Final Report

Gamma Irradiation as a Quarantine Treatment Against

*Copitarsia decolora* (Guenée) in Fresh Asparagus

Bach. Marisela Huamán Maldonado

In collaboration with:

Instituto Peruano de Espárrago y Hortaliza -- IPEH

Instituto Peruano de Energía Nuclear -- IPEN

Servicio Nacional de Sanidad Agraria -- SENASA

Agricultural Research Service -- ARS

Universidad Nacional Mayor de San Marcos -- UNMSM

[Table of Contents:]{.underline}

1.  Approach and documentation of the problem 3

> 1.2 Objective

2.  Background 4

3.  Methodology 8

> 3.1 Methodology of identification and rearing of
>
> *Copitarsia decolora* 9
>
> 3.2 Dosimetry 11
>
> 3.3 Irradiation process 12
>
> 5\. Results and Discussion 14
>
> 5.1 Preliminary test
>
> 5.2 Confirmatory test
>
> 6\. Conclusions 16
>
> 6.1 Recommendation
>
> 7\. Bibliography 17
>
> 8\. Appendices 19
>
> 9\. Illustrations 21

1.1 APPROACH AND DOCUMENTATION OF THE PROBLEM.

In recent years, asparagus production on a worldwide scale has
established itself as a booming industry, due to the flavour,
nutritional value and ease of preparation of the asparagus, of which
Peru is currently the world's largest exporter.

In 2005, fresh Peruvian asparagus reached an export volume of 80 000 MT
(approx), our principal market being the United States of America (USA),
the destination of 65-70% of all fresh (mainly green) asparagus.

Currently, as a phytosanitary requirement for the importation of
Peruvian fresh asparagus to the USA, methyl bromide fumigation is
obligatory for the control of *Copitarsia spp*.

However, methyl bromide fumigation costs the fresh asparagus industry
amongst other things: a deterioration in product quality; a reduction in
the product shelf-life; a break in the cold-chain and annual expenses
estimated at over \$2.6 million dollars.

This research proposes the application of ionizing radiation as an
alternative means of quarantine, to methyl bromide fumigation.

1.2 OBJECTIVE.

The objective of the present work is to determine the minimum effective
dose of Gamma irradiation that ensures 100% non-emergence of adults of
*Copitarsia decolora* when applied to second instar larvae, while
guaranteeing the quality and useful life of the fresh green asparagus.

2\. BACKGROUND.

In 2000, USDA-APHIS gave notice of the stopping of all shipments of
fresh asparagus from Peru because of infestations of eggs and larvae up
to the 2^nd^ instar of *Copitarsia sp*., until methyl bromide fumigation
of all shipments of fresh asparagus was established as a phytosanitary
requirement for entry into the USA.

*Copitarsia incommoda* (Lepidoptera: Noctuidae) is a polyphagous pest
that affects crops of economic significance, and its distribution ranges
from Mexico through South America (Angulo and Weigert, 1975). Sanchez
and Apaza (2000), likewise consider this insect as one of the principal
pests of the cultivation of asparagus in Peru, as much as for the damage
it causes to the foliage, as for the clutches of eggs that the females
oviposit in the shoots during harvest, thus affecting the commercial
quality of the harvested product.

Vergara, et. al. (2002) observed that the biological cycle of
*Copitarsia* *incommoda* (Walker) in laboratory conditions (25°C and 25%
relative humidity (RH)) comprises an incubation period of the eggs of 3
days, the 6-stage larval period ranged from 18 to 26 days, the pupa
period between 13 and 16 days, the pre-ovipositional period of 2 to 6
days. Oviposition occurred for 10 to 15 days and the moth live and
additional mean of 0.6 days after oviposition

According to Simmons and Pogue (2004) *Copitarsia turbata* was
misidentified as *Copitarsi incommoda*. *C. turbata* (Herrich-Schaffer)
is placed as a new synonym of *C. decolora* (Guenée).

The Normas Técnicas Peruanas (Peruvian Technical Regulations, NTP)
regulate irradiation of foodstuffs in Peru.

-   NTP 209.500:2001: Irradiation of foodstuffs. Requirements. This
    technical regulation establishes the characteristics that an
    irradiation process must fulfil with respect to the source of
    ionizing irradiation, the dose applied, the facilities used, and
    control and labelling to be applied across the nation. The scope of
    this regulation is foodstuffs treated with ionizing irradiation.

-   NTP 209.501:2001 Irradiated foodstuffs. Classification. This
    technical regulation establishes the information that all packaged
    irradiated foodstuffs destined for human consumption must display.

-   NTP 209.502:2005: Code of Practice for the treatment of foodstuffs
    to be irradiated. This technical regulation establishes the working
    practices which must be applied to ensure that the treatment of
    foodstuffs by irradiation is effective and is carried out in such a
    way as to maintain product quality and to obtain foodstuff products
    which are harmless and fit for consumption.

History of Ionizing Irradiation as a Phytosanitary Treatment

In 1986 the first commercial shipment of mangoes irradiated as a
quarantine treatment against fruit flies was sent from Puerto Rico to
the state of Florida. This marks the first time that ionizing
irradiation was used as a phytosanitary treatment on a commercial scale.
The following year a shipment of Hawaiian papayas was irradiated and
sent to be marketed in the state of California; these were met with
general consumer acceptation (Hallman, 2001).

In 1989 the dose of 0.15kGy was approved as a quarantine treatment for
the elimination of fruit flies in papayas produced in Hawaii destined
for the United States of America (Hallman, 2001b). However it nor any
other irradiation phytosanitary treatment was used on a commercial scale
until 1995 when irradiation of Hawaiian fruits began on a continuing
basis.

In May 2000 the commercial entry into California of sweetpotatoes from
Florida irradiated as quarantine control for *Cylas formicarius
elegantulus* (Summers) was carried out.

Studies with respect to the quality carried out by the University of
Michigan, concluded that irradiating asparagus with a dose of between
0.01 -- 1.0 KGy slightly delayed the sprouting of buds, but had a small
effect on the colour and texture (Markakis and Nicholas, 1972); Cast
(1984) commented that a dose between 0.05KGy -- 0.15KGy extended the
shelf-life of asparagus, owing to the inhibition of sprouting; and in
Chile, Rubio et. al. (1988) observed that ionizing radiation had no
significant effect on the chemical, microbiological, or organoleptic
parameters of neither white nor green asparagus.

Later in Peru, Linares (1989), carried out working experiments with
irradiation on white asparagus, indicating that the most appropriate
dose for preserving asparagus is 1.5KGy followed by storage between 1
and 2°C and 90-95%RH.

In Argentina, Lescano et. al. (1993) demonstrated that asparagus (Var.
Argenteuil), when packed in polyester boxes and wrapped with a PVC film
and stored at 3±2°C ±9% and 92±5% RH, doubled its shelf-life when
treated with 2KGy of gamma radiation.

Nordion (1994) concluded that asparagus can tolerate an irradiation
de-infestation and its shelf life can probably be similar or slightly
longer than non-irradiated asparagus.

In Peru Cornejo et. al. (1994) observed that irradiated at a dose of
1KGy; fresh green asparagus did not suffer noticeable changes in terms
of its organoleptic or shelf-life characteristics.

In December 2003, the main investigators from APHIS, representatives
from SENASA, the Peruvian Asparagus and Vegetables Institute (IPEH) and
the Peruvian Institute for Nuclear Energy (IPEN) joined together for a
video conference, in which it was decided that investigation should be
conducted in two stages. The preliminary, to search for the minimum
effective dose and the confirmatory stage would consist of confirming
the required dose that would need to be applied in 2^nd^ instar larvae.
The 2nd instar was targeted as it was the most developed instar found in
shipments of Peruvian fresh asparagus in the United States, and
secondly, as the most developed, it was therefore felt to be the most
resistant to quarantine treatment.

Subsequently, APHIS communicated that at least 10 000 larvae 2^nd^
instar should be used for the confirmatory test. (See Appendix 1).

In July 2005, Doctor Guy Hallman (ARS) visited Peru and supervised the
investigation. A report was issued detailing his satisfaction with the
progress. (See Appendix 2).

3\. METHODOLOGY.

The investigation consists of three stages: preliminary trial consists
of searching for the minimum effective dosage that ensured 100%
non-emergence of adult *Copitarsia decolora* from stage 2 larvae,
followed by the confirmatory stage which consisted in treating at least
10 000 larvae 2^ND^ instar with the dosage discovered, and the final
stage to determine the effects of the irradiation dose on fresh green
asparagus. This document deals with the description of these three
stages.

For the preliminary test identifications were carried out on *Copitarsia
decolora* collected in fields of asparagus, these were later bred for
three generations in two laboratories; the first at the University of
San Marcos in environmental conditions of 24±3°C and 63±9% RH and the
second at the Peruvian Institute of Nuclear Energy in environmental
conditions of 23±3°C and 63±7% RH. (Figure 1)

Before carrying out the preliminary test, a dosimeter test was performed
on 100 2^nd^ instar larvae to ensure that doses were approximated as
closely as possible.

Having performed the dosimeter test, the preliminary test was begun,
that consisted in searching for the minimum dose required to ensure a
100% non-emergence of *C. decolora* adults from larvae 2^nd^ instar.

Having estimated the minimum dose, the confirmatory test was carried out
on at least 10 000 2^nd^ instar larvae, To do this a new dosimeter test
was done, and 10 repetitions were set up, each one with approximately
1000 2^nd^ instar larvae.

3.1. Method of Identification and breeding of *Copitarsia decolora*.

[Collection.]{.underline}

Collection of the larvae of *Copitarsia decolora* was made in asparagus
fields; they were placed in two litre plastic flasks together with fresh
leaves. These were then transferred to the laboratory (Figure 2).

[Identification.]{.underline}

In the entomology laboratory the collected larvae were compared to the
taxonomical keys provided by Angulo and Weigert (1975) and Castillo and
Angulo (1991), to separate the species *Copitarsia decolora* from other
Noctuidae.

To identify the larvae, the nature of their taxonomical characteristics
was used *ad hoc.* To identify adults, characteristics of the external
morphology and the morphology of the genitalia, were used. The genitalia
and associated assemblies were extracted to undertake the identification
(Figure 3).

[Conditioning.]{.underline}

The larvae were placed in 7 liter plastic containers at 24±3°C y 63±9%
HR, in the base of which absorbent paper was placed to maintain the
humidity. The larvae were provided with their natural diet of fresh
asparagus leaves, which, before being given to the larvae was
disinfected with 5 ml of 6%Sodium Hypochlorite in 20 liter of water in
addition to 0.1 mg of tetracycline. The foliage was obtained from
asparagus fields and from the University of San Marcos, neither of which
had used insecticides, to avoid mortality in the larvae. Food was given
to the larvae daily, the upper part of the containers was covered with
an organdy cloth and secured with an elastic band to prevent the larvae
from escaping.

Pupae: When the larvae reached the pre-pupa stage, they were transferred
to new containers, the bases of which had been prepared with sand
(disinfected in an oven at 100°C for a period of two hours) to provide
the larvae with a suitable habitat. After ten days the larvae passed to
the pupal stage and they were then sexed. Using a magnifying glass, the
sexual characteristics of the distal part of the abdomen of the pupae
were observed (Figure 5).

Adults: After sexing the pupae, a single pair of *C. decoloara* was
placed in a plastic flask. Craft paper had been prepared in the sides of
the flask, so that when the adults emerged and were sexually mature, the
females would place their egg clutches on the paper facilitating their
collection. The adults were given a diet consisting of one part honey,
one part pollen and a quarter part water, this was placed in a 5-ml
plastic container with a cotton wick to facilitate feeding. feeding was
done every other day (Figure 6).

Eggs: Approximately 500 eggs were placed in one plastic 7-liter flask
covered with an organdy cloth held down with elastic tape to help
ventilation and prevent the recently emerged larvae from escaping. On
observing eclosion, the date was noted and they were given their natural
diet for their development to continue.

Larvae were separated in groups of 50 according to developmental stage
and later into groups of 25 individuals per flask to avoid cannibalism.

[Egg Collection.]{.underline}

Three days after emergence of the adult, the paper was inspected for egg
clutches and the paper was changed, Egg masses were collected every
three days. (Figure 7).

[Renewal of Biological Material.]{.underline}

After three generations (F3) the *Copitarsia decolora* colony was
renewed with individuals collected from the field, the aim being that
the individuals in the laboratory would have the same potential to
develop, survive and reproduce as would those in the wild.

3.2. Dosimetry.

The dosimetry system is used to determine the absorbed dose, and
consists of dosimeters, which can be chemical solutions, polymers or any
material which undergoes physical or chemical changes under irradiation
and equipment to measure these changes and convert them into grays.
Different dosimetry systems take as their reference the internationally
recognized standards. The irradiation dosimetry system documents that
the product has received the minimum dose for the desired process.

Fricke dosimetry was used, which is based on the International ASTM
Standard: E1026-95. The calibration and certification was performed
using the alanine reference dosimeter system of the International Atomic
Energy Agency (IAEA), with the International Dose Assurance Service
(IDAS) acting as intermediary.

-   Dosimetry Procedure for 2^nd^ instar larvae:

> Two petri dishes were placed in the center of the irradiation chamber
> of the Gammacell 220 Excel (Figure 7), in each of the dishes 50 2^nd^
> instar larvae were placed, later 4 dosimeter ampoules were placed in
> the four extremities of the dishes and irradiation was carried out for
> predetermined periods of time.
>
> Once this time had elapsed, the ampoules were removed and the
> absorption of the irradiated solution was determined at 305 nm via a
> spectrophotometer.
>
> The irradiation process should have the lowest dose uniformity ratio
> (DUR) possible. In these studies the goal is for the DUR to not exceed
> 1.2, that is, the difference between minimum and maximum dose must not
> be greater than 20%.
>
> 3.3. Irradiation Process.
>
> A Gammacell 220 irradiator (Figure 8) was used, which currently has a
> dose rate of 18 KGy, this value does not exceed the dose rate in
> normal commercial use of a million curies for older plants.
>
> **A. Preliminary Test (June -- November 2004)**
>
> 100 2^nd^ instar larvae were administered a dose of 30 Gy, on
> observing any emergence of adults, the dose was increased. This
> sequence was continued until the minimum dosage was reached which
> impeded emergence of the adult stage in 100% of the 2^nd^ instar
> larvae. The minimal dose found was 100Gy.
>
> **Irradiation Conditions.**

1.  Second instar larvae reared in the laboratory and maintained on
    their natural diet of fresh asparagus foliage, were used.

2.  The larvae were placed in two plastic petri dishes in the centre of
    the irradiator.

3.  Only individuals from the first, through third generations were
    used.

4.  Once irradiated, the larvae were placed in containers with suitable
    food and placed in the laboratory to monitor development. The same
    procedure was performed with the control.

**Post irradiation evaluation:**

Both the treatment and the control were evaluated on a daily basis by
quantifying the live and dead larvae. Food continued to be provided for
those that developed to the next stage until their death.

**B. Confirmatory Test. (May -- December 2005)**

100Gy was to be confirmed as the appropriate dose in approximately 10
000 individuals. For this test to be successful, 100% of the individuals
must not reach the adult stage.

**Dosimeter for the confirmatory test.**

Following the recommendations of Dr. Hallman to reduce the DUR, a
smaller flask was placed in the centre of the irradiation chamber
(Figure 8). In the flask were 1000 2^nd^ instar larvae and 3 dosimeter
ampoules in the extremities of the flask. The ampoules were removed, and
the absorbance of the irradiated solution was tested at 305 nm using a
spectrophotometer.

In addition two routine dosimeters were used. These involved placing
Gafchromic HD 810 films on the insides of the flask. These were read in
a spectrophotometer. The routine dose assists in verifying that the
product is receiving the required dose.

**Irradiation conditions for the confirmatory test.**

-   10 repetitions were to be performed, each one to containing 1000
    2^nd^ instar larvae.

-   In each repetition Gafchromic HD 810 film was placed on the insides
    of the flask.

-   As in the preliminary stage, larvae at the second instar, grown in
    the laboratory and fed on their natural diet of fresh asparagus
    foliage, were used.

-   The larvae were placed in the new flask in the centre of the
    irradiator (Figure 10).

-   Once irradiated, the larvae were placed in containers with suitable
    food for observation of development in the laboratory. The same
    procedure was performed with the control,.

**Post irradiation evaluation:**

As far was possible, both the treatment and the control were evaluated
on a daily basis; by quantifying the live and dead larvae, and food
continued to be provided for those that matured to their next stage
until their death.

> 5\. Results and Discussion.
>
> 5.1 Preliminary Test -- Percentage of adults surviving the
> irradiation.

  ------------ --------- ------------- ------- ------- ------- ------- ------- ------ ------ ------ ------------
  Dose         \#        Dates of                                                                   Total
               numbers   Irradiation                                                                Replicates
               treated                                                                              

                         3 Jun.        19      6 Sept. 9 Sept. 14      18      20     22     10     
                                       Agst.                   Sept.   Sept.   Oct.   Oct.   Nov.   

  30Gy.        100       37%           \*      \*      \*      \*      \*      \*     \*     \*     1
               larvae II                                                                            

  40Gy.        100       35%           \*      \*      \*      \*      \*      \*     \*     \*     1
               larvae II                                                                            

  60Gy.        100       30%           \*      \*      \*      \*      \*      \*     \*     \*     1
               larvae II                                                                            

  80Gy.        100       1%            0       0       0       0       0       \*     \*     \*     6
               larvae II                                                                            

  100Gy.       100       \*            \*      0       0       0       0       0      0      0      7
               larvae II                                                                            

  Control^1^   100       43%           44%     48%     43%     52%     34%     39%    45%    42%    9
               larvae II                                                                            
  ------------ --------- ------------- ------- ------- ------- ------- ------- ------ ------ ------ ------------

> \*no test performed.

[^1]

It can be observed that the three first doses tested have a high
percentage of emerged adults, therefore these doses were not considered
further.

Repeated tests were carried out at 80Gy, in the 6 tests performed, one
individual succeeded to mature to the flying adult stage, therefore 80
Gy was not considered further.

Tests were begun at 100Gy, 100% of the larvae treated with this dose did
not reach pupa stage after 700 larvae were treated. The most developed
individuals were pre-pupa.

**5.2 Results of the Confirmatory Test.**

5.2.1. 10 Repetitions performed on 5 different dates:

  ------------- ------------ --------- ------------ ------------ ------------ --------------
  Dates         Repetition   N° larvae \% Prepupae  \% Pupae     \% Adults    Irradiation
  irradiation                II        from         from         from         efectiveness
                             treated   irradiated   irradiated   irradiated   from Larvae II
                                       larvae II    larvae II    larvae II    

  12/05/2005    I            1000      0%           0%           0%           100%

  12/07/2005    II           1025      0%           0%           0%           100%

                III          1050      0%           0%           0%           100%

  14/07/2005    IV           1025      0,10%        0%           0%           100%

                V            1025      0,20%        0%           0%           100%

                VI           1030      0%           0%           0%           100%

  18/07/2005    VII          1000      0,10%        0%           0%           100%

                VIII         1017      0,30%        0%           0%           100%

  06/10/2005    IX           1034      0,10%        0%           0%           100%

                X            1010      0,50%        0%           0%           100%

  Total de                   10216                                            
  larvas                                                                      
  irradiadas                                                                  
  ------------- ------------ --------- ------------ ------------ ------------ --------------

Of 10216 larvae treated, none survived the irradiation, thus we conclude
that the dose of 100Gy is effective in preventing adult emergence from
irradiated 2^nd^ instars.

**5.2.2. Result of the control.**

The Control is the samples that were not irradiated.

+--------------+---------+---------+-------------------+-------------+
| **Dates**    | **N of  | **      | **%**             | **% pupa    |
|              | l       | Pupae** |                   | to**        |
|              | arvae** |         | **Adult**         |             |
|              |         |         |                   | **Adult**   |
|              |         |         | **Survivors**     |             |
+--------------+---------+---------+-------------------+-------------+
| 12/05/2005   | 100     | 61      | 55 ( adults)      | 88.50%      |
+--------------+---------+---------+-------------------+-------------+
| 12/07/2005   | 100     | 67      | 51 (adults)       | 76.10%      |
+--------------+---------+---------+-------------------+-------------+
| 14/07/2005   | 100     | 79      | 64 (adults)       | 81%         |
+--------------+---------+---------+-------------------+-------------+
| 18/07/2005   | 100     | 69      | 53 (adults)       | 76.80%      |
+--------------+---------+---------+-------------------+-------------+
| 06/10/2005   | 100     | 76      | 58 ( adults)      | 75%         |
+--------------+---------+---------+-------------------+-------------+

It can be observed that the percentage of pupae that pass into the adult
stage is in the range of 75-88.5%.

6\. CONCLUSIONS.

-   The minimum effective dose of gamma irradiation which prevents
    emergence of adult *Copitarsia decolora* from 2^nd^ instar larvae is
    100Gy.

-   Irradiation was achieved of 10 replicates of at least 1000 2^nd^
    instar larvae, or approximately 10 000 2^nd^ instar larvae, as
    agreed to by APHIS for the confirmatory test.

-   The effects of irradiation on larvae were:

    1.  Retardation of the biological cycle.

    2.  Change in the cuticle (Figure 12)

**6.1 Recommendations.**

It is recommended that APHIS approves the quarantine treatment method of
ionizing radiation at an absorbed minimum dose of 100 Gy as an
alternative to methyl bromide fumigation.

7\. BIBLIOGRAPHY

**Angulo , A. O., and T. S. Olivares.** 2003 Taxonomic update of the
species of Copitarsia Hampson ( Lepidoptera: Noctuidae: Cucullinae).
Gayana Zool. 67: 33-38

**Artigas,J. N., and A. O. Angulo** 1973 *Copítarsia consueta* (Walker)
Biología e importancia Económica en el Cultivo de Raps ( Lepidoptera:
Noctuidae) Bol.Soc.Biol.. de Concepción 46: 199-216.

**Bautista, N. H. and Flores, R.P.** 2003 Simposio Nacional Sobre
*Copitasia incommoda* memorias Mexico 65pp.

**Castillo, E. E., and A. O. Angulo** 1991 Contribution to the knowledge
of the genus Copitarsia Hampson,1906 ( Lepidopetra Glossata :
Cucullinae) Gayana Zool. 55: 227-246.

**Hallman, G. J.** 1998 Ionizing radiation quarantine treatments An.
Soc. Entomol. Brasil 27(3) 313-323.

**Hallman, G. J.** 2000 Expanding radiation quarantine treatments beyond
fruit flies Agricultural and Forest Entomology 2: 86-96

**Hallman, G. J.** 2001 Irradiation as a quarantine treatment pp.
113-130 in: Molins, RA, Fod Irradiation: Principles and Applications.
John Wiley & Sons.

**Hallman, G. J.** 2001 Ionizing irradiation quarantine treatment
against sweetpotato weevil (Coleoptera: Curculionidae) Florida
Entomologist Vol. 84 N(3) 415-417.

**Lescano, P. Narvaiz and Kairiyama** 1993 Gamma Irradiation of
Asparagus ( Asparagus officinalis, var. Argenteuil) Comisión Nacional de
Energía Atómica Gerencia de Radioisotopos y Radiaciones Vol.26 411-416 .

**Linares, A. M.** 1990 Estudio de la Radio Preservación del Espárrago (
Asparagus Officinalis) Tesis para optar el título de Ingenio químico
Universidad nacional Del Callao Lima Perú.

**Marcotte M.** 1994 Suitability of irradiation disinfestation for
produce previously deemed to be unsuitable or sensitive Nordion
International Canada 10pp.

**Poole, R.** 1989 Lepidopterum Catalogus Fáciles 118: Noctuidae, part E
J. Brill, Leiden, The Netherlands.

**Rubio, C., Espinoza, B. And A. Godoy** 1989 Efecto de la Radiación
Ionizante en la Calidad Comercial de los Espárragos de Exportación
Nucleotécnica año 9 N°16 (Abril 1989) 17-29

**Sánchez, V. And W. T. Apaza** 2000 Plagas y Enfermedades del Espárrago
en el Perú . Instituto Peruano del Espárrago Lima - Perú.

**Simmons, R. and Pogue.** 2004 Redescription of two often confused
noctuid pests, *Copitarsia decolora* and *Copitarsia incommoda* (
Lepidoptera: Noctuidae) Annals of the Entomological Society of America
vol. 97.no 6 1159-1164

XLI Convención Nacional de Entomología 2002 Resúmenes Universidad
Nacional Agraria la Molina 164 pp.

8.- APPENDICES

**Appx.1**

![](media/image1.png){width="4.083333333333333in" height="6.125in"}

**Appx2.- Report on Effort Toward an Irradiation Quarantine Treatment to
Disinfest Peruvian Asparagus of the Insect *Copitarsia decolora.***

Guy Hallman

USDA-ARS

2413 E. Highway 83

Weslaco, TX 78596 USA

<ghallman@weslaco.ars.usda.gov>

From July 12-18 I visited Peru in part to consult on a research program
to develop an irradiation quarantine treatment against the noctuid moth
*Copitarsia decolora* on asparagus so that irradiation may be offered as
an alternative to methyl bromide fumigation for exports of asparagus to
the United States of America. The research is being carried out by
Marisela Huamán Maldonado of the Universidad Nacional Mayor de San
Marcos (UNMSM) with the aid of the Intituto Peruano de Espárrago y
Hortalizas, the Instituto Peruano de Energia Nuclear (IPEN), and the
Servicio Nacional de Sanidad Agraria of Peru (SENASA).

Based on a December 2003 conference call with officials from the United
States Department of Agriculture, Animal and Plant Health Inspection
Service as well Peruvian parties from the previous-mentioned
organizations, it was decided that 2^nd^ instars were the most developed
stage of *C. decolora* found on imported asparagus and that the
criterion for efficacy would be prevention of the adult stage from
irradiated 2^nd^ instars.

The study is proceeding well, and Huamán presented an update at the
2^nd^ National Technical Congress of Asparagus held in Lima 15-16 July.
Two colonies of *C. decolora* have been established, one at IPEN and one
at UNMSM. Preliminary results indicate that about 100 Gy would be
sufficient to prevent the development of adults from irradiated 2^nd^
instars. Preliminary testing has shown that asparagus tolerates several
times this dose. The final dose would need to be confirmed by testing of
a large number of insects. That number was not agreed upon during the
conference call, but Huamán will test 10,000 2^nd^ instars.

According to what I have seen, I suspect that enough data will be
generated by the end of this calendar year to support a relatively low
dose (\~100 Gy) irradiation quarantine treatment for asparagus or any
other host of *C. decolora* for up to and including 2^nd^ instars.

9.- ILLUSTRATIONS.

Figure 1: The breeding laboratory at the UNMSM.

![](media/image2.jpeg){width="2.8493055555555555in"
height="3.267361111111111in"}

### Figure 2: Collection of the larvae in the asparagus fields 

![](media/image3.jpeg){width="2.875in" height="3.3020833333333335in"}

Figure 3 Identification based on ad hoc morphological characteristics.

![](media/image4.jpeg){width="2.4027777777777777in"
height="2.982638888888889in"}

Figure 4.-Asparagus field at the UNMSM.

![](media/image5.jpeg){width="3.3555555555555556in"
height="3.8444444444444446in"}

Figure 5: Pupae in disinfected sand.

![](media/image6.jpeg){width="2.8319444444444444in"
height="3.2472222222222222in"}

Figure 5: Adults.

![](media/image7.jpeg){width="2.658333333333333in"
height="3.053472222222222in"}

Figure 6: Egg collection.

![](media/image8.jpeg){width="2.15625in" height="2.4715277777777778in"}

Figure 7: The two petri dishes located in the centre of the irradiator
cylinder

![](media/image9.jpeg){width="2.345833333333333in"
height="3.0833333333333335in"}

Figure 8: Container for the confirmatory test.

![](media/image10.jpeg){width="3.311111111111111in"
height="2.4590277777777776in"}

Figure 9: Gammacell Excel 220 Irradiator.

![](media/image11.jpeg){width="2.2125in" height="2.5409722222222224in"}

Figure 10: Container with approximately 1000 2nd instar larvae to be
irradiated with 100Gy.

![](media/image12.jpeg){width="2.7430555555555554in"
height="1.9986111111111111in"}

Figure 11: Larvae irradiated with 80Gy.

Death of larvae due to irradiation

![](media/image13.jpeg){width="2.667361111111111in"
height="3.058333333333333in"}
![](media/image14.jpeg){width="2.511111111111111in"
height="1.8833333333333333in"}

Figure 13: Change in the cuticle.

Figure 12 Dead larvae after 100Gy.

![](media/image15.jpeg){width="2.1381944444444443in"
height="2.7381944444444444in"}

![](media/image16.jpeg){width="1.023611111111111in" height="2.625in"}

[^1]: The control gave an average of 43% survivals; however, this is
    equivalent to 84% of adults which successfully emerged from pupa
    stage. This indicates that \>50% of pupae passed from pupa to adult
    stage (the percentage recommended by Dr. Hallman).