing seeds is to have

a viable seed supply when it is needed for

regeneration. Successful storage of woody plant

seeds must be carefulIy planned, and good planning depends on an understanding of the purposes of storage, seed deterioration, and the

effwts of the storage environment on the deterioration processes.

B. Objectives

1. Learn the objectives and rationale of seed

storage.

2. Identify factors that affect seed longevity in

storage.

3. Review the general process of seed deterioration.

C. Key Points

The following points are essential to understanding seed storage principles:

1 . Longevity of seeds is a species characteristic.

2. Prestorage factors affect longevity in storage.

3. The most important factors in storage are

seed moisture content and temperature,

4. Seed deterioration begins at abscission and

involves complex physiological changes.

Objective of Storage

Once mature seeds are collected, deterioration

starts; deterioration is first manifested as

slower germination and then as death of seeds

and sometimes an’ increase in abnormal germination. The objective of seed storage is to

delay deterioration or decrease its rate until

seeds are used.

Rationale for Storage

Seed storage includes short- and long-term storage; it may be extended for long periods for

germplasm conservation.

1. Short-term storage:

ba:

C.

d.

Is used for immediate operations

Epically lasts less than 5 years

Allows carry-over of surplus production

in good seed years

Allows minimum storage environment

for most species

55
+

CONE STORAGE

(BAGS, BOXES)

1

HOT WATER DIP SEPARATION OF

SEEDS FROM DEBRIS

I

v

SEED DRIER

(CHECK h’OlSTURE TO DETERMINE NEED)

DEWINGER

(WETOR DRY) 4 I

FLOTATION OPTIONAL; I

1 I

LOELOLLY ONLY

Figure 29. -Flow chart for extracting and conditioning seeds of Pinus in a typical

extractory in the Southern United States (Banner 1991b) [no equivalent

figure in Student Outline].

2. Long-term storage:

a. Typically lasts from 5 to 10 years

b. Ensures constant seed supply for species that are irregular producers

c. Is used to save special lots that will not

be collected annually; e.g., in geographically remote areas

d. Requires very good storage environments

3. Germplasm conservation requires:

a. Very long-term storage with a goal of 50

years or more

b. The best storage environment or perhaps special technology

F. Longevity in Storage

Seed characteristics, seed handling before storage, genetics, and the storage environment

affect longevity in storage.

1. Seed characteristics

a. Basic physiology

(1) Orthodox seeds are tolerant of desiccation to low moisture contents

(usually less than 10 percent); dried

to this level, they may be stored at

subfreezing temperatures.

(2) Recalcitrant seeds are intolerant of

desiccation, usually dying if dried

below moisture levels of 25 to 30

percent or even 50 percent for some

tropical species; at these levels, subfreezing temperatures are lethal.

(3) These definitions were originated by

Roberts (1973). They are not “biologically logical,” but they are now

well established.

b. Seed structure-Thick or hard seedcoats restrict moisture uptake and gas

exchange; thin seedcoats allow too much

of both.

c. Seed chemistry - Oily seeds tend to be

harder to store than starchy seeds.

d. Stage of maturity-Immature seeds

usually will not store as well as seeds

that were fully mature when collected.

Therefore, maturity indices and time of

collection become very important.

e. Environmental stress during maturation -Drought, high temperatures, and

perhaps nutrient deficiencies may

decrease seed quality on the mother tree

and thus decrease storage potential.

5 6
2 . Seed handling before storage

a. Physiological mistreatment, such as

allowing overheating at high moisture

levels, will damage storage potential.

b. Processing can cause seedcoat cracks

and bruised tissue that can lower seed

quality, primarily by allowing invasion

of pathogens (table 9) [no equivalent

table in Student Outline].

3. Genetics- Good seed quality, and thus

greater storage potential, seems to be inherited to some degree. There are also genetic

differences among species; some are longer

lived than others.

4. Storage environment

a. Moisture content

(1)

(21

(3)

(4)

(5)

Moisture content is the most important factor.

Potential damage thresholds are

outlined in table 10 [table 5 in Student Outlinel.

The best range for orthodox seeds is

5 to 10 percent.

The best range for recalcitrant

seeds is full imbibition.

The equilibrium moisture content is

defined as the seed moisture content

when seed moisture is in equilibrium with the moisture in the

storage atmosphere (table 11) [table

6 in Student Outline].

(a) Equilibrium moisture content is

influenced by seed chemistry;

proteins are most hydroscopic,

then carbohydrates, then lipids

(figs. 30, 31) [no equivalent figure in Student Outline for figure 30; figure 31 is figure 9 in

Student Outline].

(b) Equilibrium is rarely reached

with recalcitrant seeds because

metabolism rapidly changes

seed weight and chemical condition.

(c) Sorption and desorption difference must be recognized.

b. Temperature

(1)

(2)

Generally, the cooler the seeds, the

slower the deterioration rate for

both orthodox and recalcitrant

seeds.

The safe temperature range for

orthodox seeds is related to the

moisture content of the seeds; 20

percent may be the critical upper

limit for 0 “C or lower (free water

and ice crystals), 15 percent for

Table 9. -Germination of unscarified and scarified seeds stored

under different conditions for 2 months (Lauridsen end

Stubsgaard 1987) [no equivalent table in Student Outline]

Seed storage Scarification method

Species condition Seed gun Seed burner

---------- percent ____...-_-

Acacia farnesiana Unscarifkd, 51 (211) 98 (22)

0 to 4 “C

Scarified, 30 “C, 47 (211) 91 (k3)

and 80 percent RH

Prosopis cineraria Unscarified, 75 (k3) 94 (*3)

0 to 4 “C

Scarified, 30 “C, 78 (~5) t

and 80 percent RH’

‘RH = relative humidity.

tProcess not used for this species.

Table 10. -Moisture content thresholds andpotential effects on stored

seeds [table 5 in Student Outline]

Moisture

content

Percent

>30

18 to 20

10 to 18

>9

5 to 8

<5

Effects

Germination begins

Overheating from respiration

Seed fungi become active

Insect activity

Best range for sealed storage

Desiccation damage possible in some species

Table ll.-Equilibrium moisture contents at 4 to 5 “C and three

relative humidities (Banner 1981b, Justice and Bass 1978)

[table 6 in Student Outline]

Species

Orthodox trees

Carya ovata

Juglans n&a

Liquidambar styraciflua

Liriodendron tulipifera

Picea abies

Pinus sylvestris

I? taeda

Prunus serotina

Orthodox crops

Glycine maz

Zea nays

Recalcitrant trees

Quercus alba

Q. nigra

Shorea robusta

Relative humidity

------------- percent ___________ __

2 0 4 5 9 5

Moisture content

------------- Percent _____________

I

. . . 10 1 5

1.. 11 20

. 8 20

1.. 10 19

6 8 . .

6 8 . .

10 17

. 9 17

6 8 19

a 12 20

. . . 37 50

. . . 13 29

. . . 35

*Data not available.

5 7
0

I I I I

30 4 5 60 7 5 90

RELATIVE HUMIDITY (PERCENT)

Figure 30.-Equilibrium moisture content at 25 “C for three orthodaz

species (adapted from Bonner 1972b) [no equivalent figure in Student Outline].

--- Quercus alba

-- - - Quercus muehlenbergN

~ Quercus shumordii

i= 50- ------ Quercus nigro /

z

.’

2 I/------ - ’

2

/’

h 40-

s

i SO- A--------

a

cc

__--

__--

/-+

20- ------* *_*- -___i--

IO I I I I

3 0 45 60 7 5 so

RELATIVE HUMIDITY (PERCENT)

Figure 31.-Equilibrium moisture content at 25 “C for four

recalcitrant Quercus species (adapted from Willan

1985) [figure 9 in Student Outlinel.

5 8

- 15 “C, and 13 percent for - 196 “C.

(a) Orthodox seeds at 5- to lopercent moisture can be stored

at most temperatures.

(b) Between 50 and 0 “C, every 5 “C

lowering of storage temperature

doubles the life of the seeds

(Harrington 1972).

(3) The safe temperature ranges for

recalcitrant seeds are:

(a) Temperate Zone species: - 1 to

3 “c!.

(b) Tropical species, usually above

12 to 15 “C because of chilling

injury.

c. Storage atmosphere

(1) Reduced oxygen slows metabolism

and increases longevity, but it is not

usually practical to regulate oxygen

level.

(2) Inert gases have been tested for

storage. They show no advantage in

long-term storage, but they may

help in short-term storage:

(3) In sealed containers, the COtJO,

ratio changes because seeds absorb

oxygen in metabolism.

(4) Recent work shows that Pinus radiata stored in nitrogen or carbon

dioxide retained viability long

enough for transport in the Tropics.

(5) Research in Indonesia showed that

coating Shorea pinanga seeds with

wax retained close to 50-percent

viability for 4 weeks; uncoated seeds

died during this period. The coating

reduced gas exchange.

G. Cells and Tissues During Seed Aging

The following changes occur in cells and tissues

during aging:

1 . Loss of food reserves by respiration.

2. Accumulation of metabolic byproducts from

respiration. Some are toxic.

3. Irreversible enzyme deactieation because

dried protein molecules lose their ability to

form active protoplasms on rehydration.

4. Deterioration of cell membranes: endoplasmic reticuhim and mitochondria.

5. Lipid peroxidation forming damaging free

radicals. (There is some question as to

whether peroxidation is the cause or an

effect.)

6. Alterations of DNA, causing genetic mutations and physiological damage.

H. Sources

For additional information, see Bonner and

Vozzo 1990; Warrington 1972; Justice and Bass
1978; Tang and Tamari 1973; Willan 1985, p.

129-160.

IV. Storage Applications

A. Introduction

The previous section introduced the principles

and critical factors that influence seed longevity. This section discusses how these principles

are applied in practice to store tree seeds.

B. Objectives

1. Relate seed storage principles to prescriptions for each species group.

2. Learn features of cold storage units.

3. Discuss storage constants and their application.

4. Learn basic principles of seed storage management.

C. Key Points

The following points are essential to storage

applications:

1. There are four classes of seed storage

behavior.

2. Cold storage is best but not always necessary for successful seed storage.

3. Each species, and perhaps individual populations within a species, will nearly always

respond identically to a given set of storage

conditions.

4. Good facilities and good seeds are not

enough; there must be good management

for optimum seed storage.

D. Seed Storage Classes

Experience in storing tree seeds suggests an

expansion of Robert’s definitions of seed storage

classes (Bonner 1990). There are four classes of

tree seed storage behavior (table 12) [table 7 in

Student Outline].

1. Due orthodox

a. True orthodox seeds are tolerant of

desiccation (table 13) [table 8 in Student

Outline], and:

(1) Can be dried to 5- to lo-percent

moisture levels.

(2) Can be stored at subfreezing temperatures.

(3) Are easily stored for one rotation

under these conditions.

(4) Have generally unknown upper

limits of storage.

b. Examples include most of the valuable

temperate genera (Pinus, Picea, Betula,

Prunus) and many tropical genera

(Acacia, Eucalyptus, and Casuarina).

2. Suborthodox

a. Suborthodox seeds require the same

conditions as true orthodox seeds but

are limited to shorter periods (table 14)

[table 9 in Student Outline].

(1) They are stored under the same conditions.

(2) Some limits to storage potential are

high lipid contents, thin seedcoats,

and genetic makeup.

b. Examples include:

(1) Seeds with high lipid content (Juglam, Carya, some A&s, and Pinus).

(2) Seeds with thin seedcoats (Popuhs

and Sal&).

(3) Seeds that must be dried slowly

(Fagus and Citrus).

(4) Genetics; e.g., [Have the students

suggest some].

c. These species could be classed as “true

orthodox” if the limitations were overcome. Gmelina arborea is one borderline

species.

Table 12. -Storage conditions for four storage classes of tree seeds [table 7 in Student

Outline]

Storage Storage Seed

class period moisture Temperature Container type

Years Percent “C

True orthodox <5 6-10 o-5 Airtight

>5 6-10 -18 Airtight

Suborthodox <5 6-10 o-5 Airtight

>5 6-10 - 18 Airtight

Temperate

recalcitrant <3 30-45 - 1 to -3 4-mil’ plastic, unsealed

Tropical

recalcitrant <l 30-45 12-20 4-mil plastic, unsealed

‘mil = l/1,000 inch = 0.025 mm.

59
Table 13. -Storage test results for true orthodox species (adapted from Bonner 1990)

[table 8 in Student Outline]

Species

Abies procera

Acacia leptopetala

A. mangium

A. pruinocarpa

Acer saccharum

Albizia falcataria

Araucaria cunninghamii

A. cunninghamii

Casuarina equisetifolia

C. torulosa

Liquidambar styraciflua

Pinus caribaea

var. hondurensis

F! elliottii

P merkusii

l? ponderosa

Tectona grandis

Tsuga heterophylla

T. heterophylla

Storage conditions

Seed

Temperature moisture

“C Percent

0 9

20-25 . . . *

4-8 . . .

20-25

-10 10

4-8 ..I

-15 16-23

19 7

- 3 6-16

20-25 8-12

3 5-10

Storage results

Time Viability

stored loss

-

Years Percent

7.0 11

18.0 1

1.2 6

16.0 2 0

5.5 5

1.5 10

8.0 few+

0.1 0

2.0 O-5

18.0 6

9.0 3

8 . . .

4 10

4-5 <8

0 8

0 4 =12

5 8

-18 8

2.1 k16

50.0 30

4.0 0

7.0 0

7.0 0

2.0 0

2.0 0

‘Data not available.

+Exact value not available from original source.

Table 14. -Storage test results for suborthodox species (adapted from

Bonner 1990) [table 9 in Student Outline]

Species

Citrus lirrwn

Fagus sylvatica

Gmelina

arborea

Populus

deltoides

Salk glauca

Storage conditions Storage results

Seed Time Viability

Temperature moisture stored loss

“C Percent Years Percent

-20 5 0.9 t5

-10 10 5.0 3 4

- 5 6-10 2.0 10

-20 6-10 6.0 21

-10 6-10 1.2 0

Table E.-Storage test results for temperate recalcitrant species

(adapted from Bonner 1990) [table 10 in Student Outline]

Species

Storage conditions

Seed

Temperature moisture

Storage results

Time Viability

stored loss

“C Percent Months Percent

Acer

saccharinurn - 3 50 18 8

Quercus falcata var.

pagodaefolia 3 35 30 6

Q. robur - 1 4045 29 31-61

Q. rubra -It/o - 3 3845 17 18-46

Q. virginiana 2

*

. 12 35

*Data not available.

60

3. Temperate recalcitrant

a. Temperate recalcitrant seeds are ‘intoIerant of desiccation (table 15) [table 10

in Student Outline].

(1) They cannot be dried below 20- to

30-percent moisture; thus, storage

must generally be above freezing,

although some Quercus species have

been reported to survive storage at

-2 “C.

(2) Metabolism and pregermination are

common in storage.

(3) They cannot be stored in airtight

containers; some gas exchange is

necessary (table 12) [table 7 in Student Outline].

b. Examples include Quercus (high lipid)

and Aesculus (high carbohydrate).

4. Tkopical recalcitrant

a. Tropical recalcitrant seeds have the

same moisture and gas exchange requirements as temperate recalcitrant

seeds, but they are sensitive to low temperature. Chilling damage and death

will occur below 12 to 20 “C, depending

on the species (table 16) [table 11 in

Student Outline].

b. They are the most difficult group to

store, even for short periods.

c. Examples include species of Shorea,
Table 16.-Storage test results for tropical recalcitrant species Table 17. -Storage test results for cryogenic trials of forest tree seeds

(adapted from Bonner 1990) [table 11 in Student Outline] (adapted from Bonner 1990) [Table 12 in Student Outline]

Storage conditions Storage results

Seed Time Viability

Species Temperature moisture stored loss

“C Percent Days Percent

Araucaria

hunsteinii 19.0 25-30 5 4 *30

A. hunsteinii 2.0 3 0 365 82

Azadirachta indica 26.0 lo-18 5 6 65

Hopea helferi 15.0 4 7 3 7 2

Shorea robusta 13.5 40-50 3 0 6 0

S. roxburghii 16.0 4 0 270 230

Hopea, Dipterocarpus, and even some

legumes; e.g., in Costa Rica, Pithocellobium.

E. Cryogenic Storage

Cryogenic storage is a method for very longterm storage for germplasm conservation (table

17) [table 12 in Student Outline].

1. Techniques

a. Packages of seeds are immersed in liquid nitrogen (- 196 “C) or suspended

above it in the vapor.

b. The potential for germplasm conservation is good for small quantities but not

for bulk lots.

c. The upper (maximum) time limits of

viability retention are not known. Only

a few tests have been made on tree

seeds.

2. Costs-Comparable with conventional

storage in some cases of small seeds: $0.30

to $1.41 per year per sample of 1,000 seeds.

F. Physical Facilities

The optimum storage environment requires

cold storage units, proper containers, and moisture management.

1. Cold storage units

a. Cold storage units require a reliable

power source, should not be used where

floods or earthquakes may occur, should

be located near other seed activities,

should be rodent proof, and should be on

high elevations when possible because

ambient temperatures will be cooler.

b. They should be built to hold a 5-year

supply (or whatever the operational

time).

c. For germplasm conservation, 1 liter is

needed of each sample (3,000 to 12,000

seeds) for medium-sized seeds (wheat

and rice); e.g., 85 ms should hold 22,800

samples.

Soecies

Seed

moisture

Time

stored

Viability

105s

Abies alba

A. concolor

Fagus sylvatica

Larix decidua

Picea abies

Pinus echinata

I? ponderosa

l? sylvestris

Populus tremula X

tremuloides

Ulmus pumila

Percent Days

*

. 6

<13 180

. . . 6

. . . 6

6

. 112

Cl3 180

1.1 6

. 6

. . . 112

Percent

5

0

100

5

1

0

0

0

1

0

‘Data not available.

d . Humidity control is not recommended in

the Tropics; the seeds are dried and

sealed in containers. If the power fails,

seeds stay dry; this method is less

expensive than humidity control.

e. Direct or indirect vapor-compression

refrigeration is best for the Tropics; it is

usually available and reliable.

f. Standby generators and safety alarms

are recommended.

g. When the power goes off and cooling

stops, thermal time constants of 4 to 5

days should apply in large coolers, but if

orthodox seeds are dry, serious damage

may not occur for 2 weeks.

h. Modular panel units are effective.

i. Insulation depends on ambient conditions. A reliable local refrigeration specialist should be employed; an R value of

35 (heat transfer coefficient of 0.029 or

less) is recommended.

2. Containers

a . Fiber drums are very effective. Common

capacities are 0.45 and 0.90 hL (25 and

50 kg for Pinus taeda).

b. Generally, plastic is better than glass;

plastic bags can be inserted inside glass

containers.

c. Rectangular containers utilize space

better than round ones, but air spaces

are still needed between containers.

Round containers assure air spaces.

d. Plastic bags should be 0.1 to 0.2 mm

thick in humid atmospheres.

3. Moisture management

a. Seeds will reach an equilibrium moisture content when exposed to the storage atmosphere.

6 1
b. If the storage unit has humidity control

(50 to 60 percent relative humidity),

orthodox seeds need not be sealed.

Recalcitrant seeds cannot be sealed, so

they cannot be stored in such a unit; the

low humidity would desiccate the seeds.

c. Without humidity control, relative

humidity will be 95 percent or more,

which is fine for recalcitrant seeds.

Orthodox seeds must be dried and

stored in sealed containers in such a

unit.

d. Humidity control is not recommended

for the Tropics because both orthodox

and recalcitrant seeds will be stored in

the same facility.

e. Frost-free refrigerators are an alternative for humidity control because these

systems remove the moisture from the

unit.

G. Genetic Damage in Long-term Storage

1 . Long-term storage could be devastating to

germplasm conservation, but there is no

strong evidence to date of lasting damage.

Point aberrations occur on chromosomes,

but they- are not passed along to the next

generation (fig. 32) [no equivalent figure in

Student Outlinel.

C

//I\

;ii

D

Figure 32.-Chromosome aberrations found in germinated fresh and

50-year-old Pinus echinata: (A) single bridge, (B) double

bridge, (C) single fragment, (0) multiple fragments

(Burnett and Vozzo 1985) [no equivalent figure in Student Outline].

2. Population changes are almost certain in

heterogenous seedlots; some parts of the

population die sooner in storage. The

impact of this effect is still unknown.

H. Retesting in Long-term Storage

1. Test procedures--ETA (1985) recommends 4 replications of 100. Subsequent

tests can be 2 replications of 100; if viability

has fallen 5 percent, the test is repeated

with another 2 replications of 100.

2. Test interval for orthodox seedsShould be initial year, third year, and every

fifth year thereafter.

3. Nondestructive testing- Leachate conductivity could be used.

4. Regeneration-In annual plants,

regeneration is done when viability falls to

50 percent. There may be a better plan for

trees.

I. Viability Constants in Storage

The use of viability constants in storage is

another technique introduced by Roberts

(1973).

1. Theory-Viability retention will,be the

same for a given species under a given set of

storage conditions. This property can be

expressed as a “viability constant” for each

species.

2. Practice

a. Good results have been obtained with

agricultural seeds.

b. Critics say varieties of a single species

will differ.

c. One must start with very good seeds.

d. There are few data for tree seeds.

Tompsett (1986) has developed constants for several tree species, and other

data are available for two pines, Liquidambar styruciflua, and P&anus occidentalis. The two hardwoods fit the

expected pattern, but the pines have

shown damage repair at high moisture

levels after 8 years.

e. The above species are all orthodox. A

different technique is needed to determine viability constants for short-lived

recalcitrant species.

3. Viability constants for forest species

could be useful in long-term storage for

germplasm conservation or when good storage conditions are not available.

ZBonner, Franklin T. Ln.d.1 Unpublished research notes. On file

with: U.S. Department of Agriculture, Forest Service, Southern Forest Experiment Station, Starkville, MS 39759.

62
4. Procedures

a. Store samples in many combinations of

temperatures and moisture contents,

and test frequently.

b. Plot seed death over time for each condition, using probit transformations.

c. Calculate coefficients for time, temperature, and seed moisture content.

d. See table 18 [no equivalent table in Student Outline].

J. Summary

Review table 12 [table 7 in Student Outline].

K. Sources

For additional information, see Bonner 1990,

Bonner and Vozzo 1990, Chin and Roberts

1980, Harrington 1972, International Board for

Plant Genetic Resources 1976, Justice and Bass

1978, Roberts 1973, Tang and ‘Ikmari 1973,

Willan 1985.

Table 18. -Viability equation’ coefficients for seeds of Ulmus carpinifolia and Terminalia brassii (Tornpsett 1986) and four

United States species [no equivalent table in Student

Outline]

Species K*: cw C” C”

Liquidambar styraciflua 5.3435 1.7616 0.0307 0.000869

Pinus elliottii 5.2463 0.9832 0.0508 0.000571

P taeda 3.2783 0.7300 0.0348 0.000328

P&anus occidentalis 5.1013 1.6742 0.0354 0.000838

Terminalia brassii 4.9990 2.1490 0.0350 0.000410

Ulmus carpinifolia 5.7150 2.9660 0.0340 0.000408

‘V = Ki - pl,,& - C,Jog,fl - C,t - Cptz,

where

V = probit percent viability,

Ki = initial viability,

p = storage time (days),

KE = species constant,

m = percent moisture,

t = temperature (“0, and

C, CH, C, are coefficients.

6 3

65
I. Sampling

A. Introduction

Sampling is the process of taking a small part

or quantity of something for testing or analysis;

it is the first step in seed testing. In sampling, it

is essential: (1) to obtain a sample of proper

size, and (2) to obtain a sample representative

of the main seedlot. The results of the laboratory tests can only show the quality and characteristics of the sample submitted for the

analysis; therefore, the validity of test results

for a large seedlot is determined by the success

of obtaining a representative sample. Sampling

seedlots for quality evaluation must be done

systematically, using appropriate techniques,

tools, and procedures, to ensure that the seed

sample represents the entire lot.

B. Objectives

5.

6.

least the minimum specified size and may

be comprised of either all or part of the

composite sample.

Working sample -a subsample taken

from the submitted sample in the laboratory on which one of the quality tests is

made.

Subsample- a portion of a sample

obtained by reducing the sample by recognized methods (tables 19, 20) Ino equivalent for table 19 is included in Student

Outline, table 20 is table 13 in Student Outline].

E. Sampling Intensity

1. Quantify a seedlot according to accepted

standards.

2. Determine sampling intensity according to

size and characteristics of the seedlot.

3. Learn about appropriate sampling instruments and techniques according to recognized standards.

C. Key Points

A sample is obtained by taking small portions at

random from various positions in a seedlot and

combining them. From this sample, other subsamples are obtained by one or more steps. At

each step, thorough mixing is followed either by

progressively subdividing or by selecting at

random still smaller portions and then recombining.

‘Iable 19. -Guidelines for minimum working sample weights for tree

seeds when both purity and germination are to be tested

(adapted from Bonner 1974~) [no equivalent table in Student Outline]

The following points are essential in seed

sampling:

1 . Laboratories can only measure the properties of the sample; the sampler must ensure

that the sample truly represents the seedlot.

2. Submitted samples should contain at least

2,500 seeds (except for very large seeds of

certain species).

Seeds

Per

pound

Minimum

working

sample

size

Seeds

per

am

Minimum

working

sample

size

Number Ounce Number Grams

3. Drawing the sample must be completely

random.

4. Proper packaging and labeling of the sample are essential.

D. Definition of Terms

Relevant terms are defined below:

1. Lot -a specified, physically identifiable

quantity of seeds. A lot, or seedlot, may be

small and in a single container, or large and

in several containers.

2. Primary sample- a small quantity of

seeds taken from one point in a seedlot.

3. Composite sample -formed by combining and mixing all the primary samples

taken from a seedlot. The composite sample

is usually much larger than required for

seed testing and is reduced to a smaller

sample for testing.

<2,000’ 16+ less than 5’ 500+

2,000-2,500 14-18 5-7 300-400

2,500-3,000 11-16 7-10 200-300

3,000-3,500 10-13 10-15 140-240

3,500-4,000 8.5-11.5 15-20 100-170

4,000-4,500 7.5-10.0 29-25 85-125

4,500-5,000 6.5-9.0 2530 70-100

5,000~5,500 6-8 30-35 60-90

5,500-6,000 5.5-7.5 35-40 54-75

6,000-7,000 5-7 40-50 42-65

7,000-8,000 4-6 50-60 36-54

8,000-9,000 3.75-5.50 60-70 30-46

9,000-10,000 3.25-4.75 70-80 27-40

10,000-15,000 2.25-4.25 80-90 24-35

15,000-20,000 1.75-3.00 go-100 22-32

20,000-25,000 1.50-2.25 loo-125 17-28

25,000-30,000 1.25-2.00 125-150 15-23

30,000-40,000 0.8-1.5 150-175 13-20

40,000~50,000 0.75-1.25 175-200 11-17

50,000-65,000 0.5-1.0 200-250 9-15

65,000-80,000 0.40-0.75 250-300 a12

80,000-100,000 0.3-0.6 300-350 6.5-10

100,000-150,000 0.25-0.50 350-400 5.5-8.5

150,000-200,000 0.2-0.4 400-500 4.5-7.5

200,000-300,000 0.1-2.5 500-750 3-6

~300,000 0.1 >750 3

4. Submitted sample-the sample submit- *Purity analyses are rarely required for seeds of this size.

ted to the testing laboratory. It must be at +Sample should contain at least 500 seeds.

66
Table 20. -Weights of lots and samples for shrubs and trees (ISTA

1985) [table 13 in Student Outline]

Sr3ecies

Maximum Working sample

weight of Submitted for purity

seedlot sample analysis

Kilograms Grams Grams

Acacia spp. 1,000 70 35

Ailanthus altissima 1,000 160 8 0

Alnus rubra 1,000 15 2

Castanea s&vu 5,000 500 seeds 500 seeds

Cedrela spp. 1,000 8 0 40

Eucalyptus

camaldulensis 1,000 15 5

E. glob&s 1,000 6 0 20

E. tereticornis 1,000 15 5

Morus spp. 1,000 2 0 5

Pinus halepensis 1,000 100 5 0

P wallichiana 1,000 250 125

Quercus spp. 5,000 500 seeds 500 seeds

Robinia

pseudoacacia 1,000 100 50

1. Calculating primary samples-Each

composite sample must be made up of at

least five primary samples. When more

than one sample is taken from a drum, they

are taken from well-separated points. For

lots of one to six drums, each drum is

sampled, and at least five primary samples

are taken. For lots of more than six drums,

five drums plus at least 10 percent of the

number of drums in the lot are sampled.

For example:

Number of drums in the lot

12 3 4 5 6 7 10 23 50

Number of drums to sample

12345666710

Total number of primary samples

55555666710

2. Seedlot size-For international trade in

tree seeds, a maximum size of a seedlot for

most species has been set at 1,000 kg t5

percent (table 20) [table 13 in Student Outline]. This maximum is recommended for

all domestic transactions also. If the

amount of seeds exceeds this maximum, it

should be divided into lots not larger than

the maximum, and each portion should be

given a separate lot identity.

F. Sampling Procedures

There are three common sampling tools or techniques: triers, soil dividers, and the extended

hand method.

1. Triers- Free-flowing seeds can be most

easily sampled using tools called “triers.” A

trier consists of a tube that fits inside an

outer sleeve with a pointed end. The tube

and the sleeve have slots in their walls, and

when the slots in the tube line up with slots

in the sleeve, seeds can flow into the cavity

of the tube. A half turn of the tube closes

the openings. Triers are available in various

lengths and diameters and can be inserted

into the seed containers either vertically or

horizontally. If used vertically, the trier

must have partitions dividing the tube into

a number of compartments to obtain a representative sample. The steps are:

a. Close the gates before inserting the trier

into the drum.

b. Insert the trier into the drum.

c. Open the gates and let the seeds fill the

trier.

d. Close the gates.

e. Remove the trier.

f. Dump the seeds out of the trier.

2. Soil dividers-These devices are made to

mix and divide soil samples, but they work

well on free-flowing seeds. $oil dividers are

a good tool for small lots only. The steps

are:

a. Pour the seeds through the divider several times for mixing.

b. Divide the sample into halves, quarters,

etc., until the desired size is reached.

3. Extended hand method-Requires no

special equipment. It is recommended for

chaffy, winged, or other nonflowing seeds,

but it will work on any seeds.

The steps are:

a. Extend the fingers, and insert the hand

straight into the seeds.

b. Close the hand, and withdraw a primary

sample.

G. Preparation of the Sample

1. Composite sample-The composite sample is prepared by combining all primary

samples and mixing. If it is small enough, it

can be sent as the submitted sample. If it is

too large, as is common, it can be reduced

with a soil divider. See tables 19 [no equivalent in Student Outline] and 20 [table 13

in Student Outline] for determining sample

weights.

2. Working sample-The submitted sample

is reduced to a working sample by thoroughly mixing the submitted sample and

then repeatedly halving the sample by one

of the following methods:

a. Mechanical divider method-This

method is suitable for free-flowing

seeds. An apparatus divides a sample

into two approximately equal parts. The

sample is reduced by repeated halving

6 7
until a working sample equal to or slightly

more than the prescribed size is obtained.

Some types of mechanical dividers are the

conical (or Boerner), the centrifugal (or

Garnet), and the soil divider type.

b. Random cups method- Small cups or

containers, usually not more than eight,

are randomly placed on a tray and the

seeds of the submitted sample are

poured evenly over the tray. Most seeds

fall on the tray, but those that collect in

the cups are combined in a larger container as the working sample. The process is repeated until the necessary

quantity of seeds is collected.

c. Modified halving method- In this

method, a grid of equal-sized cells is

used; all cells are open at the top, but

alternate cells have no bottom. The grid

is placed on a tray, and the submitted

sample poured evenly over it. The grid is

then lifted with the seeds collected in

the cells being used as the working sample. The process is repeated until the

desired sample size is achieved. A 50- by

50-mm cell size is useful for conifer

seeds.

d. Spoon method-After mixing, the submitted sample is poured evenly over a

tray. Small portions of seeds are

removed with a spoon and spatula from

at least five random positions until the

required amount is obtained. This

method is used only for species with

small seeds.

e. Manual halving-Chaffy, winged, or

large seeds are spread on trays and

divided in halves with the hand, fingers

extended together.

3. Disposition of the extra seeds-The

remainder of the submitted sample should

be stored to permit retesting if necessary.

International Seed Testing Association

(1985) recommends storing for 1 year.

H. Sources

For additional information, see Association of

Official Seed Analysts 1988, Edwards 1987,

International Seed Testing Association 1985.

II. Moisture Content

A. Introduction

The first measurements taken in seed testing

are moisture, purity, and weight. All of these

measurements are important, but moisture is

the most critical one. Seed moisture levels can

influence or indicate seed maturity, longevity in

storage, and the amount of pretreatment

needed for rapid germination.

B. Objectives

1 . Learn the principles of official seed testing

for moisture.

2. Apply these principles in practical exercises.

C. Key Points

The following points are essential to seed moisture content:

1 . Procedures are prescribed in detail for official testing.

2. Many tests may be unofficial, and different

methods may be used, but accuracy and

precision are still essential.

3. Large recalcitrant seeds present special

problems that official testing rules (ISTA

1985) have not yet adequately addressed.

D. Definition of Terms

Relevant terms are defined as follows:

1 . Sample, submitted-the sample of seeds

submitted to a seed-testing station; it

should be twice the size of the working

sample.

2. Sample, working- a reduced seed sample

taken from the submitted sample in the

laboratory on which some test of seed quality is made.

3. Seedlot -a specified quantity of seeds of

reasonably uniform quality; the maximum

lot size is 1,000 kg or 5,000 kg for Fagus

and larger seeds.

E. Moisture Measurements

1. Importance

a. Moisture content is the most important

factor in viability retention.

b. Insect and disease activity occurs at certain moisture levels (table 10) [table 5 in

Student Outline].

c. Moisture content influences the relationship of weight to number of seeds.

2. Frequency

a. Moisture is measured after extracting

and cleaning. The seeds may have been

too moist during cleaning.

b. Moisture is measured when seeds are

stored. The correct moisture content

must be reached, or more drying will be

necessary.

c. Periodic checks during storage reveal

whether the container seals are good.

d. Moisture is measured when seedlots are

shipped. Seeds shipped moist will lose

quality rapidly.

3. Procedures-Moisture content is measured by:

68
a. Using the submitted sample.

b. Measuring immediately on receipt.

However, if the sample bag is not moisture proof, the measurement will be

meaningless.

c. Expressing results as a percentage of

fresh weight (wet weight), not dry

weight. This is the international convention for seed moisture. If dry weight percentages are needed, nomograms are

available for conversion.

4. Methods-Moisture content is measured

by four methods:

a. Ovendrying. Critical points are:

(1) Heat samples for 17 + 1 hours at

103 lt2 “C, a good overnight schedule. This is the official ISTA

method.

(2) A forced-draft oven, not a gravity

convection oven, is used.

(3) Glass or metal containers with

rounded sides and base and closefitting tops are used.

(4) Space equal to one container’s diameter is allowed between containers

on the oven shelf. The tops are

removed when the containers are

placed in the oven.

(5) Desiccators are used to cool the

samples for 30 to 45 minutes. The

container covers are replaced before

they are transferred to the desiccators.

(6)

(7)

(8)

(9)

Ambient humidity should be less

than 70 percent in the laboratory. If

not, the samples should be cooled for

an additional 30 minutes in the

desiccator.

All weights should be to the nearest

milligram. The ideal balance is a

top-loading, electronic instrument.

The ISTA requires grinding for

Fagus, Quercus, and all other large,

recalcitrant seeds. At least 50 percent of the ground material must

pass 4.00-mm mesh sieves. However, these seeds must be predried

before grinding because of their

moisture content (except perhaps

Fagus).

Predrying is required if moisture

exceeds 17 percent in seeds that

must be ground and 30 percent in

other species. Predrying should be

at 130 “C! for 5 to 10 minutes or

overnight in a “warm place” (ISTA

1985).

(10) Tolerance is more liberal for tree

seeds than for, agricultural seeds

because of tree seeds’ large size,

higher moisture, and more natural

variation (table 21) [table 14 in Student Outline]. (See ISTA 1985, table

9D.)

b. Electric meters

(1) Electric meters are not allowed for

ISTA official tests but are useful for

quick checks during processing and

storage.

(2) They are based on electrical

resistance or capacitance and are

accurate to within t 1 percent on

most free-flowing seeds.

(3) All meters are made for grains;

thus, calibration charts must be

constructed for tree seeds. [see

exercise 71

(4) Manufacturers of electric meters

are:

(a) Motomco - based on capacitance

and very accurate.

(b) Radson (Dole or Seedburo)an

established, reliable meter in

the United States, based on

resistance.

cc> Dickey-John or Instobased on

capacitance. These meters are

portable and battery operated

only.

(d) Super-Beha - widely used in

Europe.

c. Infrared balances - These small,

infrared ovens have built-in balances.

They use a gravimetric method based on

drying time that has given good results

on tree seeds in many places in the

world.

d. Laboratory methods for research:

(1) The Karl Fischer method uses direct

measurement and is the most widely

used reference method.

Table 21. -Tolerance levels for differences between two determintiions

of moisture content of tree and shrub seeds (ZSTA 1985)

[table 14 in Student Outline]

Seed size

class Seeds per kilogram Initial moisture Tolerance

Number ----------__ percent ____________

Small seeds >5,000 Cl2 0.3

Small seeds >5,000 >12 0.5

Large seeds c5,ooo Cl2 0.4

Large seeds <5,000 12-25 0.8

Large seeds <5,000 >25 2.5

69
F .

G.

(2)

(3)

(4)

Toluene distillation was once

allowed in ISTA for oily seeds but

has been discontinued for safety

reasons.

Nuclear magnetic resonance (NMR)

requires sophisticated equipment

and must be referenced to the Karl

Fischer method; NMR can measure

moisture nondestructively in an

intact seed.

Infrared spectroscopy is a destructive test that must also be referenced to Karl Fischer measurements.

Summary

See table 22 [table 15 in Student Outline].

Sources

For additional information, see Bonner 1981b;

International Seed Testing Association 1985,

sections 9, 9A; Willan 1985, p. 227-230.

III. Purity and Weight

A.

B.

C.

Introduction

After moisture content has been determined,

the submitted sample is ready for purity and

weight determinations. These determinations

are a vital part of official seed testing and practical seed use, with legal ramifications in both

domestic and international seed trade.

Objectives

1 . Learn the principles of official seed testing

for purity and weight.

2. Apply these principles in practical exercises.

Key Points

The following points are essential to determine

seed purity and weight:

1 . The line between true seeds and trash can

be ambiguous for some tree seeds,

especially those that are dewinged.

2. Patience and good eyesight are needed.

Table 22.-Suggested test procedures for tree seed moisture (Bonner 1981b) [table 15 in Student

Outline]

Seed size class

Accurate measurement or

ISTA official test Rapid estimate

Small seeds, low oil content

(e.g., Platanus, Robinia)

Small seeds, high oil

content (e.g., Abies,

Pinus, Tsuga,

Zanthoxylum)

Large seeds, low oil content, (1) Grind or equivalent

moisture <20% (e.g., (2) Oven: 103 a2 “C for 17

Nyssa) ? 1 hours.

Sample: 4 to 5 g or

enough to equal weight

of live seeds

Large seeds, low oil content,

moisture >20%, (e.g.,

Aesculus, Quercus)

Large seeds, high oil

content (e.g., Carya,

Fagus, Juglans)

Oven: 103 C2 “C! for 17 2 1

hours.

Sample: 4 to 5 g

Oven: 103 It 2 “C for 17 C 1

hours.

Sample: 4 to 5 g

or

Toluene distillation

(1) Predry to <20% at 130

“C for 5 to 10 minutes

(2) Grind or equivalent

(3) Oven: 103 -t2 “C for 17

k 1 hours.

Sample: enough to

equal weight of five

seeds

(1) Grind or equivalent

(2) Oven: 103 t2 “C for 17

t 1 hours.

Sample: enough to

equal weight of five

seeds

01

Toluene distillation

Electric meter

Sample: SO to 200 g,

depending on type

Electric meter

Sample: 80 to 200 g,

depending on type

Microwave drying

Sample: 4 to 5 g or enough

to equal weight of five

seeds

Microwave drying

Sample: enough to equal

weight of five seeds

Microwave drying

Sample: enough to equal

weight of five seeds

70
3. The smaller the seeds, the more difficult the

purity test will be.

D. Definition of Terms

Relevant terms are defined as follows:

1. Purity-the proportion of clean, intact

seeds of the designated species in a seedlot,

usually expressed as a percentage by weight

2. Sample, submitted-the sample of seeds

submitted to a seed-testing station; it

should be twice the size of the working

sample

3. Sample, working- a reduced seed sample

taken from the submitted sample in the

laboratory on which some test of seed quality is made. For size of sample, see table 20

(ISTA 1985, table 2.A.Q or the values

based on seed size (table 19) [no equivalent

table in Student Outline].

4. Seedlot -a specified quantity of seeds of

reasonably uniform quality. The maximum

lot size is 1,000 kg (5,000 kg for Fagus and

larger seeds).

E. Purity

1. Procedure-The ISTA (1985) rules are

followed in purity testing. The steps are:

a. Reduce the submitted sample (after

mixing) to the working sample with:

(1) Mechanical dividers

(2) Random cups

(3) Modified halving

(4) Spoon method

(5) Manual halving (chaffy, winged,

and large seeds)

b . Divide the working sample into fractions

of

(1) Pure seeds

(2) Other seeds

(3) Inert matter

c . Weigh and express each as a percentage

of the total sample weight

2. Pure seed component -This component

contains:

a. Intact seed units of the desired species

b. Pieces of seed units larger than one-half

the original size, even if they are broken

3. Tree seed specifics

a. Seeds of Leguminosae, Cupressaceae,

Pinaceae, and Taxodiaceae with seedcoats entirely removed are inert matter.

b. In Abies, Larix, Libocedrus, Pinus

elliottii, l? echinata, P. rigida, P. taeda,

and Pseudotsuga, wings or wing fragments are detached and removed and

placed in the inert matter fraction.

Other pines retain wing fragments. (See

“ a” above). Normal dewinging and

cleaning should remove the wings in

these four Pinus species. Many more

species should probably be on this list.

Nursery workers want clean seeds.

c. For samaras, the wings are not removed

(e.g., Acer, Fraxinus, Cedrela, and

Swietenia).

d. For drupes, the fleshy coverings are not

removed.

e. In Eucalyptus species with small seeds,

the following simplified procedure is

used: pull out only other seeds [from 1.

b(2) above] and inert matter that is

obviously of nonseed origin. Pure seeds

will contain unfertilized and aborted

ovules. Germination of most species is

tested on weighed replicates.

f. For Leguminosae, if any portion of a

testa is present, it must be classified as

pure seed. Broken seeds must also be

larger than half the normal seed size.

For Dalbergia and other legumes that

may not be completely ‘extracted from

the pods, there are no instructions.

g. If species distinctions are impossible,

then only the genus name is given on the

certificate. This can happen with many

conifers.

F. Seed Weight

1. Determination-The ISTA (1985) rules

are followed to determine seed weight.

Either the whole working sample or replicates from it are used.

a. Working sample- The working sample

is the entire pure seed fraction of a

purity analysis carried out in accordance with ISTA (1985, chapter 3). The

working sample is put through a counting machine. Then the sample is

weighed in grams to the same number

of decimal points as in the purity analysis (ISTA 1985, rule 35.1).

b. Replicates-From the working sample, 8

replicates of 100 seeds each are counted

at random, by hand or with a mechanical counter. Each replicate is weighed in

grams to the same number of decimal

places as in the purity analysis (ISTA

1985, rule 3.51). The variance, standard deviation, and coefficient of variation are calculated as follows:

(1) Variance = n(Xx2) - (I&)2

n(n- 1)

Where n = number of replicates, x

= the sum of, and x = weight of

each replicate in grams.

71
(2) Standard deviation (a) = 

Coefficient of variation (CV)

s

= CT

i i

-

X 100 where X = mean weight

of 100 seeds.

(3) If the coefficient of variation does

not exceed 6.0 for chaffy grass seeds

or 4.0 for other seeds, the result of

the determination can be calculated.

If the coefficient of variation exceeds

whichever of these limits is appropriate, 8 more replicates are

counted, and the standard deviation

is calculated for the 16 replicates.

Any replicate is discarded that

diverges from the mean by more

than twice the standard deviation.

Two examples are:

Weight determination example 1

Lot 1 where:

2.50 n = 8

3.12 xx = 22.64

3.00 x = 2.83

2.78 ‘cx2 = 64.52

2.97 (r = 0.2530

2.42 CV = 0.2530

i I

100 = 8.9%

2.83

3.02

2.83

Weight determination example 2

Lot 2 where:

2.80 n = 8

2.78 23~ = 23.28

3.00 x = 2.91

2.94 xx2 = 67.80

2.97 u = 0.0866

3.01 cv = 0.0866 100 = 3.0%

i----i 2.91

2.88

2.90

2. &porting results-Results are reported

in one of two ways, either by a l,OOO-seed

weight or by seed per gram (or per kilogram, ounce, or pound).

a. l,OOO-seed weight

If counting is by machine, the weight of

1,000 seeds is calculated from the

weight of the whole working sample. If

counting is by replicate, from the 8 or

more weights of loo-seed replicates, the

average weight of 1,000 seeds is calculated (i.e., 10 x mean weight). The

result is expressed to the number of decimal places used in the determination

(ISTA 1985, rule 10.4).

b. Seeds per gram or per kilogram, ounce,

or pound.

Number per gram = 1,000

weight of 1,000

seeds in grams

Number per pound = 453,600

weight of 1,000

seeds in grams

Conversion is simple:

Number per gram = number per pound

x (0.002205)

Number per pound = number per gram

x (453.6)

Number per ounce = number per gram

x (28.35)

G. Sources

For additional information, see International

Seed Testing Association 1985, sect. 3, 3A, 10;

Willan 1985, p. 198-202, 221.

IX Germination Tests

A. Introduction

Good seed testing is the cornerstone of any seed

program, no matter what kind of seeds: agricultural, forestry, agroforestry, or ornamental.

The quality of the seeds used must be measured

and described. Seed testing may have legal ramifications because of its connection to seed sales.

For this reason, the International Seed Testing

Association (ISTA) coordinates international

efforts to standardize seed testing. The quality

of seeds must be known to make efficient and

effective use of them in reforestation or

afforestation programs.

B. Objectives

1. Identify the international organizations

that deal in tree seed testing and how they

derive their prescriptions.

2. Learn the principles of germination testing

and how they are applied in the laboratory

for standard conditions.

3. Practice actual germination testing in the

laboratory.

4. Learn proven techniques to analyze germination data and how these data can be

expressed.

5. Learn how to apply germination test results

to practical nursery and field conditions.

6. Learn how to make rapid estimates of seed

quality when time and/or proper facilities

are absent or limited.

C. Key Points

The following points are essential for conducting germination tests:

1. Laboratory germination tests are designed

to provide the optimum conditions for

germination and to determine the full

72
germination potential of the seeds under

these conditions.

2. The primary conditions to be considered are

temperature, light, aeration, and moisture.

3. Rapid estimates of germination are just

that - estimates; they are not as accurate as

germination tests.

4. If more than 60 days are required for a

germination test, analysts should use a

rapid estimate for official testing.

5. Germination testing in the course of

research may require different methods and

equipment from official testing.

6. No matter how standardized the test prescriptions are, the judgment of the analyst

must prevail in the laboratory. Almost every

test will produce some condition that is not

covered by the rules (ISTA 1985).

D. Definition of Terms

Relevant terms are derived from the glossary

developed by the Seed Problems Project Group

of the International Union of Forestry

Research Organizations (IUFRO) (Banner

1984a) and are defined as follows:

1. Abnormal seedlings-in seed testing,

seedlings that do not possess all normal

structures required for growth or show the

capacity for continued development

2. Filled seed- a seed with all tissues essential for germination

3. Germination - resumption of active

growth in an embryo, which results in its

emergence from the seed and development

of those structures essential to plant development

4. Germination capacity-proportion of a

seed sample that has germinated normally

in a specified test period, usually expressed

as a percentage (synonym: germination percentage)

5. Germination energy-proportion of germination that has occurred up to the time

of peak germination, the time of maximum

germination rate, or some preselected point,

usually 7 test days; the critical time of measurement can be chosen by several means

6. Germination percentage - (see germination capacity)

7 . Hard seeds- seeds that remain hard and

ungerminated at the end of a prescribed

test period because their impermeable seedcoats have prevented absorption of water

8. Peak germination- the specific time

when rate of germination is highest. It may

be derived in many ways (see germination

energy).

9. Pretreatment -any kind of treatment

applied to seeds to overcome dormancy and

hasten germination

10. Purity-proportion of clean, intact seeds

of the designated species in a seedlot, usually expressed as a percentage by weight

11. Sample, submitted-the sample of seeds

submitted to a seed-testing station

12. Sample, working- a reduced seed sample

taken from the submitted sample in the

laboratory, on which some test of seed quality is made

13. Seedlot -a specified quantity of seeds of

reasonably uniform quality

14. Seed quality-a general term that may

refer to the purity, germination capacity, or

vigor of a seedlot

15. Sound seed - a seed that contains in viable

condition all tissues necessary for germination (synonym: viable seed)

16. Tolerance -a permitted deviation (plus or

minus) from a standard. In seed testing,

the permitted difference between or among

replicated measurements beyond which the

measurements must be repeated.

17. Vigor-seed properties that determine the

potential for rapid, uniform emergence and

development of normal seedlings under a

wide range of field conditions

E. Quality Evaluation

For satisfactory evaluation of germination, the

following principles are fundamental:

1 . Sampling must be good; tests describe the

sample only. The seed manager must

ensure that the sample represents the lot.

2. Testing at standard, optimum conditions

ensures that:

a. Absolute maximum potential of the lot

is determined.

b. Standard conditions can be duplicated

by all laboratories for test comparison.

F. Methodology

1. Pure seed component- Only the pure

seed component is used (4 replications of

100 seeds each). If 400 seeds are not available, the number of seeds per replication is

reduced, not the number of replications.

2. Environmental conditions-Temperature, light, moistur,,= and medium must be

carefully controlled.

a. Temperature: Constant vs. alternating

temperature is a much debated point.

Some variation is allowed; thermogradient plate (TGP) results show broad

latitude in suitable temperatures.

b. Light: ISTA (1985) requires 750 to

73
1,250 lx (lux) (75 to 125 fc [footcandles]).

c. Medium-The germination medium

must be nontoxic; it can be either natural or synthetic.

(1) Natural materials are not used in

any type of germinator with water

circulation pumps. Materials include:

(a) Soil

(b> Sand

(c) Peat and other organic materials

(2) Synthetic materials include:

(a) Nontoxic paper products such as

blotters, paper towels, cellulose

wadding (Kimpaka), and filter

paper. (Some paper products

have mold problems.)

(b) Agar

(c) Cloth

3. Moisture-Excessive moisture is a common problem in many tests. Avoid having a

film of water around the seeds. Use the

“finger-press” test to judge moisture in

blotters. Make a depression with the tip of a

finger; if it fills with moisture, the blotter is

too wet.

4. Equipment-Reliable testing operations

must have dependable equipment.

a. Cabinet germinators - Cabinet germinators have good temperature control

and high capacity. Seeds can be placed

on moist blotters on open trays or in

small containers (see “d” below).

b. Jacobsen tables -The advantage of a

Jacobsen table is simplicity and moisture control; the disadvantage is poor

temperature control unless used in

climate-controlled rooms.

c. Walk-in rooms - Temperature control is

a problem; alternating temperatures are

almost impossible to establish.

d. Containers-Germination containers

include petri dishes (watch out for poor

aeration) and plastic boxes.

5. Test Procedures - There are many important considerations in proper germination

testing.

a. Pretreatment

(1) Micro-organism/pathogen treatment. A lo-percent sodium hypochlorite solution is a good, simple

treatment for external infection;

hydrogen peroxide (H,O,) is

another good surface sterilant (30

percent for 20 minutes).

(2) Overcoming dormancy (delayed germination)

(a) stratification (prechill): cold,

moist pretreatments are typical

for temperate species; warm,

moist treatments are also used.

In some species, paired tests are

prescribed (one with and one

without pretreatment).

(b) chemical treatment: nitrates,

H,O,, and growth regulators

are proven agents. Testing organizations disagree about the use

of chemicals in standard tests.

(c) scarification is common for hard

seeds, both by mechanical and

chemical means.

b. Placement of samples -Analysts need to

be careful in how samples are placed.

(1) Always leave spaces between seeds

on the medium.

(2) Rotate trays from bottom to top of

the cabinet in all germinators to

equalize light and temperature.

c. Counting- Certain guidelines are

needed.

(1) Be aware of definitions of a “germinated seed.”

(2) Conduct counts at least weekly. If

good germination rate data are

needed, three counts per week are

needed. Count daily if germination

is rapid (e.g., Populus, where 12-

hour counts are taken).

(3) Recognize abnormal seedlings.

(a) Typical or common abnormalities include albino seedlings, stunted roots, negative

geotropism (can be an artifact),

“endosperm” collars, and necrotic areas.

(b) Abnormalities also need to be

recorded during the test. Some,

such as those seen in Quercus

and Fraxinus, may be test

artifacts.

d. Length of the tests depends on rate of

germination. Most people use multiples

of weeks; 2, 3, and 4 weeks are the most

common. In official testing, analysts are

sometimes allowed to extend the test

period to allow complete germination.

This must be reported on the certificate,

and seed users should be alert for it in

dormant species.

e. At test’s end, cut ungerminated seeds to

determine empty, dormant (hard), or

74
rotten seeds. These counts help in quality evaluation.

6. Tolerance and retesting-Analysts

should:

a. Review the concepts for official testing.

For tolerance application, see table 23

[no equivalent table in Student Outline]

and the examples below:

ISTA procedure

Test 1 Test 2 Test 3 (retest)

80 78 81

83 80 90

88 95 84

90 98 87

-Xl = 85 r, = 88 x3 = 86

R, = 10 R, = 20 fofZand3 = 87

AOSA procedure

Test 1 Test 2 Test 3 (retest)

80 78 70

83 83 83

88 40 1 5

90 91 90

-21 = 85 X2 = 73 X, = 64

R, = 10 R, = 51 R, = 75

drop 40, drop 15,

-

x2 = 84 X3= 81

R = 13 R = 20;

retest lot

b. Analysts should be aware of other reasons for a retest:

(1) Too much dormancy; additional prechill needed.

(2) Too much fungal infection; increase

distance between seeds on blotter or

test in sand or soil.

(3) I’?‘?;rmal/abnormal distinction un-

(4) Evidence of human error.

G. Additional Testing Considerations

1. Thermogradient plates are good research

tools for screening many temperatures.

2. Greenhouse or nursery bed tests can be

useful.

3. Eucalyptus and Bet&a can be tested by

weight. See ISTA (1985).

4. Quercus and other large seeds can be cut in

half to save space and speed germination.

Table 23. --Maximum tolerated ranges between four replicates

(adapted from ISTA 1985) [no equivalent in Student

Outline]

Average percent Maximum Average percent Maximum

germination* range germination range

1 2 3 1 2 3

9 9 2 5 87-88 13-14 13

9 8 3 6 84-86 15-17 14

9 7 4 7 81-83 18-20 15

9 6 5 8 78-80 21-23 16

95 6 9 73-77 24-28 17

93-94 7-8 10 67-72 29-34 18

91-92 9-10 11 56-66 35-45 19

89-90 10-11 12 51-55 46-50 2 0

‘Calculate the average germination percentage to the nearest

whole number. Locate the average in column 1 or 2 and read the

maximum tolerated range opposite in column 3.

H. Reporting Results (figs. 33 to 36) [no equivalent

figures in Student Outline]

1. Germination capacity-should be expressed as a percentage of the total seeds in

the replication. If there are many empty

seeds, the posttest count will show this in

the test result.

2. Rate of germination- Germination

studies of many species suggest that rate of

germination is a good index of vigor.

a. Germination energy (the early count).

b. Mean germination time (MGT). Appears

to be very good when comparing treatments within lots.

c. Time for a certain proportion of germination to occur (e.g., number of days

for 50 percent or 75 percent of the seeds

to germinate).

d. Germination Value (GV).

e . Peak Value (PV>. In Pinus, PV has been

the best measure of rate of germination

(table 24) [no equivalent table in Student Outline].

I. Practical review

For suggested test conditions of tropical species,

see table 25 [no equivalent table in Student

Outline]. For practice on test results interpretation, see table 26 [no equivalent table in Student Outline].

J. Sources

For additional information, see Bonner 1984a,

1984b; Czabator 1962; Edwards 1987; International Seed Testing Association 1985, sect. 5,

5A, 11; Willan 1985, p. 202-227.

75
1108 Germination Test Sheet

Study 3.2 (11 Species Pinus taeda

Seeds\Rep. 50\2 Test Period 12-1-88 to 12-29-88

Temp. Regime 20 to 30 oc Treatment 7-vr test

Sample ID -18 OC - 10% B -18 OC - 10% C

I

DATE TEST DAY A B A B

12-12-88 11 3 4 1 3

12-14-88 13 3 5 5 9

12-16-88 15 2 4 1 8

12-19-88 18 12 14 9 10

12-26-88 25 19 13 22 10

12-28-88 27 0 1 1 0

12-29-88 28 0 0 0 0

NNS 39 41 39 40

Good, Ungerm. Seeds (ND) 0 0 0 0

Abnormal Germ. (NA) 0 0 0 0

Total No. of Seeds (NT) 50 50 50 50

Empty Seeds (NE) 6 5 5 4

Rotten Seeds (WV) 5 4 6 6

+- 1

Figure 33.-A germination test sheet showing good germination results with a sample of Pinus taeda [no equivalent figure in Student

Outline].

'76
Study 3.2 Sample ID 18102.1

ICUT = Day trial terminated

DFG = Day of first germination

NNS = Number of normal germinations

NA = Number of abnormal germinations

ND = Number not germinated by cut off but not

empty and judged capable of germination

NNV = Number not germinated by cut off but not

empty and judged not capable of germination

NT = Total number of seeds in trial

NE = Number found empty at cut off

NG = Good (NT - NE)

PUG = (NNS + ND)100

PNG = (NNS/NT) 100

PNGA = ARCSIN(SQR(PNG/lOO))

PAG =(NA/NT)lOO '

PD = Earliest day of max ((NNS(I)/NT)/I)

28

11

39

0

0

5

44

0

44

88.63636

88.63636

70.29977

0

25

PV = ((NNS(PD)/NT)/PD)lOO 3.545455

MDG = ((NNS/NT)/ICUT)lOO 3.165584

GV = (JW WDG) 11.223440

RECONSTRUCTION OF DAY-NUMBER PAIRS

Day

(1)

11 3 3

12 2 5

13 1 6

14 1 7

15 1 8

16 4 12

17 4 16

18 4 20

19 3 23

20 3 26

21 3 29

22 2 31

23 3 34

24 2 36

25 3 39

Increment Cumulative No.

INNS (I) NNS (I)

Mean 18.435900

Mode 16.000000

Variance 17.094470

Std. Var. 4.134546

Figure 34. -Computer analysis of data from figure 33, replicate A of the - 18 “C, lo-percent sample. See figure 33 for some term

definitions. Also, PUG = ultimate germination (includes dormant seeds); PNG = percentage normal germination;

PNGA = arc sine transformation of PNG; PAG = percentage abnormal germination. For Pv MDG, and Gv see

Section IV under Evaluation Quality [no equivalent figure in Student Outline].

7 7
*PNG DAE = germination percentage when dead seeds, empty seeds, and

abnormal germinants are included.

+Mean of replication A and B.

1108 Germination Test Sheet

Study 2.2 Species Pinus taeda

Seeds\Rep. 100\2 Test Period 12-4-87 to l-l-88

Temp. Regime 20 to 30 oc

Sample ID

DATE TEST DAY 01-A 01-B 02-A 02-B

12-14-87

12-23-87 19 2 12 5 6

12-26-87 22 9 10 8 3

12-28-87 24 4 6 2 1

12-30-87 26 3 5 5 5

01-01-88 28 2 5 5 5

NNS 70 72 73 75

PNG DAE'

Good, Ungerm. Seeds (ND) 20 23 11 16

Abnormal Germ. (NA) 0 0 0 0

Total No. of Seeds (NT) 100 100 100 100

Empty Seeds (NE) 0 0 1 2

Rotten Seeds (NNV) 10 5 15 7

Figure 35. -Agermination test sheet showing moderate germination results with a sample of Pinus taeda [no equiualent figure in Student

Outline].

7 8
Study 2.2 Sample ID 1.1

ICUT = Day trial terminated

DFG = Day of first germination

NNS = Number of normal germinations

NA = Number of abnormal germinations

ND = Number not germinated by cut off but not

empty and judged capable of germination

NNV = Number not germinated by cut off but not

empty and judged not capable of germination

NT = Total number of seeds in trial

NE = Number found empty at cut off

NG = Good (NT - NE)

PUG = (NNS + ND)100

PNG = (NNS/NT)lOO

PNGA = ARCSIN(SQR(PNG/lOO))

PAG = (NA/NT)lOO

PD = Earliest day of max ((NNS(I)/NT)/I)

PV = ((NNS(PD)/NT)/PD)lOO

MDG = ((NNS/NT)/ICUT)lOO

GV = (W (MDG)

RECONSTRUCTION OF DAY-NUMBER PAIRS

DaV

(1)

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

Increment Cumulative No.

INNS (I) NNS (I)

1 1

5 6

4 10

7 17

6 23

9 32

9 41

9 50

1 51

1 52

3 55

3 58

3 61

2 63

2 65

2 67

1 68

1 69

1 70

Mean 16.728570

Mode 15.000000

Variance 18.693390

Std. Var. 4.323585

28

10

70

0

20

10

100

0

100

90

70

56.78914

0

17

2.941177

2.5

7.352941

Figure 36. -Computer analysis of data from figure 35. See figure 34 for definition of terms [no equivalent figure in Student

Outline].

7 9
Table 24.-Germination of Pinus taeda: comparing data based on

total seeds with those of filled seeds only [no equivalent

table in Student Outline]

Lot

number

1

2

3

4

5

Germination

capacity

Total Filled

46 6 7

55 74

38 6 4

45 74

58 78

Pv’ G-v+

Total Filled Total Filled

2.06 3.00 3.45 7.27

2.19 2.94 4.47 8.00

1.54 2.58 2.12 5.86

2.43 4.04 3.91 10.77

2.70 3.63 6.24 10.87

6 38 8 2 1.67 4.04 2.74 12.80

7 32 78 1.48 3.63 1.73 10.09

8 40 8 6 1.65 3.52 2.39 10.64

9 2 4 73 0.98 3.03 0.90 8.82

10 24 6 5 1.05 2.81 0.99 6.98

11 19 4 6 0.90 2.18 0.69 3.64

12 5 6 77 2.55 3.50 5.10 9.63

13 22 65 1.05 3.35 1.18 10.42

14 20 91 1.06 5.05 1.01 21.86

15 45 79 2.36 4.14 4.07 12.68

16 2 0 77 1.19 9.22 1.02 13.68

17 2 4 8 4 1.48 5.39 1.69 22.11

18 4 8 9 4 3.51 6.83 8.15 30.60

*Peak value.

+Germination value.

Table 25.-Suggested test prescriptions for selected species [no equivalent table in Student Outline]

Species Suggested pretreatment Medium+ Temperature Duration Comments

Acacia nilotica

Aesculus indica

Ailanthus altissima

Albizia procera

Azadirachta indica

Bombax ceiba

Casuarina equisetifolia

Cedrus deodara

Dalbergia sissoo

Eucalyptus camaldulensis

Juglans regia

Leucoena leucocephala

Melia azedarach

Morus alba

Pinus roxburghii

I? wallichiana

Populus euphratica

Prosopis cineraria

Prunus padus

Robinia pseudoacacia SC: M, A, HW B 20/30 14

SC: M, A, Hw; CW 24 hr

CW 48 hr, cut off % scar end

CW 24 hr, remove pericarp

SC: M, HW

CW 24 to 48 hr

none

CW 24 hr

PC: 21 days at 3 to 5 “C

CW 24 hr

none

PC: 30 to 120 days

SC: HW (80 “C, 2 min), M

CW 24 to 48 hr

none; CW 24 hr

none

none

none

SC: M; CW 24 hr

PC: 3 to 4 mo at 3 to 5 “C

S, B

S

P, B

B

B, S

B

p, B

P, B

B

P

S

B

S

P, B

B

B

P, B

B

S

“C Days

20/30* 21

20/30 21

20130 21

20/30 21

25 21

25 21

20/30 14

20 21

30; 20130 21

30 14

20/30 40

30 14

20130 28

20/30 28

20/30 28

20130 28

20/30 10

30 21

20130 28

Some sources may need prechilling

(I

5

Remove seeds from pods

Yf’est by weight (0.1 g per replicate)

5

5

5

PTetrazolium or excised embryo

preferred

D

*Pretreatment codes: A = acid scar&cation; CW = cold water SO& HW = hot water treatment; M = mechanical scarification; PC = pre&ill;

SC = scarify.

+Medium codes: B = germination blotters; P = filter paper; S = sand.

*Use 16 hr at 20 “C in dark and 8 hr at 30 “C in light.

BPrescription from ISTA rules (1985).
Table ~&-HOW to interpret test results from the testing laboratory [no equivalent table in Student

Outline]

Rotten/

Normal Dormant Empty dead Abnormal PV Evaluation

___________________________ percent __________________________ Percentfday

95 3 1 0 1 6.0 Good lot; sow.

70 12 15 0 3 4.0 Too empties; reclean. many

80 0 0 13 7 2.0 Old seed? damaged?

25 2 5 6 0 8 0.2 Bad lot; don’t sow.

85 2 3 7 3 7.0 Which of these two would be

85 10 1 4 0 2.0 best for early sowing in cold soil?

50 35 5 5 5 1.5 Repeat with prechill.

6 0 12 2 3 2 3 2.5 Too abnormals; processing many

damage? genetics?

70 1 1 21 7 4.0 Too dead; abnormal total many

suggests damage; recondition to

remove dead seeds if possible.

‘Peak value.

V. Rapid Tests: Cutting, Vit.al Stains,

Excised Embyro, and Hydrogen

Peroxide

A. Introduction

The standard for judging seed quality is always

a germination test under optimum conditions.

Under certain circumstances, however, germination tests are not possible, and so-called

“rapid tests” must be used to estimate seed

quality. When performed properly, rapid tests

can furnish valuable information to seed users,

analysts, and managers.

B. Objectives

1 . Learn the different types of rapid tests and

how to perform them.

2 . Recognize the limitations of each test and

when it should be used.

3 . Examine the interpretation of test results.

C. Key Points

The following points are essential to perform

rapid tests:

1 .

2.

3.

4.

The cutting test is the quickest and simplest and can be extremely useful with

fresh seeds.

Tests with vital stains can tell the analyst

more than just potential germination, but

interpretation is subjective.

X-ray radiography is the most expensive,

but not necessarily the best, of the rapid

tests. It is very effective for some situations.

Leachate conductivity is a new and promising method.

D. Use of Rapid Tests

Rapid tests are used when one of the following

conditions occurs:

1 .

2.

3.

4.

5.

60-day rnle of ISTA-If a germination

test requires more than 60 days to complete, a rapid test should be used.

User request-Sometimes the test customer chooses not to wait for germination

test results and wants immediate evaluation.

Limited seed supply-If the lot is too

valuable to sacrifice 400 or even 200 seeds, a

nondestructive rapid test may be used. This

practice is common for research or breeding

lots.

Dnring collection-A rapid test can be

used to check the quality of the collected

seeds and adjust plans if necessary (i.e., to

increase collections if seed quality appears

to be low).

Other test objectives- When other seedlot parameters are more important than

germination (e.g., extent of mechanical

damage and viability), rapid tests may be

useful.

E. Sampling

The same sampling principles and precautions

apply as in standard germination tests; proper

sampling is essential.

F. Test Methods

There are six rapid tests that have applications

with tree seeds.

81
1. Cutting

a. Technique: Seeds are cut in half lengthwise and all tissues are examined.

b. Evaluation: Good seeds are firm, with

no apparent decayed or insect-damaged

spots, and have good color (usually

white to greenish white or ivory colored).

c. Advantages

(1) The fastest and cheapest test

(2) Can be performed in the field to

check collections as they are made

(3) Surprisingly accurate for fresh

seeds

d. Disadvantages

(1) Difficult for small seeds

(2) Poor results with stored seeds of

some species

(3) Is a destructive test

2. Vital stains

a. Technique: Embryo and storage tissues

are exposed by cutting, then immersed

in staining solutions for up to 24 hours.

The location and intensity of staining

indicate viable or dead tissue.

b. Stain options:

(1) Tetrazolium chloride (1‘2;) (2, 3,

5triphenyltetrazolium chloride) is

the most widely used stain. Active

dehydrogenase enzymes form a red

insoluble dye (formazan) from the

colorless TZ solution; live tissue

stains red. Primary development

was by Lakon in Germany. The TZ

method is widely used in Western

Europe. Dr. Robert Moore promoted

TZ strongly in the United States,

including applications for tree seeds.

(2) Indigo carmine stains dead tissues

blue. This method is common in

Eastern Europe and was developed

in Russia by Professor Nelyubon.

(3) Selenium or tellurium salt tests

were developed in Japan by Dr.

Hasegawa. This method was the

first vital stain method with seeds,

but it is not used now because of

metal toxicity from the salts.

c. Evaluation (TZ only):

Sound tissue should stain carmine.

Weak living tissue allows more rapid

penetration of the salt and stains a

darker red. Tissue that has been injured

by freezing may develop a blurred,

bluish-red stain. Because dead tissues

are not respiring and cannot produce

formazan, they do not stain.

(1) “Topographic stain” analysis is the

most accurate, but it is the most

difficult to standardize (figs. 37

through 43) [no equivalent figures

in Student Outline].

(2) The ISTA (1985) prescribes TZ for

certain dormant species and supplies evaluation guides.

d. Advantages

(1) Fast -stains can be read within 24

hours

(2) Inexpensive

(3) Equipment needs are simple

e. Disadvantages

(1) Labor-intensive, time-consuming

preparation

(2) Difficult to obtain uniform penetration of the stain, especially in seeds

with hard seedcoats

(3) Difficult to interpret the stain intensity and distribution

(4) Requires practice and experience

(5) Destructive test

3. Excised embryo

a. Technique: Seeds are cut open, and the

embryos are carefully removed for

incubation in dishes. The excised

embryo test begins as the TZ test does,

but the embryo is completely removed.

The embryos are incubated in light at 18

to 20 “C for 10 to 14 days.

b. Evaluation

(1) Viable seeds are green or white,

with some growth.

(2) Nonviable seeds are dark or moldy,

with no growth.

(3) The ISTA (1985) prescribes this test

for some dormant species.

c. Advantages

(1) Simple equipment needs

(2) Actual seed performance is tested

(3) Easy to evaluate

d. Disadvantages

(1) Labor-intensive, time-consuming

preparation

(2) Requires practice for proper excision

(3) Slow (10 to 14 days)

(4) Destructive test

4. Hydrogen peroxide

a. Technique: Seedcoats are cut to expose

the radicle and incubated in l-percent

hydrogen peroxide (HzOz) in the dark

with alternating temperatures of 20 and

30 “C. Radicle growth is measured after

3 to 4 days, and the seeds are placed in

fresh hydrogen peroxide. Radicle growth

is measured again at 7 or 8 days. Devel82
SEEDCOAT

I

FEMALE

GAMETOPHY

COTYLEDON

HYPOCOTYL

RADICLE

‘TE

S

F&are 37.- Pinus: Good stain (lined area); good seed (adapted from Krugman and Jenkinson 1974)

[no equivalent figure in Student Outlinel.

-9mm

-0

SEEDCOAT

FEMALE

GAMETOPHYTE

COTYLEDONS

HYPOCOTYL

RADICLE

Figure 38. -Pinus: Cotyledons weakly stained (lined area); almost no radicle stain; probably nongerminable or perhaps poor TZpenetration (adapted from Krugman and Jenkinson 1974)

[no equivalent figure in Student Outline].

8 3
-9 mm

SEEDCOAT

FEMALE

I GAMETO PHYTE

COTYLEDONS

HYPOCOTYL

RADICLE

Figure 39.-Pinus: Radicle stain only (lined area); nongerminable (adapted from Krugman and

Jenkinson 1974) [no equivalent figure in Student Outline].

SEEDCOAT

ENDOSPERM

COTYLEDONS

HYPOCOTYL

RADICLE

Figure 40. -Leucaena: Complete stain (lined area); good seed (adapted from Whitesell 1974) [no equivalent figure in Student Outline].

84
SEEDCOAT

ENDOSPERM

COTYLEDONS

HYPOCOTYL

RADICLE

Figure 41.-Leucaena: Unstained cotyledon tips (blank area); should germinate

(adapted from Whitesell 1974) [no equivalent figure in Student Outline].

-7 mm

0

SEEDCOAT

ENDOSPERM

COTYLEDONS

HYPOCOTYL

RADICLE

Figure 42. - Leucaena: Less than one-third of the radicle tip is unstained (blank

area); should germinate (adapted from Whitesell 1974) [no equivalent

figure in Student Outline].

85
SEEDCOAT

ENDOSPERM

COTYLEDONS

HYPOCOTYL

RADICLE

Figure 43. -Leucaena: One cotyledon unstained and more than one-half of the

radicle unstained (blank areas); nongerminable (adapted from whitesell 1974) [no equivalent figure in Student Outline].

oped on barley, the test is used on many North

American conifers.

b. Evaluation: Based on radicle growth - 5-mm growth

and up is good; 0- to 5-mm growth is uncertain; 0

growth is not viable.

c. Advantages

(1) Inexpensive and requires no elaborate equipment

(2) Objective (partially)

(3) Simple preparation

d. Disadvantages

(1) Not practical for very small seeds

(2) Tested only on conifers so far among tree

seeds

(3) Destructive test

(4) Slow (7 to 8 days)

5. X-ray radiography

a. Technique: Intact seeds are exposed to soft x rays,

and the images captured on film are examined.

(More details are presented in the next section.)

b. Evaluation: Subjective. Enhancements are possible

with contrast agents.

c. Advantages

(1) Fast

(2) Provides a permanent image

(3) Nondestructive test unless contrast agents are

used

d. Disadvantages

(1) Equipment is expensive

(2) Extensive training is required

(3) Interpretation is subjective

6. Leachate conductivity

a. ‘Ibchnique: Seeds are leached in deionized water for

24 to 48 hours, then electrical conductivity of the

leachate is measured. (More details are given in the

next section.)

b. Evaluation: Relationship of conductivity to germination must be established for each species.

c. Advantages

(1) No expensive equipment required

(2) Fast and simple

(3) Objective

(4) Nondestructive test

d. Disadvantages

(1) Indirect measurement that requires testing to

establish the relationship of conductivity to germination.

(2) Still under development; some unknown factors

cause trouble.

8 6
G. Sources

For additional information, see International

Seed Testing Association 1985, annex to chap.

6, appendix B; Leadem 1984; Willan 1985, p.

221-226.

VI. Rapid Tests: X Rays and Leachate

Conductivity

A. Introduction

Like other rapid tests, x-ray radiography offers

a quick estimate of seed quality when there is

no time for a complete germination test. The

application of x-ray radiography in seed science

is one of the few technologies that originated

with tree seeds instead of agricultural seeds. It

has not yet fulfilled its early promise, but there

are many applications with seeds. Many rapid

estimates of seed quality have major drawbacks: high cost, subjective interpretations,

excessive time, etc. The leachate conductivity

method offers a test that meets all requirements: low cost; fast, objective measurements;

easy procedures; and nondestructive. Although

relatively new, it shows great promise.

B. Objectives

1 . Review x-ray theory and see how x rays can

be used in seed radiography.

2. Learn the principles of seed radiograph

interpretation.

3. Examine the physiological basis for leachate testing.

4. Learn the leachate methodology.

5. Recognize the advantages and the disadvantages of both techniques.

C. Key Points

The following points are essential to understanding x rays and leachate conductivity.

1. Many types of seed damage can be detected

by x-ray testing.

2. Embryo development can be measured precisely, but exact correlations with germination are not possible.

3. The use of contrast agents can increase the

amount of information obtained from radiographs; however, many of these agents kill

the seeds.

4. Many special radiographic techniques are

available, but most require equipment associated with medical x-ray technology.

5. As seeds deteriorate, cellular membranes

are damaged, allowing the leaching of many

substances from the seeds.

6. Many chemical groups can be detected, but

electrolytic activity is the easiest to measure.

7 . Good estimates of quality are possible with

many species, but germination tests are

still preferred as the standard measurement of seed quality.

8. The conductivity method is promising, but

more research is needed.

D. X Rays

1. Theories

a. X rays are electromagnetic energy of

very short wavelengths. X rays penetrate

materials that absorb or reflect light,

and are themselves absorbed by the target object. The amount of absorption

depends on thickness, density, and composition of the object and the wavelength of the x ray. (The shorter the

wavelength the greater the energy,

which means more penetration) (fig. 44)

[no equivalent figure in Student Outline].

b. Radiographs are pictures of the object

formed by the x rays that pass through

the object and strike a photographic

material (film) or fluorescent screen.

(1) Radiograph quality is defined by:

(a) Contrast-degree of difference

in optical density of two adjacent fields.

(b) Density-amount of blackening of the radiograph.

cc> Definition - sharpness of

image detail.

(2) Quality is controlled by:

(a) Kilovoltage (kV) -voltage

potential between cathode and

anode; increase kV to get

shorter wavelength to lower contrast.

(b) Milliamperage (mA) -current

applied to the cathode; increasing mA increases quantity of

electrons available for x rays; too

much mA results in overexposure.

(c) Exposure time (seconds)-

the period that the object is

exposed to x rays; increases density of the image (better to

increase mA); the product of

time and mA (n&seconds) is

constant for equal radiographic

effect.

(d) Focus-film-distance (FFD) -

distance between focal spot and

film surface; increasing FFD

decreases intensity of radiation

and density.

87
c

ANODE +

/

GLASS -A I

FILAMENT CURRENT LEADS

WINDOW

BACK-SCATTERED

RADIATION

Figure 44.-Diagramatic view of an x-ray apparatus (Simak 1980) Ino equivalent figure in Student Outline].

(e) Object-film-distance (OFD)

-distance between object and

film surface; in seed work,

objects are usually placed

directly on the film; short OFD

gives better quality to the radiograph.

2. Methods

a. Equipment -Several types of x-ray

equipment are available commercially,

from both Europe and the United

States. X-ray generation of 0 to 60 kV at

3 to 10 mA is sufficient.

b. Film-Several film choices are available:

(1) Conventional film is similar to photographic film. It requires wet development and different film speeds

are available.

(2) Polaroid film provides rapid process88
ing, but is more expensive and provides less detail than conventional

film. Polaroid images fade with

time.

(3) Radiographic paper can be developed faster than film, but special

processing is required. It shows less

detail than conventional film, and

images fade with time.

c. Contrast agents - Contrast agents are

used to increase density of certain seed

tissue images on the radiograph by

treating the seeds before exposure.

Water is the simplest agent; most others

kill the seeds. The targeted tissues

absorb the contrasting agents.

(1)

(2)

Aqueous agents -primarily solutions of heavy cation salts; e.g.,

barium chloride (BaCI,) and silver

nitrate (AgNO,); seeds are soaked

for 1 hour after full imbibition, and

salts impregnate dead or damaged

tissue, thus greatly increasing the

density of the tissue image on the

radiograph. Water can be an excellent contrast agent with some

seeds.

Vaporous agents - different

vapors can be used to penetrate

either live or dead tissue; for example, 2 to 4 hours of exposure to chloroform (CHCl,) (or other halogen

derivatives of alk anes) produce high

density at dead or damaged tissues.

d. Safety is important in seed radiography.

Energy levels used in seed work are normally very low, but radiation exposure

has a cumulative effect in cells. Equipment should have approved shielding,

which must be checked periodically.

Operators should wear monitoring

devices.

3. Special Techniques

a. Stereoradiography - With two radiographs of the same seed, one taken with

the seed shifted, a three-dimensional

view can be obtained with a stereoscope.

The ratio of object shift to FFD is generally 1:lO.

b. Tomography - Radiographic images are

taken of preselected planes within the

object. The plane serves as the focal

point; tube and film are above and below

at fxed heights and are moved simultaneously during exposure. Only the

target plane stays in focus. This method

is widely used in medicine.

4.

5.

c. Xeroradiography -Xeroradiography

combines radiography and xerography, a

technique with exceptional image resolution. Instead of x-ray film, the image

is captured on a photoreceptor plate

sensitized with a charged, selenium surface.

d. Other techniques- Other special techniques are available, but none has yet

found widespread use in seed radiography.

Application in seed testing-X rays

were first used on seeds in 1903 in Sweden.

Practical development for seed testing came

in the 1950’s in Sweden.

a. Currently x rays are used to test for:

(1)

(2)

(3)

Determining seed anatomy, including embryo presence, size, and

shape. This process is good for

empty seed counts before germination tests.

Determining insect damage, including the location and extent of

damage and the growth of insect

larvae.

Determining internal mechanical

damage, including seedcoat cracks

invisible to the naked eye.

b. X rays have limited usefulness in determining viability or other physiological

attributes; the standard germination

test is better.

Summary- Show sample radiographs, if

available, on a screen.

E. Leachate Conductivity

1. Major Points

a. Deterioration: As seeds deteriorate, substances can be leached in proportion to

the degree of deterioration. Measurements of these substances can be correlated with seed quality.

b. Measurable materials:

(1) sugars

(2) Amino acids

(3) Electrolytes (the easiest to measure,

both in terms of time and expense.)

(fig. 45) [no equivalent figure in

Student Outline]

2. Techniques

a. Multiple-seed analyzer: Instruments

made in the United States, the ASAand ASAC- meters, have the same

circuits for measurement, but the

ASAC- has other features.

(1) Advantages

(a) Fast

89
/’----- -__-_

--_-_

/’

/’ ,/ Pinus polustris

/’

/’

,’

.’

--I /

/’

I t I I i I I

5 IO 1 5 2 0 2 5 3 0 3 5 L

LEACHING PERIOD (HOURS)

Figure 45.-Release of electrolytes over time from seeds of Pinus

paiustris and l? taeda (adapted from Bonner and Vozzo

1986) [no equivalent figure in Student Outline].

(b) Receives input from individual

seeds

(c) Data are printed on paper tape

(ASA-610)

(d) The ASAC- has a microprocessor that calculates some

statistics, and the data can be

stored in computer files.

(2) Disadvantages

(a) Expensive KJS$6,500)

(b) Some equipment not reliable

(c) Conductivity/germination relationship not completely understood

(d) Manufacturer’s support not

adequate

b. Single probe techniques: Single probes

measure conductivity in bulk samples

with a simple conductivity cell.

(1) The ISTA handbook of vigor test

methods (Perry 1981) includes this

method for peas.

(2) Advantages

(a) Fast

(b) Equipment inexpensive and

common in general laboratories

(c) Completely objective

o3-

IIoY = 117.87 - 3,321.99 (IhiS-25)

.

.

.

.

0.

l l

.

.

.

.

. .

0 2 0 4 0 6 0 S O 100

ESTIMATED GERMINATION (HS - 25)

Figure 46. -Relationship of actual germination to estimated germination in Pinus palustris as determined with the

ASAC- Analyzer (adapted from Bonner and Vozzo

1986) [no equivalent figure in Student Outline].

0 2 0 4 0 6 0 S O 100

ESTIMATED GERMINATION (HS-45)

Figure 41. -Relationship of actual germination to estimated germination in Pinus taeda as determined with the

ASAC- Analyzer (adapted from Bonner and Vozzo

1986) [no equivalent figure in Student Outline].

90
(d) Accuracy (within 10 percent of

germination for some species)

equals or exceeds that of the

ASAC-

(3) Disadvantage-As with the multiple-seed analyzers, some influences are not yet understood.

c. Results with United States species.

(1) ASAC-- See table 27 [no equivalent table in Student Outline] and

figures 46 and 47 [no equivalent figures in Student Outline] for examples of correlations with germination.

(2) Single probe-See figures 48

through 53 [no equivalent figures in

Student Outline] for examples of

correlations with germination.

3. Current status -The single-probe

method is promising. More research is

needed, but this method can already be

used to group seeds into high, medium, or

low quality classes. It is also easy to calibrate.

F. Sources

For additional information on x rays, see Vozzo

1978, 1988; Willan 1985, p. 224-226. For more

information on leachate conductivity, see Bonner 1991a; Perry 1981, chapter 6.

VII. Vigor Tests

A. Introduction

Standard germination tests do not adequately

measure the ability of seeds to germinate and

produce normal seedlings under field conditions

because germination tests are conducted in the

laboratory under optimum conditions. Such

conditions are seldom encountered in the field,

so germination and emergence may be much

lower than in the laboratory. Therefore, a more

sensitive measurement of seed quality has been

sought by those concerned with the planting

quality of a seedlot. This measurement of seed

quality has been referred to as seed vigor. Seed

vigor tests add supplemental information about

the quality of seeds to information obtained

through other tests.

B. Objectives

1 . Learn the concept of seed vigor and realize

how it can help the seed users.

2. Identify the types of seed vigor tests and

which ones are most suitable for tree seeds.

C. Key Points

The following points are essential in conducting

vigor tests:

1 . Vigor is a seed quality that may or may not

be indicated by a standard germination test.

2 . Vigor is most important under adverse field

conditions, and it can also indicate the storage potential of a seedlot.

3. Vigor tests usually involve either direct or

indirect measurements.

4. For many tree seeds, rate of germination is

the best expression of vigor.

D. Definition of Terms

1. Vigor

a. Association of Official Seed Analysts:

“Those seed properties which determine

the potential for rapid, uniform emergence, and development of normal seedlings under a wide range of field

conditions” (AOSA 1983).

b. International Seed Testing Association:

“The sum of the properties which determine the potential level of activity and

performance of the seed or seedlot during germination and seedling emergence” (Perry 1981).

Table 27. -Correlations of conductivity data with laboratory germination and nursery emergence and calculated error

limits for six pine species (Bonner and Vozzo 1986) [no equivalent table in Student Outline]

Number Laboratory germination Nursery emergence

of Best histogram Correlation Error Best histogram Correlation

Species seedlots segment* coefficient limit segment coefficient

r Percent r

Pinus echinata 9 35 0.7477 4.8 . ..+ .

P. elliottii 24 4 0 0.7806 7.2 45 0.6974

I? palustris 14 25 0.9252 10.9 20 0.8145

I? strobus 14 2 5 0.8546 13.6 25 0.7086

I? taeda 2 4 4 5 0.9775 6.5 4 0 0.7122

P virginiana 11 3 0 0.5342 3.1 . . .

‘Refers to that portion of the distribution frequency of current that indicates a good seed for that particular species

(See Bonner and Vozzo 1986).

+No nursery tests with these species.

91
-Or

oO-

)-

.

I.-

30

.

0

. .

0

.

.

.

.

0

.

0

.

.

.

.

:

I I I

35 40 45

CONDUCTIVITY (I/vS g-‘)

Figure 48. -Relationship of germination to leachate conductivity for

Eleagnus angustifolia [no equivalent figure in Student

Outline].

100 -

so -

60 -

40 -

20 -

l

. .

.

.

.

.

.

.

75 125 175

CONDUCTIVITY (PS g-‘)

Figure 49. -Relationship of germination to leachate conductivity for

Picea glauca as measured with a bulked sample (Bonner

and Agmata-Paliwall992) [no equivalent figure in Student Outline].

201 I I I I I I

0 5 0 100 150 200 250 300 350

CONDUCTIVITY (MS g-‘1

PNG = 106.24 - 0.1623 CONDUCTIVITY

Figure 50. -Relationship of germination to leachate conductivity for Gleditsia triacanthos samples

that were given accelerated aging to produce varying levels of quality (Hooda and others,

in press) [no equivalent figure in Student Outline].

9 2
.

G = 102 - 0.17 CONDUCTIVITY

CONDUCTIVITY (vS g-‘)

Figure 51 .-Relationship of germination to leachate conductivity for

38 mixed seedlots of Picea rubens (Conner and AgmataPaliwal1992)[no equivalent figure in Student Outline].

oooO10 :

\

Ooo -

) -

I2 0

.0). . . . l .

G = 99 - 0.40 CONDUCTIVITY

R’. 0.53

.

I I I I

6 0 100 140 180

CONDUCTIVITY (v.S 9-l)

Figure 52. -Relationship of germination to leachate conductivity for

14 seedlots of Picea rubens that were given accelerated

aging to produce varying levels of quality (Banner and

Agmata-Paliwall992) [no equivalent figure in Student

Outline].

1.0 1.5 2.0 2.5

CONDUCTIVITY (LOG IO pS g-1)

Figure 53. -Relationship of germination to leachate conductivity for

seeds from a single mother tree of Picea rubens, which

were given accelerated aging (Banner and AgmataPaliwal1992) [no equivalent figure in Student Outline].

9 3
c . International Union of Forestry Research

Organizations: “Those seed properties

which determine the potential for rapid,

uniform emergence, and development of

normal seedlings under a wide range of

field conditions” (Bonner 1984a).

2. Seed quality- “A general term that may

refer to the purity, germination capacity, or

vigor of a seedlot” (Banner 1984a).

E. Seed Vigor Concepts

1. Physiological quality- Seedlots vary

tremendously in physiological quality. This

is exemplified by the different rates of germination within a seedlot, the variation in

the growth rates and sizes of seedlings produced, and the ability of some seeds to produce seedlings under adverse conditions

while others do not. The physiological quality of seeds is commonly called seed vigor.

2. Physiological maturity- Seeds reach

their maximum germination capacity and

vigor during the maturation process at

their maximum dry weight, or the “physiological maturity” stage. Once physiological maturity has been reached,

deterioration begins and continues until the

death of the seed. The process cannot be

stopped, but the rate of deterioration can be

controlled to some extent. Different seeds

decline in vigor at different rates.

3. Deterioration - Seed vigor declines more

rapidly than does the ability to germinate.

The first sign of deterioration is a loss of

vigor. Thus, a seed may germinate even

though some of its physiological functions . may have been impaired. The ability to produce seedlings under stress conditions and

the growth and yield of plants may be

affected as vigor declines. Vigor is thus a

more encompassing measurement of seed

quality than the standard germination test.

4. Strategy-The general strategy in determining seed vigor is to measure some

aspect of seed performance or condition

that reflects the stage of deterioration or

genetic deficiency. Developing a good test

for this strategy is not easy. A practical seed

vigor test should:

a. Be reproducible

b. Be easily interpreted

c. Indicate field performance potential

d. Take a reasonable length of time

e. Not require expensive equipment

f. Not require extensive training

F. Common Seed Vigor Tests

Vigor tests can be grouped into four categories:

1. Seedling growth and evaluation

a. Seedling vigor classification-Similar to

the standard germination test, except

that normal seedlings are further classified as strong or weak based on deficiencies of roots, shoots, or cotyledons that

are symptomatic of reduced quality. Not

commonly used with tree seeds, seedling

vigor classification is closely related to

incidence of abnormal seedlings.

b. Seedling growth rate-Similar to the

standard germination test, but at the

end of the germination period, seedling

growth is measured as either linear

growth or weight. This method has been

tested on tree seeds with mixed results.

2. Stress tests

a. Accelerated aging- Seed germination is

the criterion for evaluation.

(1) In this test, seeds are subjected to

40 to 45 “C and nearly 100 percent

relative humidity for various

periods up to 6 days.

(2) Developed at Mississippi State University on agricultural seeds, this

test is now being used for tree seeds.

b. Cold test-Seeds are placed in soil (high

moisture and low temperature

[lo “Cl) for a specified period, then

transferred to favorable temperature for

germination. Used primarily for corn, it

is probably the oldest vigor test used in

the United States.

c. Cool germination test -Used commonly

with cotton. Seeds are germinated at

nearly minimal temperature (15 “C) for 7

days, then seedling length is measured.

Normal cotton seedlings of 4 cm or longer

are considered high vigor seedlings.

d. Osmotic stresses - Osmotic stresses

have been used but less than the other

stress tests.

e. Methanol treatment-This chemical

stress test was studied in Indonesia on

Heuea seeds and in the United States on

several Pinus species. It is still being

developed.

3. Biochemical tests

a. Tetrazolium chloride (TZ) staining is

primarily used to rapidly estimate germination, but some analysts interpret it

for seed vigor also. Location, rather than

intensity of the stain, is most important. It is a subjective interpretation

that takes years of practice to master.

b. Adenosine triphosphate (ATP) activity

is a laboratory test that has occasionally

shown good correlation with seed vigor

94
but is not widely used. Some trials have

been run with conifers in the United

States.

c. Glutamic acid decarboxylase activity

(GADA) is an enzyme activity test that

measures carbon dioxide output. It was

tested with hardwoods in the United

States with poor results.

d. Oxygen uptake (respiration) is a laboratory test that requires respirometers;

results have generally been mixed with

tree seeds.

e. Leachate test-Deteriorating membranes allow many cellular substances

to leach out when seeds are soaked in

water. The amount of these substances

can be related to seed vigor; for example:

(1) Sugars, tested with Picea glauca in

Canada and Pinus in the United

States.

(2) Amino acids, tested with Pinus in

Canada.

(3) Electrolytes. This method is by far

the easiest. It is probably more useful as a rapid test for estimating

germination (see previous section).

4. Germination data-Another approach is

to use germination test data, although more

frequent counts than ISTA requires may be

needed to achieve the required sensitivity.

a. Modeling. Mathematical modeling of the

germination response allows quantitative comparisons of frequency distributions.

(1) Normal distribution-If there is no

dormancy, frequency distribution of

germination should be normal (bellshaped curve). If dormancy is present, the curve will be skewed right

(Janssen 1973).

(2) Polynomial regressions for curve fitting (Goodchild and Walker 1971).

(3) Logistic function (Schimpf and

others 1977).

(4) Probit transformation - Cumulative

germination percentages are transformed to probits and plotted

against the germination rate

instead of test period in days. This

calculation should give a straight

line if no dormancy is present. Some

good results have been obtained

with conifers (Campbell and Sorensen 1979).

(5) Weibull function -A three-parameter function that can quantitatively

describe curve shape and beginning

and ending of germination on a time

scale. Research has studied both

hardwood and conifer species (Bonner 1986b, Bonner and Dell 1976,

Rink and others 1979).

b. Germination rate.

(1) Early counts-As prescribed by

ISTA, the first (early> count can be

used as a vigor indicator.

(2) Percentiles - Time required to reach

50 percent, 75 percent, etc., of germination is calculated. Similar to

mean germination time (below).

(3) Mean germination time (MGT)-

Can be useful in some cases. However, slow germination because of dormancy may inflate the value by giving

equal weight to the very last se&

lings to emerge; in the nursery, these

seedlings may never germinate.

(4) Germination value (Gv> and peak

value (PV) (Czabator 1962):

GV = PV(MDG)

where, PV equals the largest quotient of cumulative germination at

day x, divided by x, and MDG equals

the mean daily germination, or final

germination percentage divided by

total test days. Germination value

combines elements of rate and completeness of germination. These

expressions, more than any other,

are used worldwide. Peak value is a

good germination-rate term to

express vigor in temperate species.

G. Recommendations For Tree Seeds

1 . Germination rate parameters are best overall. Peak value may be best, but others are

good also.

2. Seedling growth tests are good if facilities

are available.

3. Tetrazolium (TZ) staining can be good for

large, tropical seeds, if interpretations can

be standardized.

4. Accelerated aging shows promise, but general recommendations are not yet available

for tree seeds.

5. Leachate conductivity also shows promise

and deserves more attention. Its nondestructive nature and inexpensive equipment requirements are attractive.

H. Sources For additional information, see Association of Official Seed Analysts 1983; Blanche

and others 1988; Bonner 1986b; Perry 1981;

Willan 1985, chap. 9.

95

97
I. Insects

A. Introduction

Insects are one of the greatest destroyers of tree

fruits and seeds. They reduce both quality and

quantity of seeds and affect angiosperms and

gymnosperms equally. Damage is done through

all reproductive stages, from developing buds to

cleaned seeds in storage. Losses to seed insects

are huge, and much is yet to be learned about

their complete role in the reproductive cycle of

woody plants.

B. Objectives

1. Learn the orders of insects that cause the

most damage to tree seeds and the species

they attack.

2. Recognize the types of injury that insects

cause.

3. Learn some methods of insect control and

management.

C. Key Points

The following points are essential in protecting

seeds from insects:

1. Insects of the orders Hymenoptera, Diptera, Lepidoptera, Hemiptera, Coleoptera,

Homoptera, and Thysanoptera do the most

damage to flowers, fruits, and seeds of

woody plants.

2. Damage ranges from causing reproductive

structures to abort to causing loss of seeds

in storage.

3. General types of damage include:

a. Destroying the seeds only, Hymenoptera

(wasps).

b. Forming galls and mine scales, Diptera

(flies).

c. Free feeding, Lepidoptera (moths).

d. Consuming endosperm, Hemiptera

(true bugs).

e. Mining cone axes, Coleoptera (beetles).

f. Causing cone abortion, Homoptera

(aphids and others) and Thysanoptera

(thrips and others).

4. Control methods depend on identifying the

insect’s life cycle and the host-plant relationship.

5. Some methods for reducing damage are preventive measures, insecticides, natural biological control agents, and proper

management techniques.

D. Damage

1. General concepts

a. Insects reduce seed production by infesting buds, flowers, cones, and seeds. Bud

damage causes wilting, abnormal

growth, and abortion of flowers and

immature fruit. Damaged fruits may be

deformed, worm infested, riddled with

galleries, and susceptible to secondary

infection by pathogens.

b. The most damaging insects to flowers,

fruits, and seeds of most trees are

largely restricted to six orders: Lepidoptera (moths and butterflies), Diptera

(flies), Coleoptera (beetles), Hymenoptera (wasps), Hemiptera (true bugs), and

Thysanoptera (thrips).

2. Specific concepts

a. Coleoptera (beetles) are the most damaging group in arid and semiarid zones.

(1) Bruchidae (bruchid beetles) are the

most important by far for

Leguminosae.

(a) Amblycorus, Bruchidius, Caryedon, and others feed on Prosopis, ?Ermarindus, Acacia, Parkinsonia, Gleditsia, and Cordia.

(b) Eggs are deposited on developing fruits, and the larvae eat

seed tissues. One group emerges

at fruit maturity (trees and

shrubs). A second group infests

pods at later stages of development, emerges, and reinfests

the same seeds in storage (e.g.,

peas and beans, Fabaceae).

(c) Bruchid beetles are serious seed

predators of Acacia in arid

zones.

(2) Curculionidae (weevils) lay their

eggs on developing fruits.

(a) Conotrachelus are important

cone worms in Mexican Pinus

and other subtropical pines.

(b) Curculio and Conotrachelus are

the common acorn weevils in

Quercus.

(c) Thysanocnemis are important

seed weevils in Fraxinus.

(d) Nanophyes can destroy 60 percent of the seed crop of Terminatia ivorensis.

(e) Apion ghanaense destroy many

flowers and seeds of Triplochiton.

b. Lepidoptera (moths and butterflies) can

damage stored seeds.

(1) Pyralidae attack many tropical

legumes, primarily the developing

pods and seeds. This family also

contains the very damaging cone

worms of Pinus (Dioryctrial. Larvae

destroy seeds, cone scales, and cone

axes (60 percent loss).

9 8
(2) Melissopus and Valentinia are

moths whose larvae destroy developing fruits of Quercus.

(3) Agathiphaga can destroy more than

half the seeds in Agathis cones in

the western Pacific area.

(4) Gelechiidae is a family of small

moths that attack Juniper-us cones;

they are very destructive.

c. Hemiptera (true bugs) feed on seeds with

specialized sucking mouth parts. Saliva is

injected into seed tissue to digest it.

(1) Coreidae attack Erythrina seeds in

India and some Acacia species in

Africa. The bugs reportedly move to

trees when crops are sprayed. This

family also includes Leptoglossus,

the very damaging pine seedbug on

tropical and subtropical Pinus, and

Leptocentrus, which damages

flowers and fruits on teak.

(2) Pentatomidae, the stink bug family,

feed on seeds of Pinus.

d. Hymenoptera (wasps) includes many

beneficial insects, but also some that

feed on tree seeds.

(1) Torymidae includes a major conifer

seed pest, Megastigmus spp. Its larvae feed on Pinus, Abies, and

Pseudotsuga.

(2) Eurytomidae includes the genus

Bruchophagus, whose larvae feed

on Acacia seeds; most feed on

smaller legumes than Acacia

because Acacia pods are so tough.

e. Homoptera includes aphids, cicadas, and

scales. Scales damage some Mexican

pines, but they are not a major predator.

Some damage to cones and seeds of

Pinus pinaster is caused by aphids in

South Africa; in one instance, seed yield

was reduced by 72 percent.

f. Thysanoptera (thrips) cause damage,

but little is known about the important

predators. However, Gnophothrips

damage or kill female buds and flowers

of Pinus (Mexican and other subtropical

species).

E. Controlling Insects

Control measures must be guided by the species

and ecology of the insect to be controlled. The

first step is to identify the problem and its

source insect. Seeds with insects feeding internally are collected and placed in plastic bags to

allow the insects to mature. The resulting information on the life cycle and insect-host relationship determines control measures.

1. Prevention - The insect may be prevented

from reaching the seeds. For example, in

The Gambia, long sleeves are put over the

flowering branches to exclude pests in

Acacia seed orchards. Such measures are

feasible only in very valuable seed orchards.

2. Chemical control must be applied when

the insect is at a vulnerable point in its life

cycle. For example, if the insect spends

most of its life inside of, and protected by,

seed tissues, chemical sprays will not be

effective during that period.

a. Foliar sprays: l-percent azinphosmethyl

spray for cone worms and seedbugs was

once common in Southern United States

pine seed orchards, but environmental

concerns reduced this practice. Sprays

of nuvacron and endosulfan have controlled insect damage in teak seed

orchards.

b. Systemic poisons: Carbofuran applied

as granules in the soil (4.5 g per centimer of diameter at breast height

[d.b.h.]) gives good coqtrol of cone

worms, borers, and seedbugs, but it is

not effective against seed chalcids.

c. Light traps: Can be used in seed

orchards to reduce the population of

egg-laying adults.

d. Chemical traps: Baited with sex attractants, chemical traps may reduce breeding populations.

e. Carbon dioxide: Treatment with carbon

dioxide is a promising new measure to

kill larvae in seeds. Shipping seeds in a

carbon dioxide-rich atmosphere is also

promising.

3. Natural enemies-The target insect’s life

cycle and history should reveal its natural

enemies.

a. The bruchid beetles have natural parasitoids that attack the egg, larval, and

pupal stages. The parasitoids are mostly

from the insect order Hymenoptera.

Mass-rearing the parasitoids to control

the beetles may be possible if circumstances warrant.

b. In Hawaii, seed beetles have been successfully controlled biologically by introducing wasps to attack eggs, larvae, and

pupae. The seed beetles lay their eggs on

the outer surface of the pods and are,

therefore, easily controlled by wasp

predators. Larvae and pupae, however,

are more difficult to control. Adult seed

beetles have such short lives they are not

susceptible to parasitoids.

99
c. A species of mite (Pymotes) will also feed

on seed beetles. However, laboratory

results do not corroborate field data, so

this control method is in question.

4. Collection- Good seed collection is the

first step in minimizing losses incurred in

storage. Removing infested seeds during

cleaning is the second step. Water flotation

and density separation methods can be

extremely effective. If good seeds were collected, they must be stored properly to control insects. Both low temperature and low

moisture content will subdue insect infestations; however, it is much better to avoid

transporting the insects to storage initially.

‘&vo weeks at - 18 “C will kill most larvae.

Drying seeds at 40 to 42 “C will kill insects.

Heating seeds in warm water and fumigating with serafume, methyl bromide, or carbon bisulphide will also control insects.

Chemical treatments should be avoided,

however, because they can reduce viability if

applied improperly. High moisture content

makes recalcitrant seeds especially susceptible to damage.

F. Sources

For additional information, see Cibrian-Tovar

and others 1986, Johnson 1983, Schopmeyer

1974, Southgate 1983.

II. Pathogens

A. Introduction

Pathogenic organisms (fungi, bacteria, and

viruses) cause great economic losses. Not only

are seeds the victim of pathogens, but they also

are passive carriers (vectors) of pathogens that

may not directly affect the seeds but may

endanger other organisms. This fact is the basis

of plant quarantine regulations that include

seeds in the import and export restrictions on

plant material.

B. Objectives

1. Learn the major types of seed pathogens

and the typical damage that they cause.

2. Identify steps to decrease losses to seed

pathogens.

3. Review documented occurrence of microorganisms associated with tree seeds.

C. Key Points

The following points are essential to preventing

seed pathogens:

1. The major disease-causing organisms are

fungi, bacteria, and viruses.

2. All tree seeds carry micro-organisms, primarily on the surface of their seedcoats.

3. All seed micro-organisms are not pathogenic; some may even be beneficial.

4. Pathology of tree seeds has not been studied extensively; much work remains to be

done.

D. Types of Pathogens

1. Viruses

a. Viruses are known to cause seven kinds

of seed damage:

(1) abortion

(2) flower sterility

(3) seedcoat wrinkling

(4) shriveling

(5) chalky endosperm

(6) staining

(7) necrosis

b. In legumes, embryo-borne viruses

reduce seed viability. Seedcoats shrivel

and crack, shrinking the seeds. The

seeds usually survive, but they remain

small and, most significantly, transmit

the virus to other seeds. In addition, the

weakened state of the seeds allows secondary infection from other pathogens

to reduce viability.

c. In agricultural legumes, a higher incidence of triploidy is associated with

seed-borne viral infections. These triploid seeds germinate slower with less

overall vigor than diploid seeds, and

their seedlings usually do not survive.

d. There are economic complications in

viral infections of agricultural seeds not

important to tree seeds. For example,

many virus infections discolor the seedcoat. Discoloration does not always

mean poor seed viability, but it can

influence the market price of seeds.

e. Although viruses live in seeds for very

long periods (up to 30 years in bean

seeds), it is not known how long viralinfected seeds will retain viability. The

virus will usually outlive the seed.

2. Bacteria-Bacterial infections account for

four kinds of seed damage:

a. Seed abortion includes problems ranging from shriveled seeds to size reduction to seed formation interruption; in

each case, it will significantly reduce

seed yields. The primary cause is

Xanthomonas.

b. Seed rot usually begins in water-soaked

lesions on the seedcoat. In young seeds,

there is a general, overall rotting of

tissue that then decomposes into bacterial slime. In mature seeds, the infection is usually localized in the seedcoat.

100
3.

Latent symptoms are rapidly rotting

seedlings. Infested seeds have discolored

bacterial pendants attached. Seeds are

sometimes so infested that cotyledons of

surviving seedlings are covered by black

lesions. .X&zthomonasis aIs responsible

for seed rot and is sometimes accompanied by rot fungi, such as Colletotrichum.

c. Seed discoloration is caused by pathogenic bacteria invading seedcoat lesions.

The lesions usually begin as slight

depressions caused by Xanthomonas or

Pseudomonas. Common colors are usually reddish brown or yellow. In extreme

cases, the seed eventually rots.

d. Slime disease (known also as tundu disease or tannau disease in Asia) is widespread (from Denmark to New Zealand)

and of prime significance in agricultural

seeds. It causes initial rotting, then a

massive slime coat when wet, which

dries to cover the seed with a varnishlike substance. Some symptoms

also begin as slime and result in seed

abortion. All causative agents are not

clearly identified, but include several

Corynebacterium species.

Fungi are a serious threat to seed health

simply because of the great numbers of species known as seed pathogens. In addition

to being lethal infections, some fungi drastically reduce the quality of seeds even

though they do not kill them. There are

eight kinds of fungal diseases, and two or

more commonly combine to attack seeds.

a. Seed abortion is the result of infection

by smut fungi that infest host flower

parts and replace them with fungal

fruiting parts. Flowers and young seed

structures are particularly susceptible

to Fungi Imperfecti, but as the host

matures, it can often withstand the

pathogen. Other species infect but do

not damage the seeds.

b. Shrunken seeds typically have shriveled

seedcoats. Fungal infections of stems

and leaves may cause seed shrinkage,

thus reducing seed yield. Rust fungi are

an example of this problem. Certain

other fungi reduce the oil content in

sunflowers.

c. Seed rot during germination is commonly caused by Fusarium, which produces dry rot. A large genus causing

severe problems for many tree hosts is

Botrytis. Also of significance are

Ciboria, Sclerotinia, Phomopsis, Valsa,

and Gloeosporium. Cones of many conifers are particulary susceptible to

Schizophyllum. It is also important to

control rot in seed storage. Mucor usually comes from the soil and then penetrates seedcoat cracks.

d. Sclerotization and stromatization are

defined as the transforming of host

flowers and seeds into sclerotia or

stromata of invading fungi. Ciboria and

Phomopsis cause a commonly known

“popcorn” disease in many forest tree

seeds.

e. Seed necrosis refers to dead tissue

accumulation, usually attributed to

seed-rot fungi. However, the penetration

of the fungi is usually superficial, extending only into the epidermal tissue.

In legumes, however, necrosis will

extend into lesions of the cotyledons.

f. Seed discoloration is not the problem for

forest tree seeds that it is for agricu1tura.I seeds. ‘lJ-ee seeds are usually

not subject to standards of form and

color that many edible seeds must meet.

Still, tree seeds are discolored by lesions,

fungal coatings, and pigmentation. Not

all fungi identified as seed-discoloring

fungi will produce pigmentation on

seeds, even though the fungi have

infected the seeds.

g. Lowered germination capacity is a broad

category, and perhaps any pathogen

could he placed here. However, some species seem to affect germination specifically. Ustilago may be present in the

embryo but remains dormant until the

seed itself germinates. Some reports

attribute the cause to fungal-produced

toxins.

h. The metabolites of some infecting fungi

have adverse physiological effects in the

seed. These metabolites are sometimes

not harmful to the seeds but are harmful to the animals that eat the seeds.

E. Control Mechanisms

Seed pathogens can be controlled by reducing

infection and treating seeds in laboratories,

storage, and nurseries.

1. Infection reduction - Infections in

orchards can be reduced by:

a. Locating seed orchards in areas of low

infection risk.

b. Removing alternate host plants from

seed production areas or orchards.

c. Practicing good sanitation in orchards

101
and destroying infected trees, old fruits

and cones, etc.

d. Applying fungicides (cone rust in

Pinus).

e. Using good cone- and fruit-handling

procedures; e.g.; reducing cone storage

time and removing old cones. (Enormous amounts of infection can occur

during extraction.)

2. Seed treatment in laboratories

a. Surface sterilization with hydrogen peroxide (HzOe) (30 percent ‘for 20 minutes), sodium hypochlorite (NaOCl) (lopercent solution of commerical bleach),

or ethanol (CzHsOH) (75 percent is

somewhat effective).

b. Fungicides come in numerous types and

concentrations; length of treatment and

concentration must be tested before

large lots are treated.

c. A hot water soak such as 50 “C for 15

minutes is used on rice to kill both external and some internal pathogens.

This method has not been tested for

tree seeds, but longer and hotter treatments might be effective on hard-seeded

species.

3. Seed treatment in storage is common

for some agricultural seeds but not for forestry seeds. Many fumigants or liquid,

organic fungicides are likely to decrease

seed viability in storage. Dry treatments

with fungicidal dusts are recommended for

tree seeds if a fungicide is needed.

4. Seed treatment in nurseries

a. Damping-off requires treatment.

(1) Soil is fumigated before planting.

(2) Seeds are treated with fungicide.

Thiram is used in pines of the

Southern United States.

(3) Soil is drenched with fungicide in

container operations.

b. Seedling diseases can be controlled by

treating seeds with the systemic

fungicide Bayleton (800 mg/liter).

F. Micro-organisms Found on ?‘ree Seeds

See Anderson (1986a) to identify those organisms that have been documented on the species of interest.

G. Sources

For additional information, see Anderson

1986a, International Seed Testing Association

1966, Neergard 1977, Sutherland and others

1987.

102
103
I. Production Systems

A. Introduction

This course is not intended to cover all aspects

of nursery establishment and management.

However, a few nursery problems involve seeds

and seed management practices. The type of

nursery system, size of the nursery, and location are important for seeds. It was once

believed that all seedling production and planting in the Tropics had to be done in containers.

This is not true; in general, however, bare-root

production systems predominate in the ‘lkmperate Zones and container production systems

predominate in the Tropics.

B. Objectives

1. Recognize different nursery systems and

the conditions most favorable for each.

2. Learn the relationship of nursery systems

to national seed program management.

3. Review basic seed technology for sowing in

each system.

C. Key Points

The following points are essential in understanding seed basics for nurseries:

1. Bare-root systems are more common in

Temperate Zones; container systems are

more common in the Tropics.

~

2. Bare-root production is possible in the

‘Ikopics with some pines and for stump production of selected species.

3. In container systems, large seeds are usually sown directly in containers, whereas

small seeds are sown in germination beds or

trays and transplanted (pricked out).

4. Tray mobility is an advantage in caring for

and protecting young seedlings.

5. In small nurseries, seed treatments for germination are usually done by hand.

D. Core Material

1 . ‘&pe of Nursery

a. Bare-root systems

(1) Are suitable in countries with largescale planting programs.

(2) Can produce seedlings or stumps.

(3) Require same-day planting in tropical environments when seedlings

are lifted; thus, distribution of seedlings to many planting locations

may not be feasible.

(4) Used with Pinus caribaea in Venezuela and stump plantings of

Gmelina, Dalbergia sissoo, and

Cassia siamea.

b. Container production systems

(1) Are usually preferred in most tropical locations because

(a) They can be small, laborintensive operations.

(b) Containerized seedlings can

stand the harsh environments

during transport to the planting

sites.

(2) System options are:

(a) A large centralized nursery with

0.5 to 1.0 million seedling production

(b) Numerous small nurseries with

10,000 to 100,000 seedlings

each

(3) Containers can also be used for

“wildings,” natural seedlings that

are dug up and transplanted. Wildings are useful for species with

recalcitrant seeds that cannot be

collected and delivered alive to nurseries for planting (e.g., Shorea spp.

in Asia).

(4) Movement of a large number of

seedlings at a time is difficult

because of container bulk, but the

system is suitable for many small

nurseries in dispersed locations.

c. Seed program considerations

(1) If most production is in a large, centrally located nursery, seed cleaning

and storage should be located

nearby.

(2) If most production is in small, dispersed nurseries, cleaning and

short-term storage should be in a

regional center. Long-term storage

should still be at a “national” seed

center.

(3) In small nurseries, much seed collecting, extracting, and cleaning are

done locally. This centralization

decreases the need for mechanical

operations and favors the use of

labor-intensive manual methods.

(4) Localized collection forces the use of

local seed sources.

(5) When seedlings of tropical recalcitrants are grown, small local nurseries must be used to avoid losing

seed viability during long transport.

(6) In most successful reforestation

programs, a combination of approaches will probably evolve,

depending on species, transportation systems, personnel, and politics.

2. Bare-Root Production

a. Small seeds-For small seeds, such as

104
those of Pinus, mechanized sowing and

culture are typical.

b. Large seeds-For large seeds, such as

those of Quercus, Juglans, and Melia,

sowing by hand in furrows across the

beds is typical.

c. Covering

(1) Small seeds-Press into the soil surface and cover with a light mulch (2

to 3 mm>.

(2) Large seeds -Place on their sides,

press into the soil, and cover with 5

mm of soil (or not more than three

times the seed diameter). Add mulch

if moisture conditions require it.

Some species, such as Swietenia

macrophylla, should not be pressed

into the soil because pressing can

cause J-root formation.

3. Container Production

Seeds can be sown directly into containers

or sown into seedbeds or seed trays and

transplanted later (pricking out).

a. Sowing into containers

(1) Sowing into containers is good for

the root systems because damage or

distortion from transplanting is

avoided.

(2) Sowing directly into containers is

used for:

(a) Large seeds that can be handled

individually

(b) Seedlots with expected high germination

(3) Suggested sowing rates are shown

in table 28 [table 16 in Student Outline].

(4) “Doubles” are pricked out to fill in

blanks.

(5) Only one seedling should be grown

per container.

(6) Seed needs are calculated as follows.

If the following conditions exist: the

Table 28. -Suggested sowing rates for seedling production in containers (Napier and Robbins 1989) [table 16 in Student

Outline]

Expected germination

Percent

80

60-79

40-59

<40

Seeds per container

Number

1 or 2’

2

3

use seedbeds

‘Sow half the containers with one seed and half the containers (b) Small seeds are broadcast,

with two seeds. pressed lightly into the soil, and

goal is 4,000 seedlings, seeds per

kilogram are 3,500, germination is

70 percent, plant survival is 85 percent, and plantable seedlings are 80

percent, then 5,882 containers are

needed. (Four thousand seedlings

divided by 0.8 times 0.85 equals

5,882 containers). With 70-percent

germination, 2 seeds per container

are sown (5,882 containers times 2

seeds per container divided by 3,500

kg equals 3.4 kg of seeds needed).

b. Sowing into seedbeds or seed trays

(1) Sowing into seedbeds or seed trays

concentrates germination in small

areas, allowing better protection

and care.

(2) This method is used for:

(a) Seedlots with expected germination of less than 40 percent

cb) Seedlots with slow germination

(c> Species that produce several

seedlings from one seed unit

(d) Very small seeds that are hard

to handle

(e) Scarce or expensive seedlots

(3) Seed tray mobility can be an advantage because it allows trays to be

moved in and out of shade or protected from heavy rains. Trays can

be made cheaply from local materials.

(4) The steps in sowing into seedbeds or

seed trays are:

(a) Use a sand:topsoil mix of 1: 1 for

slow germinators and growers.

They need nutrients from the

soil for 4 to 8 weeks.

(b) Use pure sand for Pinus,

Eucalyptus, etc., that germinate

quickly and are pricked out

after only a few days.

(c> Press seeds into the soil and

barely cover with washed sand;

then mulch lightly with pine

needles, rice husks, and water.

(Washed sand will not crust

when it dries.)

(d) Monitor closely to maintain

proper moisture level.

(5) There are other considerations

when using seedbeds.

(a) Seedbeds must be well drained,

and the soil mix should be at

least half sand.

105
covered lightly with sand and

mulch.

(c) If rodents are a problem, the

beds are covered with wire

mesh.

(d) For very small seeds (e.g.,

Eucalyptus and AnthocephaZus), the seeds are mixed with

fine sand and shaken from a salt

shaker to distribute them

evenly in the beds.

(6) Rates for sowing in seedbeds:

(a) Desired densities are 2,000

seedlings per square meter for

seedbeds and 4,000 seedlings

D. Sources

per square meter for seed trays.

(b) If there are 100,000 seeds per

kilogram and if the expected

germination is 40 percent, then

1 g of seeds should produce 40

seedlings. In seedbeds, 2,000

divided by 40 equals 50 g of

seeds to be sown per square

meter of seedbed. This rate is

doubled (100 g> for sowing in

seed trays.

For additional information, see Lantz 1985,

Liege1 and Venator 1987, Napier and Robbins

1989, Willan 1985.

106
107
I. National Programs

A. Introduction

National seed programs are necessary to support national reforestation and afforestation

efforts by ensuring an adequate supply of highquality seeds of suitable species and sources.

Countries of the Association of Southeast Asian

Nations are losing over 1.2 million hectares of

forest lands annually to other uses. Deforestation of “officially” designated forest lands in

India has not been excessive since the 1950’s

(about 3 percent of the lands under the Forest

Department), but more than 10 times this area

of wastelands, small groves, etc., has been

denuded. Reforestation efforts on a national

scale are needed. Within the framework of

national programs, State or Provincial seed programs may also be needed. A national forestseed program can serve many functions.

B. Objectives

1 . Learn the general functions of a national

forest-seed program.

2. Examine possible administrative structures of a national program.

3. Examine an existing national program as a

case study.

C. Key Points

The following points are important in national

seed programs:

1 . The primary function of a national forestseed program is to ensure an adequate supply of suitable tree seeds.

2. National programs can serve many other

important functions.

3. National programs should serve the needs

of all tree planting: industrial wood plantations, watershed protection, social forestry

plantings, agroforestry, etc.

D. Tree-Planting Activities

Many purposes are served by a national forestseed program.

1 . Industrial wood products, such as lumber, pulp, veneer, and speciality products,

are made from softwoods or fast-growing

hardwoods.

2. F’uelwood and charcoal are made from

short-rotation hardwoods (e.g., Acacia,

Eucalyptus, and Prosopis). These are good,

multipurpose species for:

a. Village forests

b. Individual landowners

c. Commercial production in small-scale

plantations on nonarable land

3. Watershed protection, especially around

reservoirs, mine spoils, and dune stabilization areas, is essential.

4. Windbreaks or shelterbelts

5. Urban planting can be primarily ornamental, but can include planting for climate

modification.

6. Wildlife habitat and food plantings

occur in parks and game preserves.

7 . Agroforestry planting is done for fodder

production, nitrogen fixation, and fuelwood.

8. Social forestry includes planting on roadsides, canal banks, and other common public areas.

9. Conservation of genetic resources can

be achieved by ex situ planting in clone

banks or “gene archives.”

E. Scope Of The Program

1 . The population distribution must be known.

2. Physiographic characteristics of the country or State must be identified.

3. Available land area and ownership are

important.

4. Annual goals must be realistic; the first

year’s may be modest to allow controlled

growth and expansion into a comprehensive

program.

5. Seed storage goals may include 2-, 3-, or

5-year supplies. Regional seed banks may

be needed. There may be sales to other

countries.

6. Distribution of indigenous species and their

potential seed collection zones must be

mapped.

F. Species Choices

Choice of species to be included in the program

is a critical step.

1 . Indigenous species and land races may be

best and are usually favored.

2. Exotics are often discouraged, and caution

should be used with them; however, there is

a definite place for good exotics.

3. Seed source choices are extremely important. If possible, these decisions should be

based on results of provenance tests.

4. Plantings should follow the natural plant

succession where possible. Pioneer species

should be planted first, with mycorrhizae or

nitrogen-fling bacteria innoculation, site

preparation, fire control, and protection

from livestock. These plants will form

nurse trees for the second group of shadetolerant species. The type of planting will

dictate the approach.

G. Administrative Structure

Many government agencies or ministries may

be involved because the same seeds may be used

for several purposes (agroforestry, watershed

protection, etc.). In the United States, the U.S.

108
Department of Agriculture, Forest Service and

Soil Conservation Service; the US. Department

of the Interior, National Park Service and

Bureau of Land Management; and State and

local counterparts are involved.

1 . Forestry ministry levels - The administrative structure can be:

a. National

b. Provincial or State

c. Village or other local structures

2. Comprehensive natural resource

agencies may combine forestry and wildlife.

3. Agricultural agencies -Indian Council

of Agricultural Research (ICAR) in India is

an example for agroforestry.

4. Military departments-in the United

States, military bases manage their own

forests.

5. Division of responsibilities can be made

for the various functions that are required:

a. Overall planning must consider total

production, seed collection zones, potential seed crops, collection vs. purchase,

source of funds, etc.

,

b. Seed acquisition and distribution can be

concentrated at one central location or

dispersed to regional centers. One

strong national seed center for research,

testing, and long-term storage is

strongly recommended. Dispersed secondary centers are usually needed to collect and clean seeds. In large countries,

such as India or Brazil, regional centers

as well as subregional centers are necessary. Important elements of this function are:

(1) Collecting and cleaning of seeds.

Seed production areas and seed

orchards may influence the location

of this work.

(2) Testing should be required on all

lots.

(3) Storage characteristics must be recognized. Workers must know which

species can or cannot be stored and

thus plan accordingly.

(4) Certification is usually not feasible

early in a program, but it may come

later if needed. It should be well

planned in advance.

(5) Record keeping is an important

function that should be centralized.

(6) If there will be sales to other countries, national needs must be satisfied first.

(7) If sales are within countries, sales

policies must be established. Villages or individuals may or may not

be charged for seeds and seedlings;

seeds could be supplied at cost.

c. Seedling production can be a part of

national seed programs. It usually is

not, but much depends on the government organization and the size of the

country. There are several options:

(1) National or State nurseries

(2) Village nurseries

(3) Private nurseries owned by individuals. (In some countries, farmers

can grow tree seedlings as a cash

crop.)

(4) Commercial nurseries. (Government nurseries must not conflict

with free enterprise.)

d. Plantation care is not usually a function

of the seed program unless seed stands

are involved.

(1) Protection, primarily from animals,

fire, and people is usually needed.

(2) Survival and early growth should be

documented because early plantations can be used for seed source

evaluation and then be turned into

seedling seed orchards.

e. Research may or may not be within the

structure of the national program, but

the program must help set research

priorities.

(1) Some common seed problems

include maturity indices, pretreatment, storage, and testing. One ultimate goal is a seed manual for each

country, either a comprehensive

book like the USDA manual (Schopmeyer 1974) or a smaller manual

that gives only basic seed handling

and treatment information (tables

29 and 30) [no equivalent tables in

Student Outline].

(2) Species, site, and seed source evaluations are critical, but they usually

fall under the tree improvement

program.

H. Critical Steps and Decisions

1 . Planting goals must establish what, where,

and how much to plant.

2. Availability of seed supply

a. Is the crop potential for indigenous species sufficient?

b. Are there suitable commercial sources?

Many commercial operations deal pri109
Table 29. -Cone and seed production characteristics of conifers in British Columbia (adapted from Dobbs and others 1976) [no equivalent table in

Student Outline]

Species

Period Minimum filled

Cone between seed count for

bearing collectible Cone yield collection

age Cone crops per mature Cones per 0.5 sect. per cone

begins length Avg. l?awe tree he&liter Collection period

Abies amabilis

A. grandis

A. lasiocarpa

Chamaecyparis nootkatensis

Larix laricina

L. lyalli

L. occidentalis

Picea engelmanni

I? glauca

I! mariana

P sitchensis

Pinus albicaulis

I? contorta

P. flezilis

I? monticola

I? ponderosa

Pseudotsuga menziesii

Thuja plicata

Tsuga heterwphylla

T. mertensiana

Years cm

2 0 9-13

2 0 5-12

2 0 6-12

15 0.5-1.5

4 0 1.0-1.5

3 0 4-5

2 5 3-4

15 3-8

2 0 3-6

10 l-4

2 0 6-10

2 0 4-8

10 3-5

2 0 a20

10 lo-25

15 8-15

15 5-10

15 l-2

2 0 2-3

2 0 2-6

--- Years ---

2-3 ..:

5-6 3-8

3 2-5

2-4 t..

5-6 . . .

. . . . . .

5-6 . . .

5 2-10

6 2-12

4-5 2-6

3-4 2-5

. . . 3-5

3 2-4

3 2-4

4 3-7

3 2-5

5 2-10

2-3 l-4

3-4 2-8

. . . 1-5

hL. ________-_---- Number --------------

. . . 700

. . 700

. . . 850

. . . 130,000

. . .

. . . . . .

. . . 11,000

. . . 8,300

. . . 11,000

. . . . . .

0.5-1.0 4,700

. . . I..

0.5-1.0 8,300

. . . . . .

0.2-0.5 280

1.0-1.5 700

0.5-1.0 2,800

I.. 110,000

. . . 83,000

. . . . . .

. . . .

. . . 1..

. . . . . .

. 2

. . . . . .

. . .

6-8 4 0...

7-10 . . .

7-10 . . .

. . .

7-10 :::

. . . . .

. . . 2 0

. . . . . .

. . . 9 0

. . . 75

5-7 *..

. . .

3-4 :::

. . . . . .

Late Aug. to mid-Sept.

Late Aug. to mid-Sept.

Mid-Sept. to mid-Oct.

Aug. to Oct.

Aug. to Sept.

Aug. to Sept.

Aug. to Sept.

Mid-Aug. to Sept.

Mid-Aug. to Sept.

Sept.

Sept.

Aug. to Sept.

Oct. to Mar.

Late July, Aug. to Sept.

Late Aug. to early Sept.

Late Aug. to early Sept.

Mid-Aug. to early Sept.

Aug. to Sept.

Sept. to Oct.

Sept. to Oct.

‘No data given

Table 30.-Selected data on seed quality of some Colombian tree species (adapted from aujillo 1986)

[no equivalent table in Student Outline]

Species

Average

purity

Average Range of

Pure viable Average germination

seeds/kg seeds/kg germination start End

Cordia allidora

C. gerascanthus

Cupressus lusitanica

Delonix regia

Didimopanax monototonii

Erythina glauca

Hymenaea courbaril

Jacaranda copaia

Ochroma lagopus

Samanea saman

Tecoma spectabilis

Percent

92.1

89.2

97.7

98.9

95.9

100.0

99.5

84.5

98.7

96.0

95.8

35.0

64.0

143.0

2.3

2.9

2.2

2.6

65.5

137.0

5.3

120.0

-- Number

27,098

32,459

24,778

1,394

11,305

1,374

142

12,097

61,423

4,485

97,239

63.3

65.5

13.8

58.1

35.3

63.5

54.5

20.0

69.0

94.5

77.6

--_ Days -___

9 2 2

6 16

10 2 3

5 16

69 8 6

5 21

22 3 2

11 2 5

10 4 0

4 14

5 1 2

marily with exotics (See table 31 for a

list of commercial seed dealers) [no

equivalent table in Student Outline].

3. Collection crews

a. Equipment and transportation must be

on hand.

b. Training of all crews will be required.

c. Legal assistance, such as permission

from landowners, government agencies, or tribal administrators will be

needed.

4. Nursery administration requires:

a. A suitable site

b. Trained personnel

c. Adequate equipment for all operations

5. Collection goals are related to planting

goals, potential seed crops, and storage

potential of the seeds. (Will more than 1

year’s supply be collected?)

6. Seed centers-A national seed center and/

or State or regional seed centers should be

established.

110
Table 31. -International seed dealers [no equivalent table in Student

Outline

Aggarwal Nursery and Seed Stores International Paper Co.

Panditwari Seed Center

P.O. Prem Nagar Nacogodches, TX 75962

Dahra Dun 248007, U.P. U.S.A.

India Telephone: 409/569-1069

Roger Anderson, Ltd.

P.O. Box 51-325 Pakuranga

Auckland

New Zealand

Telex: NZ 21721 (Roger A.)

Lawyer Nursery, Inc.

950 Hwy 200 West

Plains, Montana 59859

U.S.A.

Telephone: 406/826-3881

Telex: 31-9547

Asmer Seeds

Asmer House

Ash Street

Leicester

United Kingdom

Telephone: (0533) 26733

Soren Levinsen

Post Box 86

Kollerod

Denmark

Barilli and Biagi

l-40.100

Bologna

Italy

Louisiana Forest Seed Co., Inc.

Rt. 2, Box 123

Lecompte, LA 71346

U.S.A.

Telephone: 318/443-5026

EFG (Nurseries), Ltd.

Maelor Nursery, Conery Lane

Bronington nr Whitchurch

Shropshire

United Kingdom

Telephone: 094873 301

Nova %ee Seed Co., Inc.

Rt. 2, Middle Musquodoboit

Nova Scotia BON IX0

Canada

Florsilva Ansaloni

C.P. 2100

40100 Bologna

Italy

Telephone: 516255218

FAX 51-6525-6857

The Old Farm Nurseries

H.den Ouden and Son b.v.

P.O. Box 1

2770 AA Boskoop

Netherlands

Telex: 39810

The Forest Research Centre

P.O. Box HG595

Highlands

Halale

Zimbabwe

D. Oriell-Seed Exporters, Unit 11

10 Golfview St.

Mt. Yokine

6060 W. Australia

A.J. Frost

7080 Borkop

Denmark

Pal Seed Traders

7-Araghar

Dehra Dun, UP.

India

Reid, Collins, & Assoc.

Vancouver, BC

The Inland and Foreign Trading Co. Canada

(PTE) Ltd.

P.O. Box No. 2098

Maxwell Road Post Office

Singapore 9040

Telephones: 27222711, 2721801, 2782193

Telex: RS25254 IFTCO

International Forest Seed Co.

P.O. Box 290

Odenville, AL 35120

U.S.A.

Telephone: 205/629-5749

Renz Nachf GmbH and Co.

D-7270 Nagold-Emmingen

Germany

F.W. Schumacher Co.

36 Spring Hill Road

Sandwich, MA 02563

U.S.A.

Telephone: 617/888-0659

111
Table 31. -International seed dealers [no equivalent table in Student

Outline] (Continued)

Seed Branch G.J. Steingaesser and Company

Forestry Commission Research Station Postf. 1765

Alice H&t Lodge 8760 Miltenburg

Wrecclesham Germany

Farnham, Surrey

United Kingdom

Telephone: 0420 22255

Timmers and Leyer

P.O. Box 17

21OOAA Heemstede

Netherlands

lblex: 41754 flori nl

Seed Export Company

7 Bellata St.

The Gap

Queensland 4061

Australia

Telex: AA 42027

Setropa Ltd.

P.O. Box 203

1400 AE Bussum

Netherlands

Sheffield’s Seed Co., Inc.

273-Auburn Rd., Rt. 34

Locke, NY 13092

U.S.A.

Telephone: 3X/497-1058

Silvaseed Company

P.O. Box 118

Roy, WA 98580

U.S.A.

Southern Seed Company, Inc.

Baldwin, GA 30511

U.S.A.

Telephone: 4041778-4852

Telex: 4611041

Southpine, Inc.

P.O. Box 7404

Birmingham, AL 35253

U.S.A.

Telephone: 2051879-1099

Telex: 703040

I. Other Considerations

1. Continuity of operations-Well-trained

people should be kept at the same jobs for 3

to 5 years, preferably longer.

2. Ykaining of all personnel is a key to success.

3. Multiple functions sometimes occur.

a. In Morocco, government foresters also

grow and distribute fruit trees to the

farmers.

b. Some countries have only one seed laboratory so the same people must test

agricultural and tree seeds in the same

laboratory.

Tree Seeds

The Boat House, Potters Lane

Samlesbury, Preston

Lancashire

United Kingdom

Telephone: 077477 213

University of Hawaii at Manoa

Dept. of Agronomy & Soil Science

190 East-West Rd.

Honolulu, HI 96822

U.S.A.

Telephone: 808/948-7530

Van Dijk and Company

Einhuizen

Netherlands

Versepuy

43000 Le Puy

France

Vilmorin-Andrieux

La Menitre

49250 Beaufort-en-Vallee

France

Yamato-Ya, Ltd.

16-6, 1-Chome

Ginza, Chuo-ku

Tokyo

Japan

4. International organizations-The following organizations can help seed programs.

a . ISTA - ISTA Secretariat

Reckenholz, P. 0. Box 412

CH-8046 Zurich

Switzerland

b. IUFRO-IUFRO Secretariat

Schonbrunn

A-1131 Vienna

Austria

c. FAO-Forest Resources Development

Branch

Forest Resources Division

Forestry Dept., FAO

Via delle Terme di Caracalla

I-00100 Rome, Italy

112
d. ICRAF-International Council for

Research in Agroforestry

P.O. Box 30677

Nairobi, Kenya

5. Agricultural seeds- The relationship of

agricultural seeds to tree seeds must be

kept in perspective. Except for conifer seed

extraction, dewinging, and x rays, all the

tree seed technology is borrowed from agriculture: cleaning, storing, testing, etc. Seed

maturation and general biochemistry are

similar, and much can be learned from agricultural seed technologists.

J. Case Study

See Robbins and Shrestha (in press) for an

example from Nepal.

K. Summary

The functions of a national seed center are to:

1 . Further develop taxonomy and aids to species identification.

2. Collect and disseminate data on the ecology

of individual species, thus enhancing understanding of the performance of species.

3. Promote measures, as necessary, to conserve

the genetic resources of important species.

4. Develop optimum seed collection strategies

based on knowledge of breeding systems.

5. Maintain existing seed collections and

ensure their future development as programs evolve to use promising species and

provenances.

6. Assist collectors from other countries

within the framework of national policy;

some countries (e.g., Brazil and Indonesia)

restrict collections by foreign nationals.

7. Provide information on the physical and

physiological characteristics of seeds, and

any diseases that might be borne by seeds.

8. Encourage quarantine practices that minimize the chances of domestic insects

becoming established in other countries.

9. Disseminate information by providing

appropriate training, symposia, and publications.

10. Disseminate seed samples for research or

species trials to other institutions or countries on a cost or exchange basis.

L. Sources

For additional information see Gregg 1983,

Hellum (in press), Robbins and Shrestha (in

press), Rudolf 1974.

II. Seed Centers

A. Introduction

National forest-seed programs require some

sort of national tree-seed center, institute,

or laboratory. Dedicated facilities and some centralized authority are suggested for tree-seed

centers. Their level of technology may vary with

the country’s needs, but these centers should

serve as the focal point or hub of seed activities.

B. Objectives

1. Learn the general functions of national

tree-seed centers and how they support

national seed programs.

2. Examine several options for center design.

C. Key Points

The following points are essential to designing

and operating successful seed centers:

1 . The primary function of a seed center is to

support the national forest-seed program.

2. Seed centers provide seed services, research

on seed problems, training of seed workers,

and extension activities for seed users.

3. Many countries will require regional or subcenters for efficient operation.

D. Functions

The functions of seed centers include seed services, seed research, and training and extension

programs.

1 . Services -A seed center:

a. Coordinates seed collection by establishing seed zones, setting collection quotas,

and training crews.

b. Conditions seed collections by extracting, cleaning, and upgrading.

(1) All operations may be centrally

located.

(2) Drying and extracting can be at

regional centers, and seeds shipped

to a national center for final cleaning and storage.

c. Stores the seeds for the following purposes:

(1) Operational storage for nearby

users

(2) Long-term storage of surplus stocks

(3) Very long-term storage for germplasm conservation

d. Tests the seeds of:

(1) National seed program collections

(2) Other in-country users (e.g., universities) that could generate income

(3) Third parties (to settle legal disputes) that could also generate

income

e. Assists in certification-If a program is

established, seed center staff must be

involved, although primary responsibility rests with the “designated certification authority,” not usually the seed

center staff.

113
2. Seed research

a. Applied research on problems that

hinder efficient and economical seedling

production is vital; e.g., scarification

techniques, maturity indices, and treatment of seed pathogens.

b. Basic research is better done in cooperation with universities, but a basic

research group in the seed center would

have advantages; e.g., studying critical

drying rates for recalcitrant seeds, models of storage potential, and fatty acid

metabolism during storage.

3. lfraining and extension programs

a. Centers should train seed collectors,

analysts, and other personnel for specific programs.

b. Extension programs for nursery

workers and farmers to teach efficient

seed utilization are vital to national seed

programs and should be staffed by

trained extension workers. The training

of such people is beyond the scope of this

course.

E. National or Regional Centers

The decision to establish a national vs. a

regional center depends on the following considerations:

1 . National centers can be more responsive

to political realities and capitalize on

national pride.

2. Regional centers can expand scope and

function by pooling resources, an attractive

feature to donors. Regional centers may be

necessary because of the high cost of transporting large collections or because of the

use of short-lived recalcitrant seeds that

would not survive long trips in stressful

environments.

31 Compromise-National centers can be

used for storage, testing, and research;

regional centers can be used for collecting

and cleaning.

F. Location Concerns

1 . Proximity to seeds-The centers should be

located near major seed production zones.

2. Transportation - Good transportation

links (roads, rail, and water) are necessary.

3. Isolation - Complete isolation from population centers makes it hard to recruit and

retain good people.

4. Technical help-Because university affliation helps in many ways, location near a

campus is desirable.

5. Disaster potential-Flood or earthquake

zones should be avoided for the safety of the

facility and the workers.

G. Center Design

Seed center designs depend on the following

factors.

1 . Activity zones include the following areas:

a. Loading dock

b. Drying area

c. Extraction equipment (There may be

problems with dust and trash.)

d. Cleaning equipment

e. Conditioning equipment

f Seed storage

g. Testing laboratory

h. Offices for records and supervision

i. Supply storeroom

2. Building design - See figures 54 and 55

[no equivalent figures in Student Outline]

for the floor plans of Seedlabs 2000 and

5000 by ISTA.

3. Equipment

a. Commercial sources are best, but much

can he made locally.

b. Spare part sources are crucial for commercial equipment.

c. Maintenance must be available, either

from equipment suppliers or from local

people. If local people are used, then

they must be trained.

d. Electrical supply must be dependable.

e. Seed centers should get the best equipment because the size of the operation

will demand it. Subcenters and village

operations can get by with simple equipment because of their small size.

4. Staffing- Supervisors should have

defined areas of work; see figure 56 [no

equivalent figure in Student Outline]. The

staff should be made up of the following:

a. The director coordinates seed supply

needs and distribution.

b. The collection supervisor directs collection teams and coordinates State/

Province collections.

c. An extraction/cleaning supervisor and

two to five technicians.

d. A testing supervisor and two analysts

(one for purity and one for germination).

Seasonal workers will also be needed.

e. A storage/shipping supervisor, one technician, and one clerk.

5. Training-All staff members should be

trained in their specialties by university

staff, special short courses, or on-the-job

training at an established center. If personnel change jobs, the new people must be

trained immediately. The skills of long-time

staff should be updated as new methods are

developed.

114
l---

possqe

(lyxtension 1

I

I

I I I ’ 1

I I I I’ I

1 prevailing wind direction

Figure 54. -Suggested floor plan for a small testing laboratory (adapted from van der Burg

and others 1983). Dimensions are in centimeters [no equivalent figure in Student

Outline].

I f I I I I I I

l..u i germination lab

,:-kcpos

-.

Figure 55. -Suggested floor plan for a large testing laboratory (adapted from van der Burg and others 1983). Dimensions are in centimeters [no

equivalent figure in Student Outline].

115
SEED COLLECllON SUPERVISOR SEED HANDLING SUPERVISOR SEED TESTlNG SUPERVISOR

DELINEATES SEED ZONES LOGS IN COLLECTIONS SAMPLES AND TESTS ALL

INCOMING LOTS

DIRECTS REGIONAL COLLECTIONS EXTRACTS AND CLEANS SEEDS

TRAINS COLLECTORS LABELS AND STORES LOTS RETESTS IF NECESSARY

COLLECTS SEEDS MAINTAINS INVENTORY RECORDS MAINTAINS TEST RECORDS

j cEc:F;:E:WITH cmPERATIVE ) js;;;~;;~~~rL%~~~Es 1 IP~~~~~~~~~ 1

Figure 56.-A suggested staff organization for a small seed bank [no equivaht figure in Student Outline].

H. Sources

For additional information, see van der Burg

and others 1983.

III. Labeling and Certification

A. Introduction

When forest reproductive materials (seeds,

seedlings, and vegetative propagules) are not

collected or grown by the user, that user should

have reasonable assurance of the identity and

quality of the material he is buying. Many seedlabeling laws require detailed labeling to assure

the buyer of the seeds’ identity, purity, viability,

and freedom from pests; i.e., the physiological

quality of the seedlot. Certification is more than

labeling required by seed laws; it is a statement

about the genetic quality and identity of the

seedlot.

B. Objectives

1 . Understand the purpose of certification.

2. Identify the general elements of a certification program.

3. Describe the four certification categories

used in the Organization for Economic

Cooperation and Development (OECD)

standards for international trade.

C. Key Points

The following points are essential to understanding labeling and certification of forest

reproductive materials:

1. Certification is the guarantee by an officially recognized organization that forest

reproductive materials of identified varieties have been grown, collected, processed,

and distributed in a manner to maintain

high quality and genetic identity.

2. A certification program requires a certification agency, a producer who wishes to sell

certified material, records of the breeding

program, certification standards, independent inspections, and certification labels.

3. The four certification categories used by

OECD are:

a. source-identified (yellow tag)

b. selected (green tag>

c. untested seed orchards (pink tag)

d. tested reproduction material (blue tag)

4. Certification usually requires inspections of

the production unit prior to pollination, a

crop inspection before harvest, inspections

during the collection-to-storage phases, and

inspections at the time of packaging materials for sale.

D. Certification

1. Definition- the guarantee of character

and quality of reproductive materials by an

officially recognized organization, usually

evidenced by a color-coded tag and a certificate containing such information as certification category, genuineness of species

and variety, year of collection, origin, purity, soundness, and germinative capacity.

2. Purpose-Certification is more than just

labeling. The purpose of certification is to

maintain and make available to the public

high-quality seeds and propagating materials of superior crop plant varieties. For

agricultural and woody plant material,

the word “certified” implies genetic

improvement.

116
3. International aspect -An international

scheme for certifying forest reproductive

material was developed by the Organization

of Economic Cooperation and Development

(OECD). Its features are now being incorporated into forestry certification schemes in

North America.

E. Definition of Terms

The following definitions are for terms used in

the OECD Scheme (Organization for Economic

Cooperation and Development 1974):

1. Forest reproductive material

a. Seeds: cones, fruits, and seeds intended

for the production of plants

b. Parts of plants: stem, leaf, and root

cuttings, scions and layers intended for

the reproduction of plants

c. Plants: plants raised by means of seeds

or parts of plants; also includes natural

regeneration

2. Clone-a genetically uniform assemblage

of individuals derived originally from a single individual by vegetative propagation,

such as by cuttings, divisions, grafts, layers,

or apomixis

3. Cultivar - an assemblage of cultivated

individuals, which is distinguished by any

characters (morphological, physiological,

cytological, chemical, or others) significant

for the purposes of agriculture, forestry, or

horticulture and which, when reproduced

(sexually or asexually), retains its distinguishing features

4. Provenance- the place in which any

stand of trees is growing; the stand may be

indigenous or nonindigenous. (This is the

location of the seed source.)

5. Origin- for indigenous stands of trees, the

origin is the place in which trees are growing; for nonindigenous stands, the origin is

the place from which the seeds or plants

were originally introduced

6. Designated authority- an organization

or institution designated by and responsible

to the government of a country participating in the OECD scheme for the purpose of implementing the rules of the

scheme on its behalf

F. General Elements of a Certification Program

1 . The designated authority sets standards for

certification and provides inspectors and

certification tags and must have legal

standing.

2. A producer wishing to sell certified material

applies to the designated authority for qualification.

3. The history of the material (provenance,

seed source, and breeding program) must

be supplied to the designated authority who

determines if the material meets certification standards for genetic quality and

identity.

4. If the reproductive material is eligible for

certification, production of the material for

sale is supervised by the designated authority through independent inspections at

intervals during the growing, collecting,

and processing of the material.

5. The material to be sold must meet certification standards for viability, purity, and, in

some programs, freedom from pests.

6. Certification labels are placed on each container of reproductive material at the time

of packaging, and certificates of variety

identity are given to the buyer.

7 . The producer or seller is justified in charging more for certified material because certification ensures known genetic quality,

provenance identity, viability, purity, and

freedom from pests.

8. In summary, certification requires supervision, independent inspection, labeling, and

record keeping while allowing the seller to

sell at a higher price ‘or at a competitive

advantage.

G. Standards for Certification

1 . Certification classes for forest reproductive material differ from classes for agricultural seeds. Forestry programs typically

use the following OECD standards for forest reproductive material that includes

“tested reproductive material,” which is

the equivalent of the “certified” category

for agricultural seeds, and three additional

classes of less rigid genetic control. The four

classes and their requirements are:

a. Source-identified reproductive material

(yellow tag) comes from stands within

an identified seed collection zone. It is

required that:

( 1) Seed source and/or provenance must

be defined and registered with the

designated authority. (See “seed collection zones” below.)

(2) Seeds must be collected, processed,

and stored under inspection by the

designated authority.

b. Selected reproductive material (green

tag> comes from phenotypically selected

stands and cultivars. These stands and

cultivars have not been tested for

genetic quality, but they must:

(1) Be isolated by distance from poor

stands.

117
(2) Show normal variation among trees

within a stand.

(3) Be large enough for adequate crosspollination.

(4) Be old enough and developed enough

to allow evaluation of phenotypes.

(5) Exhibit phenotypic superiority in

some desirable quality, such as volume, wood quality, form or growth

habit, resistance to disease, fodder

production, or fruit production.

c. Reproductive material from untested

seed orchards (pink tag) comes from

phenotypically selected parent trees in a

seed orchard or from the progenies of

such trees.

d. Tested reproductive material (blue tag)

must come from seed orchards, stands,

or cultivars whose genetic superiority in

at least one desirable quality has been

proven in tests approved by the designated authority. Superiority can only be

certified in terms of the environment

and the age of the test.

e. Forestry certification classes can be

applied to parts of plants (cuttings) as

well as seeds.

f. Agricultural seed classes are different:

‘breeder seeds and foundation seeds have

white tags; registered seeds have purple

tags; and certified seeds have blue tags.

2. Seed collection zones

a. Seed collection zones, or regions of

provenance, should be delimited by

administrative and geographic boundaries and, where applicable, by elevation

and other appropriate boundaries.

b. Maps showing boundaries and reference

numbers of seed collection zones should

be established and published by the designated authority.

c. Seed collection zones are necessary for

“source-identified reproductive material.” The specific location and site characteristics of seed sources should be

used for the other three subclasses.

3. Other requirements of certification

a. Basic information-The originator,

developer, owner, agent, or producer

must request certification and must provide the following information:

(1) Name of the variety

(2) Statement of the variety’s origin

and the breeding procedure used in

its development

(3) Detailed description of any morphological, physiological, and other

important characteristics that distinguish the variety

(4) Evidence ‘of performance, including

comparative yield data, insect and

disease resistance, and other factors

supporting the identity of the variety (not necessary for sourceidentified reproductive material)

(5) Statement on the suggested area of

adaptation and the purpose for

which the variety will be used,

including a description of the

regions where the variety has been

tested and is recommended to be

grob. Inspections may include:

(1) Initial field inspections when certification is first requested are conducted before pollination to check

pollen-dilution zone and to remove

phenotypically or genetically poor

parent material.

(2) Crop inspections come just before

harvest to estimate the amount of

material that will be certified.

(3) Inspections during collection, conditioning, and storage of material

determine that the genetic identities

and viability are being maintained.

(4) Inspection at the time of packaging

for sale to check for “off-types,” disease, insects, viability, purity, and

germinative capacity are conducted

before tags are attached.

c. Fees-In the United States, certification

is financed by fees producers pay to the

designated authority.

H. Other Documentation

1. Labels-Some countries or other political

entities require labels for commercial sales

with identity (species), purity, germination,

etc., on the labels. No certification is

implied.

2. Phytosanitary certificate- Phytosanitary certification is required by most

countries to stop the spread of insects and

pathogens. It certifies that the seeds have

been inspected and/or treated (fig. 57) [no

equivalent figure in Student Outline].

I. Sources

For additional information, see Bonner 1981a,

Organization for Economic Cooperation and

Development 1974, Rudolf 1974.

118
THIS IS A SAMPLE

NO phyt”,d”ltary cerr,fKale <A” be lll”ed ““,#I a” applG%mn I, wnplek-d (I 0 H ,531 tee rc”CfIC for addll‘ond OMtl tnformatlon

“N,TED STATES DEPARTMENT OF AQRICULTURE fOR OFFICIAL “SE ONLY

ANIMAL AND PLANT HEALTH INSPECTION SERVICE

PLANT PROTECTION AND OUARANTINE PLACE OF ,SS”E

PHYTOSANITARY CERTIFICATE

~,

NEW ORLEANS, LOUISIANA

NO.: FPC

TO: THE PLANT PROTECTION ORGANIZATION(S) OF 134236

DATE INSPECTED

I I

CERTIFICATION

~hk b b ce&fl that. the plants or plant producra described below have been inspected accordingto appropriate procedures and are considered to be free

from quarantine pests, and practically free from other injurious pests; and that they are considered to conform with the current phytosanitary

I

1

t

3

5

-eguJat.ionsoftheimportingCQWtry.

. DATE

. CHEMICAL (acre ingrr3dienu

. CONCENTRATION

Imsif4fEsTATloN AND/OR DISINFECTION TREATMENT

2. TREATMENT

4. DURATtON AND TEMPERATURE

6. ADDITIONAL iNFORMATION

DESCRIPTION OF THE CONSIGNMENT T .

I

NAMEANDAGORESSOFTHEU(~~R 6. OECLAREO NAME AND ADDRESS OF THE CONSIGNEE

9. NAME OF PRODUCE AND GUANTITV DECLARED

10 . SOTANICA‘. NAME OF PLANTS 1,. NUMBER AND DE!XRIPTlON OF PACKAGES

I

(2. D(STINGUISHING MARKS

13. PLACE OF ORIGIN 14. DECLARE0 MEANS OF CONVEYANCE 16. DECLARED POINT OF ENTRV

Any intentional false statement in this phytosanltary certificate or misrepresentation relative to this phytosanitary

Clertificate Is a violation of law, punishable by a fine of not more than $10,000, or Imprisonment of not more than 5 years,

01 r both. (18 U.S.C. ~1001)

ADDITIONAL DECLAfIATlON

16. DATE lS.!WED (1. NAME OF AUTifORKED OFFICER (Type W WntJ $6. SMNANRE OF AUMORUED OFFICER

I I N CI liability shall attach to the United States Department of Agriculture or to any oflicer or representative of the Department with respect to this

I certificate. I

PPQ FORM 577 Previousedihns may be used. PART I - SHIPPER’S ORIGINAL

(SEP 92)

Figure 57. -A phytosanitary certificate used in the United States [no equivalent figure in Student Outline].

119
IV; Germplasm Conservation

A. Introduction

Loss of forests around the world is widely

deplored for many reasons, some valid and some

not so valid. One consequence of deforestation is

the loss of valuable germplasm that could be

used in artificial regeneration and future breeding programs. The Food and Agriculture Organization of the United Nations (FAO) lists more

than 300 tree species or provenances as

endangered. Fortunately, there are steps that

can be taken to conserve this germplasm.

B. Objectives

1. Recognize the consequences of excessive

loss of germplasm of forest trees.

2. Learn the strategies available to conserve

germplasm.

C. Key Points

The following points are essential to understanding germplasm conservation:

1 . The ideal practice would be extensive in situ

preservation.

2. Ex situ conservation is widely practiced

already, but “passport data” on planted

material need to be maintained.

3. Seed storage can play a critical role in

germplasm conservation.

4. National programs of conservation should

be carefully planned and established.

D. Importance of the Problem

1. The deforestation to replanting ratio is

about 30: 1 worldwide, although some of the

deforested areas regenerate naturally.

2. Insects and diseases can eliminate certain

gene pools or sometimes entire species. For

example, Castanea dent&a, Ulmus americana, and Abies fraseri are almost gone in

North America.

3. Global climate changes could eventually

change distribution of species through

changing selection pressures. Some gene

pools could be lost, or at least, gene frequencies would change.

4. According to FAO statistics, more than 300

species or special provenances are endangered. Complete species loss is rare, but

some provenances could be easily lost.

E. Available Technologies for Conservation

The following strategies are options for

germplasm conservation:

1 . In situ conservation is good for tropical

hardwoods; 100 species can be found in 0.4

ha. Land use pressures and economics

make this option increasingly difficult,

however.

2. Ex situ conservation is widely used for

3.

fast-growing plantation species. “It is well

suited to international cooperative efforts

because seeds can be shipped to available

sites. Blocks of 10 ha on several sites are

recommended. Current recommendations

are to save 50 to 400 individuals per geographic race or provenance.

Conventional seed storage plays a supporting role for tree germplasm because

regeneration of seed supplies requires long

periods of time. Storage is becoming more

popular because of its low cost. Genetic

damage during storage is possible, but this

is largely unproven in tree seeds.

a .

b.

C .

d.

7% orthodox seeds can be stored for

long periods (longer than one rotation)

at subfreezing temperatures.

Suborthodox seeds can be stored the

same as true orthodox seeds but for

shorter periods because of high lipid

content (e.g., Cava) or thin seedcoats

(e.g., Populus).

Temperate recalcitrant seeds can be

stored up to 3 years at a high moisture

content and temperatures just above

freezing.

Tropical recalcitrant seeds can be stored

the same as temperate recalcitrant

seeds, but chilling damage occurs below

15 to 20 “C, and seeds live for only a few

months.

4. Cryogenic storage is storage in liquid

nitrogen at - 196 “C!. This method is potentially very useful, but more research is

needed. Limits for true orthodox seeds are

unknown. Cost is about the same as for

conventional storage for small seeds (e.g.,

Eucalyptus and some Pinus spp.), but too

high for larger seeds.

5. Storage of pollen has little potential,

except as micropropagation tissues. Haploid

tissue, which may be useful in breeding,

could be stored this way. Storage life is

shorter than for seeds.

6. Micropropagation tissues have potential

for long-term storage of suspension cultures. This concept is intriguing, but

research is in the early stages. It could

help most with endangered, tropical, recalcitrant species. Technology will soon be

available, but cost may be high.

F. Current Efforts

1. The FAO, Forest Resources Division,

through the Panel of Experts on Forest

Gene Resources, promotes national and

international efforts.

120
2. Supported by FAO, the International Board of Forest Research, an Australian organiza3.

4.

5.

6.

7 .

for Plant Genetic Resources (IBPGR) is an

autonomous scientific organization of 13

countries. The board has recently added

forest trees to its agenda.

The Central America and Mexico Coniferous Resource Cooperative (CAMCORE)

is directed from North Carolina State University in Raleigh. Members include forest

industries and governments; the Cooperative includes hardwoods in its efforts.

The Oxford Forestry Institute (OFI) (formerly, the Commonwealth Forestry Institute) in the United Kingdom is active in

seed source and tree improvement for tropical species.

Danish International Development Agency

(DANIDA) operates the Forest Seed Centre

in Humblaek, Denmark. This agency is

active in Asia.

The Centre Technique Forestier Tropical

(CTFT) is a French organization that is

very active in Africa but less so in Asia.

Commonwealth Scientific and Industrial

Research Organization (CSIRO), Division

tion, is active in Asia and eastern Africa.

8. Numerous countries’ seed banks concentrate on indigenous species. Current efforts

at forest tree germplasm conservation

include seed storage at several international

and national centers (table 32) [table 17 in

Student Outline]. In addition, many countries have national seed storage facilities

that serve their domestic needs. It is questionable how much of this seed storage is

for long-term preservation of germplasm;

most of it is designed for short-term storage

to establish provenance trials and ex situ

conservation stands.

G. Recommendations for Action

1 . In situ conservation efforts should increase,

especially in tropical forests, both moist and

dry. The extent of genetic variation in natural stands must be determined.

2. Assistance in international efforts for more

ex situ conservation planting, similar to

that provided by FAO and OFI, is needed.

3. Research should continue on modeling conventional seed storage of true orthodox and

Table 32.-Some major international seed storage centers’ [table 17 in Student Outline]

Center Country

Approximate

size of collection

Species sources

_____ Number _____

Reference

United States Forest

Tree Seed Center

National Seed

Storage Laboratory

Petawawa National

Forestry Institute

DANIDA Forest

Seed Centre

CSIRO Tree Seed

Centre

OF1 United

Oxford, UK Kingdom

Banco Latinoamericano

de Semillas Forestales

Banco de Semillas

COHDEFOR

United States

United States

Canada 118 2,130 Janas (1984)

Denmark 4 6 187 Anonymous (1985)

Australia

Costa Rica

Honduras

6 7 197 Karrfalt (1985)+

18 41 Bass (1985)+

900

t

153

4

4,000 Turnbull and Doran (in press)

* *

308 Anon. (1983)

46 Gustav0 (1985)+

‘Bonner, F.T. 1986. Unpublished report. On file with: USAID Science and Technology Office,

Washington, DC. [Number of pages unknown].

+Personal communication from center directors.

*Data not available.

121
suborthodox seeds to predict just how long

they can be stored. Increased efforts with

these species in cryogenic storage is also

needed.

4. More research effort to conserve species

with recalcitrant seeds is crucial. Highpriority areas are reproductive biology, conventional seed storage, and micropropagation techniques.

5. More seed banks must be established now

for forest species. Rangewide collections can

be stored as seed reserves for ex situ plantings and as cellular reserves for micropropagation. These will also provide data on

conventional seed storage.

V. Applied Research

A. Introduction

Many seed problems can be solved locally without sophisticated research equipment that is

costly to acquire and operate. Some investigations furnish answers without statistical treatment; others need statistical work to

demonstrate their reliability. Simple designs are

usually satisfactory in seed work, including

completely randomized treatments and factorials. The main requirements are curiosity

and dedication.

B. Objectives

1. Learn a few principles of simple research

studies.

2. Review case study examples of applied seed

research.

C. Key Points

The following points are essential to applied

seed research:

1. Problems can often, but not always, be

solved with simple tests and experiments.

2. Standard procedures are always used when

they are available; e.g., ISTA (1985) rules

for germination testing.

3. Treatments are always replicated with several seed sources or in different seed years.

4. The limitations of the procedures in use

must be recognized; e.g., electric seed moisture meters cannot be accurate to 0.1 percent.

D. General Considerations

1. Replication- The “standard” for seed

work is usually 4 replicates of 100 seeds

each (4 x loo), although in research, 50 (or

even 25) are acceptable. If seeds are limited,

the number in each replicate is decreased,

but not the number of replicates. If the

material is variable, five or six replicates

would be satisfactory. If possible, more than

one seed source should be used or more than

one seed year from the same trees. Half-sib

collections are also good for certain tests,

such as determining maturity indices.

2. Documentation - Complete records are

essential. Entries should be in ink in bound

notebooks, if possible. Carbon or photocopies stored in a different place are also

desirable. Computer files are extremely useful, but not essential. The key is to make a

complete record of all treatment details and

results.

3 . Statistics-Because this is not a statistics

course, experimental design will not be

taught. However, it is important to:

a. Design studies to allow statistical analysis.

b. Use simple designs whenever possible,

but be sure that they are valid.

c. Temper statistics with common sense

because statistical significance may not

always equate with practical significance.

4 . Publication - Publication of results is

usually desirable because it disseminates

knowledge to others who may face the same

problems. However, it should never be the

primary goal of research.

E. Case Studies

1. Maturity indices of fruits or seedsThe steps for this research are:

a. Use a minimum of five trees.

b. Sample over a reasonable period.

c. Collect 10 to 15 fruits per tree.

d. Take color photographs if possible.

e. Take the following measurements:

(1) Size (length and diameter)

(2) Weight (wet and dry; dried at 103 “C

for 15 to 24 hours)

(3) Moisture content

(4) Germination

(5) Chemical analyses (optional)

(6) Histochemistry (an alternative)

f. Plot means on a time scale. (See the

example in the “Seed Maturity” section.)

g. Repeat at least twice to cover three seed

crops.

2. Extracting and cleaning methodsThe steps for this research are:

a. Make pertinent comparisons

(1) Sun drying vs. shade drying

(2) Hand extraction vs. machine extraction

(3) Any mechanical action vs. hand

cleaning

I 122
(4) Seed size (size into three groups and

test germination)

(5) Dewinging vs. sowing winged seeds

b. Replicate each treatment 5 times; test

each replicate with 4 samples of 50 seeds

each.

c. Retests unusual results.

d. Use the “t” tests for two treatments

and complete randomization for more

than two treatments.

3. Pretreatment for germination - The

steps in pretreatment are:

a. Make pertinent comparisons (tests)

(1) Hand scarification vs. mechanical

scarification

(2) Hot vs. cold water soak

(3) Stratification (time and temperature parameters)

(4) Chemical stimulation

b. Use the same general directions as for

extracting and cleaning. Factorial

designs will be more common for treatment combinations (e.g., time vs. temperature and soak time vs. chemical

level).

4. Storage conditions-The steps for storage tests are:

a. Make pertinent comparisons (tests).

(1) Room temperature vs. refrigerated

conditions

(2) Different refrigeration temperatures

(3) Seed moisture levels

(4) Qpe of storage containers

b. Use large enough replicates to allow

sampling over time.

c. Test orthodox seeds at 0.5, 1.0, 2.0,3.0,

4.0, and 5.0 years, and test recalcitrant

seeds at 1,2,4, 8, 12, 18, and 24 months,

then every 6 months thereafter.

d. Use at least four replicates.

5. Testing for recalcitrance-The steps to

test for recalcitrance are:

a. Bring the seedlot to full imbibition.

b. Start drying with at least two rates

(slow and fast), either indoors or outdoors, shade or sun.

c. Take periodic samples for moisture content and germination. Collect samples

every 2 to 4 hours for the first 24 hours,

then double that time in the next 24

hours, and double again in the next 48

hours.

d. Maintain the drying range from full

imbibition to lo-percent moisture or

until death of the seeds.

e. Designate seeds that cannot be dried

below 20 percent as recalcitrant.

f. Repeat this test to confirm recalcitrance; never trust just one measurement cycle. Tests of additional seedlots

are desirable.

g. Check chilling injury at 0 to 5 “C by

exposing fully imbibed seeds to this

temperature for 24 hours.

h. Keep statistics in perspective. Realize

that they are not as important as common sense in interpretation of results.

123
124
125
Exercise 1- Seed Structure

Objective:

To learn basic seed structures and their function in important seed types.

Methods:

1. Presoak seed samples in tapwater at room temperature (or 27 “C) for 15 to 24 hours. The imbibition will

soften tissues and facilitate dissection.

2. Using a knife, clippers, or a single-edged razor blade and depending on type of seeds, carefully cut the seeds

in one of two different ways:

cross section (transverse)

ba: lengthwise (longitudinal)

Several cuts may be necessary to expose the embryo and other internal tissues.

3. Examine the tissues exposed by the cuts and label them on freehand sketches of the cut material. Determine

which tissues are for embryo protection, storage of food reserves, etc. Look for abnormal structures, insect

damage, etc.

4. On at least one seed of each species, try to remove the embryo without damage, sketch it, and label the parts.

Supplies:

Clippers (or knives), single-edged razor blades, dissecting needles, a small magnifying glass (or hand lens), pencil

and paper, and seed samples of five tree species.

Exercise 2 - Seed Crop Estimation

Objective:

To predict seed crops in advance of collection by estimating the number of:

1. Good seeds per fruit

2. Fruits per tree

Methods:

1. Good seeds per fruit

a. Choose a multiple-seed fruit and collect 15 fruits prior to maturity.

b. Cut fruits in half lengthwise and count good seeds visible on each half

c. Dry the fruit halves in an oven (40 to 50 “C) to extract seeds and to obtain actual counts of good seeds.

d. Calculate regression equations to predict total seeds from fruit cross-section counts.

2. Fruits per tree-Visit nearby trees and estimate fruit crops by:

a. Total count

b. One-fourth crown count

c. Sample branch count

d. Any other known ways

3. Combine results of both methods to estimate size of the seed crop.

Supplies:

Cone cutters or sharp blades, an oven, drying containers, and binoculars.

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Exercise 3a-Cone Drying and Seed Extraction (Central America)

Objective:

To learn how to calculate seed and fruit needs for a planting program.

Assnmptions:

Area to plant-2,000 hectares (ha) at 1,700 trees per hectare

Species - Pinus caribaea

1. All moisture contents are percentage of wet weight.

2. There are 800 closed cones per hectoliter (hectoliter) (40 kg).

3. Moisture content of closed cones is 40 percent.

4. Cones double in size when open.

5. Yield averages 400 grams (g> of pure seeds per he&liter of closed cones.

6. There are 68,200 seeds per kilogram (kg).

7 . Laboratory germination is 80 percent; 50 percent of the germinated seeds produce plantable seedlings.

8. Cones are put into the kiln when they reach 25percent moisture content.

9. Drying trays hold 0.5 hL of closed cones; each stack of eight drying trays holds 4.0 hL; eight stacks can fit in

the kiln at once.

10. It takes 12 hours to dry a full charge to the lo-percent cone moisture needed for the cones to open fully.

11. It takes 700 kilocalories (Kcal) to heat 1 hL of cones for 1 hour.

12. Fuel value for wood of Casuarina equisetifolia is 4,950 Kcal per kilogram; for I! caribaea cones, 4,500.

13. Open I? caribaea cones weigh 104 g per liter (L).

Questions:

1. How many cones must be collected to meet the planting goal?

2. How much total moisture must be lost in predrying (prior to entering kiln)?

3. How many drying stacks will be needed to predry everything at once?

4. How many kiln charges will be needed?

5. How long will it take to open all cones?

6. How much fuel will be needed with C. equisetifolia wood? with P. caribaea cones?

7 . Have enough cones been collected to heat the kiln?

Answers:

1 . 312.5 hL

seeds required = (1,700) 2,000 = 8,500,OOO; or

(0.8) (0.5) i i

8,500,OOO = 125 kg

68,200

0.4 kg pure seed/h];: 125 = 312.5

0.4

2. 2,500 kg of moisture (312.5) (8)

1 hL = 40 kg; 16 kg I-I,0 and 24 kg dry matter

At 25%, = 0.25 x

24+x

x=8 kg

Since there were originally 16 kg, 8 kg must be lost.

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3. 78+

312.5 = 78.12

4

4. 10

78.12 stacks = 9.8

8 stacks/charge

5 . 120 hr

(12 hr) (10 charges) = 120

6 . Casuarina equisetifolia = 540 kg;

Pinus caribaea = 600 kg

Casuarina equisetifolia:

(32 hL/charge) (12 hr/charge) = 384 hLhr

(384) (700 Kcal) = 268,800 KcNcharge

268,800 = (54 kg/charge) (10 charges) = 540 kg

4,950

Pinus caribaea:

268,800 = (60 kg/charge) (10 charges) = 600 kg

4,500

7 . Yes

Open cones weigh 10.4 kg/hL

At 2 x expansion, we have (312.5) (2) = 625 hL of open cones

(625) (10.4) = 6,500 kg

Exercise 3b-Cone Drying and Seed Extraction (India/Pakistan)

Assumptions:

Area to plant-2,000 ha at 1,700 trees per hectare

Species - Pinus roxburghii

1. All moisture contents are expressed as a percentage of wet weight.

2. There are 400 closed cones per hectoliter (hL) (40 kg).

3. Moisture content of closed cones is 40 percent.

4. Cones double in size when open.

Questions:

5. Yield- averages 1.2 kilograms (kg) pure seeds per hectoliter of closed cones.

6. There are 12,000 seeds per kilogram.

7 . Laboratory germination is 80 percent; 50 percent of the germinated seeds produce plantable seedlings.

8. Cones are put into the kiln when they reach 25-percent moisture content.

9. Drying trays hold 0.5 hL of closed cones; each stack of eight drying trays holds 4.0 hL; eight stacks can fit in

the kiln at once.

10. It takes 12 hours to dry a full charge to the lo-percent cone moisture needed for cones to open fully.

11. It takes 700 Kcal to heat 1 hL of cones for 1 hour.

12. Fuel value of wood of Casuarina equisetifolia is 4,950 Kcal per kilogram; for r! roxburghii cones, 4,500. Open

P. roxburghii cones weigh 110 g per liter (L).

1. How many cones must be collected to meet the planting goal?

2. How much total moisture must be lost in predrying (prior to entering kiln)?

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3. How many drying stacks will be needed to predry everything at once?

4. How many kiln charges will be needed?

5. How long will it take to open all cones?

6. How much fuel will be needed with C. equisetifolia wood? With I? roxburghii cones?

7. Have enough cones been collected to heat the kiln?

Answers:

1. 590 hL

seeds required = (1,700) 2,000 = 8,500,OOO; or

(0.8) (0.5) i i

or 8,500,OOO = 708 kg

12,000

1.2 kg pure seed&L: 708 = 590 hL

1.2

2. 4,720 kg of moisture (590) (8)

1 hL = 40 kg; 16 kg Ha0 and 24 kg dry matter

At 25%, x = 0.25

24+x

x=Skg

Since there were originally 16 kg, 8 kg must be lost.

3. 147+

590 = 147.5

4

4. 19

147.5 stacks = 18.4

8 stacks/charge

5. 228 hr

(12 hr) (19 charges) = 228

6. Casuarina equisetifolia = 1,026 kg;

Pinus roxburghii = 1,140 kg

Casuarina equisetifolia:

(32 hL/charge) (12 hr/charge) = 384 hL-hr

(384) (700 Kcal) = 268,800 K&/charge

268,800 = (54 kg/charge) (19 charges) = 1,026 kg

4,950

Pinus roxburghii:

268,800 = (60 kg/charge) (19 charges) = 1,140 kg

4,500

7. Yes

Open cones weigh 11.0 kg/hL

At 2 x expansion, we have (590) (2) = 1,180 hL of open cones

(1,180) (11.0) = 12,980 kg

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Exercise 4 - Storage Space Requirements

Objectives:

Once annual seed requirements are known, space requirements for cold storage must be calculated. A decision

must also be made as to how many years’ supply of seeds will be maintained as a safety margin: 1 year’s?

3 years’?

Assumptions:

1. You must grow 2 million Acacia nilotica and 3 million Pinus wallichiana seedlings each year.

2. A 3-year supply of seeds will be stored.

3. Number of seeds per kilogram (kg) is 7,000 for A. nilotica and 26,000 for l? wallichiana.

4. For every three seeds planted, only two will produce a plantable seedling.

5. The seeds will be stored in large plastic bottles that hold 10 kg each. The bottles are 80 centimeters (cm) tall

and 40 cm in diameter.

6. Ten percent of the cold storage space is in aisles, etc.

C ah&ate:

1. How many kilograms of each species are to be stored?

2. How many bottles and cubic meters of storage space will be required?

3. Repeat calculation 2 if storage is in boxes 40 by 40 by 40 cm. Each box will hold 6.5 kg of seeds.

4. What are the minimum cold storage dimensions needed to store the seeds in calculations 2 and 3 above?

Answers:

1. A. nilotica: (2,000,OOO) (3) t (0.67) = 8,955,224

8,955,224 + 7,000 = 1,279.3 kg

I? wallichiana: (3,000,OOO) (3) + (0.67) = 13,432,836

13,432,836 + 26,000 = 516.6 kg

2. A. nilotica: 128 bottles (127. 93)

Bottle volume = (0.1257) (0.8) = 0.10 ms,

Space needed = (0.4) (0.4) (0.8) (128) = 16.4 me

P. wallichiana: 52 bottles (51.7)

Space needed = 6.6 m3

3. A. nilotica: 197 boxes (196.8)

Space needed = (0.4) (0.4) (0.4) (197) = 12.6 ma

I? wallichiana: 80 boxes (79.5)

Space needed = 5.1 m3

But square boxes need air space. A lo-cm space on four sides increases space needed to 19.7 ms and 8.0 ms.

4. Bottles: (16.4 + 6.6). + 0.9 = 25.6 m3

Boxes: (12.6 + 5.1) + 0.9 = 19.7 ms; 30.8 m3 with spaces.

Exercise 5 - Sampling

Objective:

To learn the basic methods of sampling bulk lots and some special applications for tree seeds.

Methods:

1 . Mix each lot thoroughly either with a mechanical mixer or by hand. To do the latter, spread the seeds out on a

smooth surface and mix by scooping from side to side. Then pour back and forth between two containers.

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2. Determine the proper size of the submitted sample (twice the working sample).

3. Draw samples using the following equipment/methods:

a. Seed trier

b. Mechanical divider

c. Division

d. Extended hand

4. Weigh each sample to the nearest gram, place in a plastic bag, and label.

5. Save these bagged samples for later measurements of purity, weight, and moisture.

Supplies:

A seed trier, a mechanical divider, a spatula, a spoon, plastic bags (15 by 15 cm), marking pens, and laboratory

balances.

Exercise 6 -Moisture, Purity, and Weight

Objective:

To carry out the basic steps in measuring moisture, purity, and weight of a submitted sample.

Methods:

1. Moisture

a. Use submitted samples drawn in the sampling exercise.

b. Use a spoon or spatula to draw two subsamples of 4 to 5 g each. Place samples in drying cans.

Follow guidelines in Bonner (1981b).

c. Weigh to the nearest 0.01 g and dry in ovens for 17 hours at 103 “C.

d. Cool in desiccators and reweigh. If desiccators are not available, use rapid-weigh techniques to

obtain dry weight.

e. Calculate moisture as a percentage of wet weight:

percent moisture = wet wt. - dry wt. (100)

wet wt.

2. Purity

a. Reduce the remainder of the submitted sample to the proper working sample size. To determine the

proper size, take at least 2,500 seeds up to a maximum of 1,000 g. ‘Iable 19 [no equivalent table in

Student Outline] may be used to determine sample size also.

b. Weigh the working sample (see 3.5.1.A in ISTA 1985).

c. Divide the sample into the following components:

(1) Pure seeds

(2) Other seeds (other species)

(3) Inert matter (includes seed parts)

d. Weigh each component and express as a percentage of the working sample weight:

percent pure seed = wt. of pure seed (100)

wt. of entire sample

3. Weight

a. Use the pure seed component from the purity test.

b. Either weigh and count the entire pure seed component or use smaller replicates (the usual

method).

c. Replicate method:

(1) Randomly count out 8 replicates of 100 seeds each.

(2) Weigh each replicate to the same number of decimal places used in the purity determination.

(3) Obtain the mean weight of 100 seeds and multiply by 10 for l,OOO-seed weight.

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(4) Convert to pure seeds per kilogram as follows:

1,000,000 = seeds per kg

wt. of 1,000 seeds

d. In official testing, variation would be estimated as follows:

(1) Variance = n(Cx2) -(%J2

n(n-1)

(2) Standard deviation (a) = J&&&n&

(3) Coefficient of variation (CV) = f (100) f-1

(X = mean wt. of 100 seeds, see 3.c above)

(4) If CV is 4.0 or less, the answer in “3.~” above is acceptable. If CV is more than 4.0, take 8 more

replicates and repeat the process, using all 16 replicates in the calculations.

Supplies:

A spatula, a spoon, laboratory balances, an oven, drying cans, desiccators, forceps, and pencil and paper,

Exercise 7 - Calibration of Electric Moisture Meters

Objective:

To demonstrate a simple method for developing calibration charts for electric moisture meters. These methods

will work with any type of meter.

Methods:

1. Draw 4 to 5 kg of seeds from a bulk lot of the desired species; mix well.

2. Separate into 10 random samples of about 400 g each.

3. Adjust moisture in these samples to span the range of moisture content that will be encountered

(approximately 5 to 20 percent). Do this by drying several samples (vary drying conditions) and by

adding water to others (vary the amount of water).

4. Place each sample in a plastic bag and place the bag in a cooler for 1 week to allow complete moisture

equilibration.

5. After 1 week, remove the samples and let them come to room temperature (2 to 3 hours).

6. Take a meter reading on the driest lot according to the manufacturer’s instructions. Record the value

and immediately draw two 5-g subsamples for oven determinations of moisture content. Follow

previous instructions.

7 . Repeat step 6 with the other samples in the order of ascending moisture content.

8. Plot data on a graph: oven moisture percentage vs. meter reading. Use this curve to relate future

meter readings to actual moisture content for this particular species only.

9. For more accurate calibration, fit a regression curve (oven moisture percentage on meter readings) and

calculate values for a calibration table. More than 10 observations should be available for a regression,

so another 10 samples should be drawn for a repeat of the entire process.

10. This procedure must be done separately for each species to be tested.

Supplies:

An electric meter, a spoon, laboratory balances, an oven, weighing dishes, desiccators, plastic bags, and graph

paper.

132
Exercise 8 - Germination Tests

Objective:

To learn the basic steps of a germination test and to carry out simple tests on some important species. Because of

the length of this course, a full test may not be possible. By starting a few samples before talking about testing at

length, some germination should occur and be available for evaluation.

Methods:

1.

2 .

3.

4 .

5.

6 .

7 .

Presoak seed samples in tapwater at room temperature (27 “C) for 15 to 24 hours.

Divide samples into 2 replicates of 20 to 50 seeds each, depending on the species. If Eucalyptus spp.

seeds are used, weigh out two replicates according to ISTA (1985) rules.

Half the class will surface-sterilize their samples with a lo-percent chlorine bleach solution, and the

other half will not treat theirs. Treatment will consist of a 5- to lo-minute soak, followed by rinsing in

running tapwater.

Hard-seeded species (e.g., Acacia) will be scarified with a knife, file, or sandpaper on the radicle end as

determined in Exercise 1.

Place replicates in glass or plastic dishes on moist filter paper or other suitable media. Paper should be

moist, but not moist enough to leave “free water” in a depression made by mashing down on the paper

with a finger. Put dish covers on; if there are no covers, use plastic wrap.

Label all dishes and place them in a germinator or constant temperature room if available. If these

facilities are not available, place the dishes on a table under lights in the center of the room. If good

lights are not available, place the dishes near windows that allow good natural light.

Check dishes every day for moisture; add water if they dry out. Germination may become evident in

about 7 days. Record normal germination, abnormal germination, and evidence of insect or disease

problems.

Supplies:

A knife, small file, or sandpaper for scarification, chlorine bleach, germination blotters, dishes (10 per student), a

glass-marking pen, and laboratory balances. A germinator or constant temperature room is desirable but not

necessary.

Suggested species:

Pinus, Acacia or another legume, Eucalyptus, and two indigenous species.

Exercise 9 - Scarification

Objective:

To demonstrate the relative effectiveness of simple scarification techniques that can be used in seed testing.

Methods:

1. Count out 120 seeds of a hard-seeded species and divide them into 8 samples of 15 seeds each.

2. Scarify 2 replicates of 15 by each of the following procedures:

a. Rub hand files or similar abrasive devices across the seedcoat enough to cut a notch in the seed.

b. Use hand clippers, shears, or a knife to cut through the seedcoat along one side.

c. Sandpaper the seed enough to cut through the seedcoat on the radicle end.

d. The other two samples will be the untreated controls.

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3 . Place the scarified samples on moist blotters in dishes and cover as in the germination test. Place all

dishes in the germinator if there is space. If not, place them on a table with good lighting and leave

them for observation through the rest of the course.

4 . Periodically count the number of germinating seeds and the number of swollen seeds. This latter

condition confirms that water uptake has occurred, but something else may be blocking germination.

Report both conditions as a percentage of the total number of seeds in the test.

Supplies:

Four glass or plastic dishes, germination blotters, a hand file, clippers or shears, rough sandpaper, and marking

pens.

Exercise 10 -Rapid Test: Tetrazolium Staining

Objective:

To learn basic techniques of the tetrazolium (TZ) stain test for viability.

Methods:

1 . Draw two 50-seed samples from seeds that have been soaking in tapwater for 24 hours.

2 . Prepare a l-percent solution of TZ by dissolving 10 g of a TZ salt (chloride or bromide) in 1,000 mL of

distilled water (pH 6.5 to 7.0). If the pH of the water is outside this range, a buffered solution must be

prepared as follows:

a. Prepare two solutions:

(1) Solution 1 -Dissolve 9.078 g KH;PO, in 1,000 mL of water.

(2) Solution 2 -Dissolve 11.876 g NazHP04 a 2HsO in 1,000 mL of water.

b. Mix two parts of solution 1 with three parts of solution 2.

c . Dissolve 10 g of TZ salt in 1,000 ml; of the buffer solution to make a l-percent solution.

3 . Carefully cut open the imbibed seeds to fully expose the embryo. The embryo may be completely

removed, as in the excised embryo test.

4 . Completely immerse the embryos in TZ solution in dishes and incubate them in the dark at 30 “C for

15 to 24 hours (depending on the species and seed condition).

5 . For evaluation, decant the TZ solution, rinse seeds in water, and examine the embryos on a wet

surface. Moderate red staining generally indicates viable tissues, heavy red staining indicates damaged

tissues, and the absence of any staining indicates nonviable tissues. Stain interpretations may vary by

species. See ISTA (1985) for guidelines.

6 . Compare results with other rapid test results or germination test results.

Supplies:

Dissecting equipment, dishes, tetrazolium salt, buffers (if necessary), and a constant-temperature dark incubator.

Exercise ll-Rapid Tests: Cutting and Excised Embryo

Objective:

To learn techniques of the cutting test for viability estimation and of embryo removal for the embryo excision

test.

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Methods:

1. Cutting Test

a. Draw samples of 50 seeds from each of several seedlots and divide them into sublots of 25 seeds each.

b. Cut the seeds in half, using a transverse cut through the center of the seeds. Categorize seeds as either

viable, damaged by insects or disease, or empty. Average the results from the sublots.

2. Excised Embryo Test

a. Draw samples of 50 seeds from each of several seedlots that have been soaking in tapwater for 24 to 48

hours at room temperature and divide into sublots of 25 as before.

b. Using razor blades or scalpels, carefully cut through each seedcoat and endosperm (if present), and

expose the embryo.

c. Carefully “tease” the embryo out of the surrounding tissues with dissecting needles or other sharppointed instruments. Avoid damaging the embryo.

d. Carefully place the excised embryo on moist filter paper in a covered dish, such as a petri dish.

Maintain at 20 “C in light until an evaluation can be made (usually within 14 days).

e. Diseased or damaged embryos should not be placed in the dishes. Empty seeds should be categorized as

such and not replaced in the test.

f The working surface and all instruments should be disinfected to reduce mold infections with a 50-

percent ethanol solution. Instruments should be “dipped” between each dissection.

g. Embryos should be categorized within 14 days as follows:

(1) Viable

(a) germinating embryos

(b) embryos with one or more cotyledons exhibiting growth or greening

(c) embryos remaining firm, slightly enlarged, and either white or yellow according to species

(2) Nonviable

(a) embryos that rapidly develop severe mold, deteriorate, and decay

(b) degenerated embryos

(c) embryos exhibiting extreme brown or black discoloration, an off-gray color, or white watery

appearance

(d) seeds in which the embryo is dead, missing, or deformed

h. Compare your results with the cutting test results.

Supplies:

Single-edged razor blades, scalpels, dissecting needles, dishes, filter paper, and ethanol.

Exercise 12 - Seed Health Testing

Objective:

To learn basic techniques of seed health testing.

Background:

Health testing of seeds is important for three reasons:

1 . Seed-borne inoculum may cause diseases in the field.

2. Imported seedlots may introduce new diseases, so tests to meet quarantine regulations may be required.

3. Seed health testing may aid in seedling evaluation and help determine causes for poor germination or field

establishment. It supplements the germination test.

Seed Health refers primarily to the presence or absence of disease-causing organisms (e.g., fungi, bacteria,

and viruses) and animal pests (e.g., eelworms and insects). However, physiological conditions such as trace

element deficiency may be involved.

Incubation maintains seeds in an environment favorable to the development of pathogens or symptoms.

135
Pretreatment is any physical or chemical laboratory treatment of the working sample preceding incubation that is

done solely to facilitate testing.

neatment is any process, physical or chemical, to which a seedlot is submitted.

Sample:

1 . Entire submitted sample may be the working sample, depending on the test.

2. The working sample is normally 400 pure seeds or an equivalent weight.

3. Sampling rules are followed.

4. Replicates containing a specified number of seeds, if required, are taken at random for a subsample after

thorough mixing.

General directions:

1 . Use different methods of testing depending on factors such as pathogen or condition being investigated,

species of seeds, and purpose of test. See ISTA (1966, 1985).

2. Examine the working sample with or without incubation.

a. Examine without incubation. (This method provides no indication of the viability of the pathogen.)

(1) Examine the sample with a stereomicroscope for general evidence of diseases or pests.

(2) Examine imbibed seeds. Immerse the working sample to make fruiting bodies, symptoms, or pests

more easily visible and to encourage the release of spores. Examine with stereomicroscope after

imbibition.

(3) Examine organisms removed by washing. Immerse the working sample in water with a wetting

agent, or in alcohol, and shake to remove spores, hyphae, nematodes, etc. Examine the excess

liquid with a compound microscope.

b. Examine after incubation.

(1) After a specific period of incubation, examine the working sample. Note the presence of disease

organisms or pests on or in seeds or seedlings. Use blotters, sand, or agar for incubation media.

(2) Use blotters when required to grow the pathogens from the seeds or to examine the seedlings.

Seeds may or may not be pretreated. Space widely to avoid secondary spread of organisms. Use

light as necessary to stimulate sporulation. Examine with a microscope.

(3) Sand or artificial composts can be used for certain pathogens. Seeds are not usually pretreated,

but they are widely spaced on the medium. Incubation is favorable for symptom expression.

(4) Use agar plates to obtain identifiable growth of organisms from seeds.

(a) Sterility is required; seeds are normally pretreated and spaced.

(b) Identify characteristic colonies and spores by microscopy.

(c) Use lighting and germination inhibitors.

3. Examine growing plants. Grow plants from seeds and examine them for disease symptoms to determine

the presence of bacteria, fungi, or viruses. Use inoculum from the test seedlot to test for infection of

healthy seedlings.

Calculations and Expression of Results:

1 . Express results as a percentage of seeds affected or as number of organisms in the weight of sample

examined.

2. Report results on the ISTA certificate.

a. Report test method.

b. Report pretreatments.

c. Absence of health test does not imply satisfactory health condition.

Specific Test Example-Pitch Canker Fungus:

1 . Adapted from Anderson (1986b).

2. Blotter Method, 400-seed sample.

136
a. Pentachloronitrobenzene (PCNB) Broth

Combine peptone, 15 g; MgSO, 7H,O, 5 g; KHsPO,, 1 g; terraclor, 1 g with 1 L of distilled H,O. Stir

well using magnetic stirrer. Autoclave for 15 minutes. After autoclaving, place flask on magnetic stirrer

and stir slowly until solution cools to room temperature or slightly warmer. Add 1 g of streptomycin

sulfate and 1 to 2 g of neomycin sulfate under sterile conditions, and stir.

b. Place 25 seeds on blue blotter paper in plastic containers. Crush the seeds with a sterilized piece of

plastic cut to fit the plastic box opening. Spray seeds and blotter paper with PCNB broth.

c. Incubate 14 days at 20 “C or until colonies are 2 cm in diameter.

d. Inspect all seeds for slow-growing, granular white colonies. Check each suspected colony using a light

microscope at 100 to 400 magnification for microconidia and polyphialids. Select fungus from seed

surface, not the blotter surface. Split the seeds into 4 groups of 100 for reporting purposes.

3. Agar Method

a . Prepare fresh potato dextrose agar (PDA) (makes 1 L)

(1) Clean and dice one medium-sized potato.

(2) Put diced potato in beaker with 500 mL of distilled H,O. Run through autoclave.

(3) In flask, add 20 g of dextrose and 17 g of agar to 500 mL of distilled H,O.

(4) Put dextrose/agar solution on magnetic stirrer and low heat.

(5) Strain cooked potatoes through two layers of cheesecloth to obtain at least 200 mL of slurry.

(6) Note amount of slurry, and pour slurry in flask with dextrose/agar solution.

(7) Add enough distilled He0 to make total slurry solution amount to 500 mL (i.e., if there are 200

mL of slurry, add 300 mL of distilled H,O).

(8) Put solution in autoclave and run for 15 minutes. To acidify media, add 20 drops of 50 percent

lactic acid to obtain a pH of 4.7.

b. Isolating external seed fungi, 25-seed sample

(1) Place the whole seeds on acidified PDA (pH 4.7).

(2) Incubate 14 days at 20 “C.

(3) If possible, observe fungal growth daily and identify the fungi.

c. Isolating internal seed fungi, 25seed sample

(1) Surface sterilize the whole seeds in 70 percent ethanol for 10 minutes. Stir the seeds every 2

minutes.

(2) Under sterile conditions, cut each seed open and remove half the center material.

(3) Place the seed half (center material) on acidified PDA (pH 4.7) using sterile technique.

(4) Incubate 14 days at 20 “C.

(5) If possible, observe fungal growth daily and identify the fungi.

137

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May 22-26; Bangkok, Thailand. Bangkok: Kasetsart

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van der Burg, W.J.; Bekendam, J.; van Geffen, A. [and

others]. 1983. Project seed laboratory 2000-5000.

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Vozzo, J.A. 1988. Seed radiography. Materials Evaluation. 46: 1,450-1,455.

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Zobel, Bruce; ‘Iblbert, John. 1984. Applied forest tree

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144
-

145
Course name and number

SHORT COURSE EVALUATION FORM

PRECOURSE INFORMATION

1. Who discussed the general objectives of the course with you before you arrived at the course

site?

2 . How clear were the course objectives? (Please circle answer.)

1 2 3 4 5

Very clear Clear Somewhat Somewhat Unclear

clear unclear

In what ways did the course differ from what you expected?

4 . Please describe how you were selected for participation in this course.

ORIENTATION

How helpful was your initial orientation at the course location?

Not helpful 1 2 3 4 5 Extremely helpful

Did not attend

Comments on precourse information and orientation:

146
COURSE ADMINISTF?ATION/LOGISTlCS

Please indicate your satisfaction with the following support arrangements at the course location:

Not at ail Extremely

satisfied 1 2 3 4 5 satisfied

Housing accommodations

Training facilities

Transportation

1 2 3 4 5

1 2 3 4 5

1 2 3 4 5

Administrative and

logistic help 1 2 3 4 5

Arranged social activities 1 2 3 4 5

Please comment on the above arrangements:

147
FIELD TRIP ADMINISTRATION/LOGISTlCS

How adequate were the following field trip arrangements?

Poor 1 2 3 4 5 Excellent

Preparatory information 1 2 3 4 5

Transportation

Lodging

Helpfulness of instructors

accompanying you

Helpfulness of people

you met in the field

Overall coordination

1 2 3 4 5

1 2 3 4 5

1 2 3 4 5

1 2 3 4 5

1 2 3 4 5

Please comment on the above arrangements.

148
COURSE CONTENT

1. At the beginning of the course or during the course, did you discuss with the instructors how

the course content would meet your specific needs?

-Yes ---No

If yes, did you find the discusson helpful?

-Y- ---No

2 . To what extent did you reach the following objectives through this course?

Not

achieved

Fully

1 2 3 4 5 achieved

Objective 1

Objective 2

Objective 3

Objective 4

Objective 5

1 2 3 4 5

1 2 3 4 5

1 2 3 4 5

1 2 3 4 5

1 2 3 4 5

149
3. Which objectives were most appropriate for your professional ctevelopment? Please explain:

4.

5. Which aspects of the subject matter covered in the course will you use most when you return to

Was the level of presentation of the subject matter:

simple? Too - About right? complex? Too

your job? Please explain:

6. Which aspects of the subject matter will you use least when returning to your job? Please

explain:

150
7 . Based on your own needs, what course topics would you recommend be expanded?

Shortened?

Omitted?

Added?

8 . What aspects of the course were most relevant to your country’s condiiions and to your role in

your country’s development? Please explain:

9 . Did the field trip provide you with practical applications of the course content? Please explain:

151
COURSE DESIGN AND DELIVERY

1.

2 .

3 .

During the course, was the daily schedule: Too About Too

-short? -right? -.-My?

Too About Too

Was the overall length of the course: ~Short? -right? -long?

Which of the following training methods were used to help you learn in this course? (Please

circle those that were used.)

a . Lecturettes

b . Large group discussions

C . Small group discussions

d . Case studies

8. Practical hands-on experience in the field

f. Classroom/laboratory exercises and experiments

9. Role plays/simulation

h . Individual consultations with instructors

i. Other

152
4 . Which of these methods most helped your learning and ability to use the skills? (List in the

order of priority for helping you learn.)

a .

b.

C .

d .

e.

5. Were any of these methods used too much? -Y= -No

If yes, please explain:

Were any of these methods used too little?

If yes, please explain:

--Ye --No

7. Could the course have been presented more effectively to meet your needs? Yes -No

If yes, please explain:

153
8. How useful were the following types of materials used in the course? (Please circle the

appropriate number.)

Not at all

useful 1 2

Written materials

(manuals, handouts, texts) 1 2

Audiovisual materials

Computer-assisted

instruction (ii used)

1 2

2

3 4

3 4

3 4

3 4

5

5

5

5

Extremely

useful

9. Please give examples of the most effective materials.

IO. Please give examples of the least effective materials.

154
INSTRUCTORS

Name of first instructor:

Please rate this instructor in the following areas:

Poor I 2 3 4 5 Excellent

Knowledge of subject matter 1 2 3 4 5

Training ability 1 2 3 4 5

Ability to relate material

to your country

Overall effectiveness

1 2 3 4 5

1 2 3 4 5

Comments:
Name of second instructor:

Please rate this instructor in the following areas:

Poor 1 2 3 4 5 Excellent

Knowledge of subject matter 1 2 3 4 5

Training ability 1 2 3 4 5

Ability to relate material

to your country 1 2 3 4 5

Overall effectiveness 1 2 3 4 5

Comments:

156
Name of third instructor:

Please rate this instructor in the following areas:

Poor 1 2 3 4 5 Excellent

Knowledge of subject matter

Training ability

1 2 3 4 5

1 2 3 4 5

Ability to relate material

to your country

Overall effectiveness

1 2 3 4 5

1 2 3 4 5

Comments:

157
Name of fourth instructor:

Please rate this instructor in the following areas:

Poor 1 2 3 4 5 Excellent

Knowledge o f subject matter 1 2 3 4 5

Training ability 1 2 3 4 5

Ability to relate material

to your country 1 2 3 4 5

Overall effectiveness 1 2 3 4 5

Comments:

158
OVERALL COURSE SATISFACTION

1. Would you recommend this course to other individuals with a background similar to

yours? -Yes -No

Please explain why:

2 . Please rate your overall satisfaction with the participation of your classmates.

Not at all satisfied 1 2 3 4 5 Extremely satisfied

Comments:

3 . Please rate your overall satisfaction with this course:

Not a t all satisfied 1 2 3 4 5 Extremely Satisfied

4 . What final comments or suggestions do you have on this course?

159
PARTICIPANT DATA

Home country: Gender: -Male -Female

Main field of education:

Highest degree achieved: Secondary (BNBS) (MA/MS) (Ph.D.)

Other:

Year in which highest degree was obtained:

Type of position currently occupied:

Scientific Administrative Technical Other

If you have managerial or administrative responsibilities, please describe them:

How long have you been working in your current position?.

Did you find language to be a problem in the course? Yes No.

If yes, please describe:

160
Bonner, F.T.; Vozzo, J.A.; Elam, W.W.; Land, S.B., Jr. 1994. Tree seed

technology training course. Instructor’s manual. Gen. Tech. Rep.

SO-106. New Orleans, LA: U.S. Department of Agriculture, Forest

Service, Southern Forest Experiment Station. 160 p.

A training manual that covers all aspects of tree seed technology from

collection to sowing.
This publication reports research involving pesticides. It does not contain recommendations

for their use, nor does it imply that the uses discussed here have been registered. All uses of

pesticides must be registered by approrpiate State and/or Federal Agencies before they can be

recommended.

Caution: Pesticides can be injurious to humans, domestic animals, desirable plants, and

fish or other wildlife -if they are not handled or applied properly. Use all pesticides selectively

and carefully. Follow recommended practices for the disposal of surplus pesticides and

pesticide containers.

The use of trade or firm names in this publication is for reader

information and does not imply endorsement by the U.S. Department of Agriculture of any product or service.