Characteristic

expression

of

twelve

rice

family

genes

in

response

to

pathogen

infection

wounding

and

defense-related

signal

compounds

-LRB-

-RRB-

Pathogenesis-related

-LRB-

PR

-RRB-

proteins

have

been

used

as

markers

of

plant

defense

responses

and

are

classified

into

families

However

precise

information

on

the

majority

members

in

specific

PR

families

is

still

limited

We

were

interested

in

the

individual

characteristics

of

rice

family

genes

and

selected

putatively

active

genes

using

rice

genome

databases

for

expressed

genes

All

were

upregulated

upon

compatible

and/or

incompatible

rice-blast

fungus

interactions

three

were

upregulated

in

the

early

infection

period

and

four

in

the

late

infection

period

Upon

compatible

rice

bacterial

blight

interaction

four

genes

were

upregulated

six

were

not

affected

and

one

was

downregulated

These

results

are

in

striking

contrast

to

those

among

genes

where

only

one

gene

was

pathogen-inducible

The

responses

of

individual

genes

to

salicylic

acid

jasmonic

acid

and

ethylene

induced

defense

signaling

pathways

in

rice

are

likely

to

be

different

from

those

in

dicot

plants

Transcript

levels

in

healthy

leaves

roots

and

flowers

varied

according

to

each

gene

Analysis

of

the

partially

overlapping

expression

patterns

of

rice

genes

in

healthy

tissues

and

in

response

to

pathogens

and

other

stresses

would

be

useful

to

understand

their

possible

functions

and

for

use

as

characteristic

markers

for

defense-related

studies

in

rice

Introduction

Pathogenesis-related

-LRB-

PR

-RRB-

proteins

which

are

classified

into

families

accumulate

after

pathogen

infection

or

related

situations

in

many

plant

species

-LRB-

van

Loon

et

al.

-RRB-

Among

the

PR

gene

family

genes

have

been

frequently

used

as

marker

genes

for

systemic

acquired

resistance

in

many

plant

species

However

only

a

small

proportion

of

genes

have

been

studied

among

the

many

members

in

this

family

and

information

on

the

other

members

is

limited

For

example

b

and

c

genes

for

acidic

proteins

and

-LRB-

-RRB-

and

genes

for

basic

proteins

were

reported

in

tobacco

-LRB-

Nicotiana

tabacum

-RRB-

-LRB-

van

Loon

and

van

Strien

-RRB-

but

little

information

is

known

on

other

members

To

understand

the

characteristics

and

redundancy

of

the

majority

of

family

members

genome-based

studies

are

necessary

For

such

studies

dicot

Arabidopsis

and

monocot

rice

-LRB-

Oryza

sativa

L.

-RRB-

plants

have

been

used

as

the

model

plants

In

Arabidopsis

genes

are

listed

as

predicted

genes

that

encode

homologous

proteins

to

tobacco

protein

which

was

first

reported

as

an

acidic

protein

in

tobacco

leaves

infected

with

Tobacco

mosaic

virus

-LRB-

TMV

-RRB-

-LRB-

van

Loon

et

al.

-RRB-

Among

the

genes

only

one

gene

-LRB-

-RRB-

which

encodes

a

basic

protein

is

known

to

be

pathogen-responsive

and

the

other

genes

reportedly

did

not

respond

to

either

bacterial

or

fungal

pathogens

-LRB-

van

Loon

et

al.

-RRB-

From

these

results

we

tend

to

suppose

that

only

one

gene

relates

to

pathogen

resistance

in

Arabidopsis

and

the

others

contribute

to

other

functions

In

rice

the

induction

of

two

genes

and

by

blast

fungus

infection

was

reported

-LRB-

Agrawal

et

al.

-RRB-

They

encode

putative

acidic

and

basic

proteins

respectively

and

also

responded

to

environmental

stresses

and

treatments

with

some

chemicals

-LRB-

Agrawal

et

al.

b

-RRB-

However

for

other

rice

gene

family

members

there

is

only

limited

information

except

for

their

presence

and

expression

-LRB-

-RRB-

at

least

signals

for

possible

rice

proteins

responsive

to

anti-tobacco

antibodies

were

found

in

an

extract

of

blast

fungus-infected

rice

leaves

-LRB-

Schweizer

et

al.

Iwai

et

al.

-RRB-

and

-LRB-

-RRB-

predicted

genes

were

proposed

to

be

present

in

the

rice

genome

-LRB-

van

Loon

et

al.

-RRB-

To

study

the

response

of

individual

rice

genes

to

pathogens

we

selected

active

rice

genes

from

the

rice

genome

databases

for

expressed

genes

and

studied

their

induced

expression

by

real

time

RT-PCR

-LRB-

qPCR

-RRB-

In

striking

contrast

with

the

result

in

Arabidopsis

all

rice

genes

selected

here

were

upregulated

by

blast

fungus

infection

The

levels

of

constitutive

expression

in

organs

and

induced

expression

by

different

treatments

varied

according

to

the

gene

In

addition

to

the

data

on

tissue-specific

expression

in

transgenic

rice

with

a

promoter

β-glucuronidase

-LRB-

GUS

-RRB-

fusion

gene

we

list

the

characteristics

of

the

rice

genes

This

is

the

first

example

of

a

comparison

of

the

expression

of

the

majority

of

members

of

a

monocot

PR

family

to

our

knowledge

This

information

will

be

useful

for

further

studies

on

PR

genes

and

on

resistance

mechanism

in

rice

plants

Materials

and

methods

Plant

materials

Wild-type

-LRB-

WT

-RRB-

rice

-LRB-

Oryza

sativa

cv

Nipponbare

-RRB-

and

the

near

isogenic

line

-LRB-

Ise

and

Horisue

-RRB-

which

contains

the

R

gene

Pi-i

against

blast

fungus

-LRB-

Magnaporthe

grisea

-RRB-

race

-LRB-

Yamada

et

al.

-RRB-

were

grown

for

days

in

soil

-LRB-

Bonsol

No.

Sumitomo

Chemicals

L.

Japan

-RRB-

in

a

greenhouse

at

°C

in

the

day

time

and

°C

at

night

The

fourth

leaves

of

plants

at

the

stage

were

mainly

used

as

the

experimental

material

The

seeds

of

transformants

with

GUS

were

germinated

on

agar

medium

containing

μg

ml

hygromycin

transferred

to

soil

at

days

after

imbibition

and

grown

in

the

greenhouse

Five-day-old

seedlings

on

agar

medium

and

plants

in

the

greenhouse

were

used

for

GUS-staining

assays

Infection

with

pathogens

Magneporthe

grisea

race

-LRB-

isolate

-RRB-

was

grown

on

oat-meal

medium

-LRB-

Difco

-RRB-

for

weeks

and

conidia

were

induced

under

BLB

light

-LRB-

BLB

Toshiba

-RRB-

for

days

at

°C

The

rice

seedlings

of

Nipponbare

and

plants

at

the

stage

were

spray-inoculated

with

a

conidia

suspension

-LRB-

conidia

ml

-RRB-

containing

Tween

and

the

inoculated

plants

were

incubated

under

high

humidity

in

the

dark

for

h

and

then

moved

to

the

greenhouse

Under

these

conditions

about

local

lesions

were

induced

per

leaf

on

Nipponbare

and

For

bacterial

blight

infection

Nipponbare

plants

which

are

compatible

with

Xanthomonas

oryzae

pv

oryzae

-LRB-

Xoo

-RRB-

strain

-LRB-

race

I

MAFF

-RRB-

were

inoculated

by

cutting

the

leaf

top

with

scissors

that

had

been

dipped

in

a

suspension

containing

cfu/ml

of

Xoo

and

incubated

in

the

greenhouse

Treatment

with

chemicals

Plastic

pots

-LRB-

cm

-RRB-

with

rice

seedlings

at

the

stage

were

dipped

in

ml

solutions

containing

mM

acid

-LRB-

ACC

-RRB-

or

mM

sodium

salicylate

-LRB-

SA

-RRB-

solution

and

incubated

for

h

in

the

greenhouse

For

jasmonic

acid

-LRB-

JA

-RRB-

treatment

pots

with

seedlings

each

was

put

in

an

air-tight

clear

plastic

box

and

a

cotton

pad

with

volatile

methyl

jasmonate

-LRB-

MeJA

-RRB-

dissolved

in

EtOH

was

put

at

the

corner

of

the

box

to

give

a

final

concentration

of

μM

and

incubated

for

h

The

fourth

leaves

were

used

for

RNA

extraction

Quantitative

real-time

RT-PCR

To

analyze

the

response

of

genes

to

biotic

and

abiotic

stresses

quantitative

real-time

RT-PCR

-LRB-

qPCR

-RRB-

was

conducted

using

iQ

SYBR

Green

Supermix

-LRB-

BioRad

Hercules

CA

USA

-RRB-

and

an

iCycler

-LRB-

BioRad

-RRB-

according

to

the

manufacturers

instructions

At

least

three

independent

biological

samples

were

used

with

specific

primers

for

each

individual

gene

-LRB-

Table

-RRB-

The

data

were

normalized

by

the

value

of

an

actin

gene

-LRB-

-RRB-

and

fold

change

in

the

expression

level

was

calculated

compared

with

that

of

healthy

fourth

leaves

and

standard

deviation

-LRB-

SD

-RRB-

values

are

shown

Construction

of

plasmids

for

transformation

of

rice

The

promoter

region

of

the

gene

-LRB-

in

this

work

-RRB-

was

obtained

through

PCR

amplification

from

the

Nipponbare

genome

using

a

GenomeWalkerTM

kit

-LRB-

Invitrogen

CA

USA

-RRB-

in

accordance

with

the

manufacturer

's

instructions

-LRB-

Accession

no

Kawahigashi

et

al.

-RRB-

The

amplified

fragment

was

digested

by

NotI

and

BamHI

and

inserted

into

the

vector

to

construct

the

GUS

plasmid

-LRB-

Fig.

-RRB-

The

GUS

plasmid

contained

bp

of

the

promoter

and

the

coding

region

for

amino

acids

at

the

N-terminus

of

the

gene

to

express

a

fused

GUS

protein

effectively

-LRB-

Kawahigashi

et

al.

-RRB-

Transformation

of

rice

Agrobacterium

tumefaciens

was

transformed

with

the

constructed

vector

-LRB-

Kawahigashi

et

al.

-RRB-

The

transformation

of

rice

was

performed

by

Agrobacterium

infection

as

described

by

Toki

et

al.

-LRB-

-RRB-

Histological

GUS

analysis

Histological

analysis

for

GUS

activity

was

performed

at

°C

essentially

as

described

by

Ohshima

et

al

-LRB-

-RRB-

using

a

modified

reaction

mixture

mM

phosphate

buffer

-LRB-

pH

-RRB-

containing

mM

glucuronide

-LRB-

X-gluc

-RRB-

methanol

μg/ml

cycloheximide

and

mM

dithiothreitol

The

reaction

was

stopped

by

the

addition

of

ethanol

Results

Characterization

of

the

rice

gene

family

Using

the

amino

acid

sequence

of

the

mature

tobacco

acidic

protein

-LRB-

amino

acids

Ohshima

et

al.

-RRB-

as

the

probe

we

searched

for

expressed

genes

in

rice

-LRB-

Oryza

sativa

cv

Nipponbare

-RRB-

encoding

homologous

proteins

with

a

similar

size

range

-LRB-

within

amino

acids

-RRB-

using

a

full-length

cDNA

database

-LRB-

-RRB-

and

rice

EST

database

-LRB-

-RRB-

The

presence

and

location

of

these

genes

were

confirmed

in

the

genome

database

of

rice

-LRB-

-RRB-

After

removing

repetitive

clones

and

confirmation

of

the

sequences

finally

genes

were

selected

as

candidates

for

active

rice

genes

which

encode

proteins

with

sequences

identical

to

that

of

tobacco

protein

at

least

in

the

C-terminal

amino

acids

Previously

two

rice

genes

were

reported

to

encode

an

acidic

protein

and

a

basic

protein

-LRB-

Agrawal

et

al.

-RRB-

which

were

upregulated

by

blast

fungus

infection

In

addition

to

the

two

we

found

another

active

genes

on

chromosomes

and

-LRB-

Table

-RRB-

Proposed

names

were

given

to

distinguish

each

gene

to

avoid

confusion

in

this

study

according

to

the

location

on

the

chromosomes

For

example

-LRB-

-RRB-

located

on

the

side

of

the

gene

for

-LRB-

-RRB-

on

chromosome

was

designated

as

and

was

On

chromosome

four

genes

and

were

clustered

in

tandem

from

the

side

to

the

side

in

the

sense

orientation

in

this

order

The

predicted

isoelectric

point

-LRB-

pI

-RRB-

of

each

mature

protein

is

shown

in

Table

for

reference

The

amino

acid

sequences

of

the

predicted

mature

proteins

for

the

genes

were

compared

with

those

of

representative

acidic

and

basic

proteins

from

Arabidopsis

-LRB-

and

-RRB-

and

tobacco

-LRB-

and

-RRB-

respectively

-LRB-

Fig.

-RRB-

The

sequences

of

the

and

listed

here

are

well

conserved

As

the

N-termini

of

and

are

not

conserved

these

regions

were

eliminated

for

the

alignment

in

Fig.

Ten

amino

acid

residues

-LRB-

closed

circles

-RRB-

are

completely

conserved

among

the

rice

Arabidopsis

and

tobacco

genes

indicating

they

are

important

for

the

predicted

roles

of

proteins

Table

of

rice

protein

genesPlantAccession

No

-LRB-

Symbol

name

-RRB-

Locus

nameReferencesPIProposed

-LRB-

-RRB-

et

al.

-LRB-

-RRB-

-LRB-

-RRB-

et

al.

-LRB-

-RRB-

-LRB-

like

-RRB-

et

al.

-LRB-

-RRB-

-LRB-

-RRB-

et

al.

-LRB-

-RRB-

-LRB-

-RRB-

-LRB-

-RRB-

Ohshima

et

al.

-LRB-

-RRB-

-LRB-

-RRB-

Eyal

et

al.

-LRB-

-RRB-

Fig.

of

the

gene

family

in

rice

plants

a

Amino

acid

sequence

alignment

of

putative

proteins

from

rice

The

predicted

mature

proteins

encoded

by

these

genes

were

aligned

by

GENETICS

-LRB-

Software

Development

Co.

Tokyo

-RRB-

Because

the

N-termini

of

and

are

considerably

different

from

those

of

the

other

these

regions

were

eliminated

from

the

figure

and

the

conserved

C-termini

among

the

were

aligned

The

acidic

and

basic

proteins

from

Arabidopsis

-LRB-

and

-RRB-

and

tobacco

-LRB-

and

-RRB-

were

used

as

representatives

of

acidic

and

basic

proteins

respectively

The

conserved

amino

acid

residues

among

the

proteins

are

marked

with

black

circles

Identical

amino

acids

are

boxed

b

Phylogenetic

analysis

of

rice

proteins

based

on

the

mature

proteins

using

UPGMA

method

The

predicted

isoelectric

point

-LRB-

pI

-RRB-

for

each

mature

protein

is

shown

on

the

right

Acidic

basic

and

neutral

proteins

are

shown

in

red

blue

and

green

respectively

Refer

to

the

description

in

a

for

details

Using

the

data

of

Fig.

a

phylogenetic

tree

was

constructed

-LRB-

Fig.

-RRB-

Among

the

proteins

acidic

and

basic

protein

members

were

found

-LRB-

Fig.

-RRB-

The

C-terminal

region

of

was

indicated

to

be

neutral

The

proteins

from

Arabidopsis

and

tobacco

grouped

in

one

clade

-LRB-

framed

-RRB-

and

proteins

belonged

to

different

clades

Responses

of

genes

to

blast-fungus

infection

in

young

rice

plants

Compatible

interaction

The

response

of

individual

genes

to

blast-fungus

infection

was

analyzed

by

qPCR

using

a

set

of

primers

specific

for

each

gene

-LRB-

Table

-RRB-

A

conidia

suspension

of

M.

grisea

race

was

sprayed

on

the

seedlings

of

Nipponbare

at

the

four-leaf

stage

In

this

system

the

fungus

infection

results

in

the

formation

of

whitish

expanding

lesions

at

days

post

inoculation

-LRB-

dpi

-RRB-

and

increasing

lesion

size

thereafter

with

conidia

formation

The

infected

plants

were

severely

wilted

and

died

within

days

indicating

a

typical

compatible

host

parasite

interaction

-LRB-

Sasaki

et

al.

Iwai

et

al.

-RRB-

Using

triplicate

cDNA

samples

originating

from

independently

prepared

RNA

templates

from

inoculated

fourth

leaves

the

expression

levels

of

each

gene

were

compared

with

that

of

an

actin

gene

which

was

selected

as

a

control

gene

Although

expression

levels

of

genes

in

healthy

Nipponbare

leaves

varied

according

to

each

gene

-LRB-

dpi

white

column

in

Fig.

-RRB-

all

genes

tested

were

up-regulated

by

blast-fungus

inoculation

at

and/or

dpi

-LRB-

black

column

-RRB-

When

compared

with

mock-inoculation

-LRB-

gray

column

-RRB-

the

enhanced

expression

by

the

fungus

was

significant

for

each

gene

at

dpi

at

which

lesions

were

vigorously

expanding

The

transcripts

of

and

had

already

accumulated

to

near

maximal

levels

at

dpi

maintaining

almost

the

same

levels

at

dpi

indicating

they

are

rapid

response

genes

to

blast

fungus

infection

However

the

transcripts

of

and

had

not

increased

significantly

at

dpi

compared

with

mock-inoculated

controls

indicating

these

are

late

response

genes

Blast

fungus

infection

induced

around

transcript

accumulation

at

dpi

compared

with

dpi

in

genes

and

Table

for

qPCR

of

and

actin

genesNameForward

primerReverse

074GTATGCTATGCTACGTGTTTATGCGCAAATACGGCTGACAGTACAGOsPR1

011ACGCCTTCACGGTCCATACAAACAGAAAGAAACAGAGGGAGTACOsPR1

071CTTTAACTATGTATGGAGTATGATATAAATGTGTTATTTTCTTCTTTTATTCGAACGACAACOsPR1

012CGCTGTGTGTTTGTGTTATGTCCGTGGTTTTGTCTTTATTTCAATCCOsPR1

073TTATATATGTATGTTCGTATGTATGTATGCTGATGTACTTATTCCATCCGACACOsPR1

021CGCAGCAACCAACCAATCTTGACAGTTGTAGTACTCTTGTAACATCATCOsPR1

022CCACAGAGTTTGTCAGGATTGTCCAGATTGCACACACCTGATTCCOsPR1

052AGCTACCTGTCATTTCTTCATTTCTGCTACTCCAGAAGGAAATTAAAAGOsPR1

072AATTAATACTGGAGTAGATGCATGTACACGAATAACGTACTGTATTCTGTATGOsPR1

121ACCATCGTCGTCGTCTCATCAGCCTCTAGGGCATATCACTAACOsPR1

101TCGCTGCCGCTAGTACATTTCATTAAGATCATTACATGCTTTATTGTTCACOsPR1

051CCTGCCTGCCTTCCTCATTCAGTGAAGATTTGGTTTCCATTGTATTGActinGAGTATGATGAGTCGGGTCCAGACACCAACAATCCCAAACAGAGFig

of

genes

to

pathogen

infection

a

Blast

fungus

infection

A

conidia

suspension

of

blast

fungus

race

was

spray-inoculated

onto

Nipponbare

-LRB-

compatible

upper

panel

-RRB-

and

-LRB-

incompatible

lower

panel

-RRB-

The

inoculated

fourth

leaves

were

harvested

at

and

days

post

inoculation

-LRB-

dpi

-RRB-

for

RNA

extraction

To

semi-quantify

the

transcript

level

RNA

samples

were

subjected

to

qPCR

and

the

relative

expression

levels

compared

with

that

of

a

control

actin

gene

are

shown

The

transcript

levels

at

time

mock-inoculated

and

blast-fungus

inoculated

leaves

were

shown

as

white

gray

and

black

columns

respectively

Bars

indicate

mean

SD

The

up-regulated

genes

by

blast

fungus-infection

are

marked

with

arrowheads

Gray

arrowheads

in

the

lower

panel

indicate

early

responsive

genes

b

Bacterial

blight

infection

The

fourth

leaves

of

Nipponbare

rice

plants

were

inoculated

with

a

suspension

of

Xoo

for

bacterial

blight

disease

and

the

leaves

were

harvested

at

and

dpi

for

RNA

extraction

The

transcript

level

was

analyzed

by

qPCR

The

relative

expression

levels

compared

with

that

of

the

control

actin

gene

are

shown

Transcript

levels

at

time

mock-inoculated

and

bacteria-blight

inoculated

leaves

are

shown

as

white

gray

and

black

columns

respectively

Xoo-upregulated

and

downregulated

genes

are

marked

with

forward

and

reverse

arrowheads

respectively

Bars

indicate

mean

SD

Incompatible

interaction

is

a

near

isogenic

line

of

Nipponbare

into

which

the

resistance

gene

Pi-i

for

blast

fungus

race

has

been

introduced

In

fungus-inoculated

small

HR

lesions

were

induced

at

dpi

with

no

remarkable

fungal

development

thereafter

indicating

a

typical

incompatible

interaction

-LRB-

Sasaki

et

al.

Iwai

et

al.

-RRB-

The

transcript

levels

of

and

were

maximized

at

dpi

and

slightly

reduced

at

dpi

-LRB-

Fig.

lower

part

gray

arrowheads

-RRB-

indicating

they

are

early

responsive

genes

to

blast

fungus

infection

The

genes

had

been

recognized

as

early

responsive

genes

in

compatible

interaction

The

levels

of

and

transcripts

increased

by

about

at

dpi

The

transcripts

of

and

were

higher

at

at

dpi

than

at

dpi

indicating

they

are

late

responsive

genes

-LRB-

black

arrowheads

-RRB-

The

level

of

was

suppressed

by

mock-inoculation

but

the

level

was

not

changed

by

incompatible

interaction

whose

transcript

increased

in

the

compatible

interaction

at

dpi

was

not

clearly

upregulated

in

the

incompatible

interaction

Responses

of

genes

to

bacterial

blight

infection

in

young

rice

plants

To

study

the

response

of

the

genes

to

a

bacterial

pathogen

the

fourth

leaves

of

Nipponbare

were

inoculated

with

Xoo

for

leaf

blight

disease

In

the

compatible

rice

bacterium

interaction

the

infected

region

was

visualized

as

wilted

necrosis

which

spread

from

the

cross

cut

end

to

the

leaf

base

with

time

and

the

area

of

necrosis

grew

to

cm

long

at

dpi

At

and

dpi

cm

long

leaf

pieces

from

the

inoculated

point

were

used

to

prepare

RNA

samples

Transcripts

of

and

which

are

early

responsive

genes

to

blast

fungus

-LRB-

Fig.

-RRB-

accumulated

remarkably

at

both

and

dpi

compared

with

dpi

or

a

mock-inoculated

control

-LRB-

Fig.

-RRB-

indicating

they

were

responsive

to

Xoo

infection

-LRB-

closed

forward

arrowheads

-RRB-

Expression

of

and

were

slightly

suppressed

after

the

infection

compared

with

mock-inoculated

leaves

-LRB-

closed

reverse

arrowheads

-RRB-

Expression

of

and

were

upregulated

by

Xoo

infection

but

other

genes

did

not

respond

or

rather

suppressed

while

all

genes

were

upregulated

in

compatible

blast

fungus

interaction

-LRB-

Fig.

-RRB-

Wound-induced

expression

of

genes

in

young

rice

plants

To

study

the

response

of

genes

to

wounding

the

fourth

leaves

were

wounded

by

cross

cutting

Triplicate

samples

were

extracted

at

and

h

after

wounding

and

used

for

qPCR

Among

the

genes

was

wound-inducible

with

peak

accumulation

of

the

transcript

at

h

after

wounding

and

was

found

to

be

an

early

wound-responsive

gene

-LRB-

Fig.

forward

arrowheads

-RRB-

The

and

transcripts

decreased

in

response

to

wounding

-LRB-

reverse

arrowheads

-RRB-

-LRB-

Fig.

-RRB-

Fig.

of

genes

to

wounding

The

fourth

leaves

were

cut

into

cm

length

sections

for

wounding

and

floated

on

water

for

and

h

under

light

and

then

the

leaf

pieces

were

homogenized

for

RNA

extraction

Relative

expression

levels

compared

with

that

of

the

control

actin

gene

are

shown

The

wound-upregulated

and

downregulated

genes

are

marked

with

forward

or

reverse

arrowheads

respectively

Bars

indicate

mean

SDFig

expression

of

genes

Leaves

and

roots

of

the

seedlings

at

the

fourth

leaf

stage

-LRB-

left

panel

-RRB-

and

flowers

of

adult

plants

-LRB-

right

panel

-RRB-

were

subjected

to

qPCR

analysis

Relative

expression

levels

compared

with

that

of

the

control

actin

gene

are

shown

The

genes

that

express

at

a

high

level

in

roots

and

flowers

are

marked

with

white

and

gray

arrowheads

respectively

Bars

indicate

mean

SD

Organ

specific

expression

of

genes

The

basal

transcript

levels

in

various

organs

varied

according

to

each

gene

in

the

roots

of

healthy

young

rice

plants

at

the

stage

the

transcript

levels

of

the

genes

and

were

clearly

higher

than

in

the

fourth

leaves

-LRB-

Fig.

-RRB-

-LRB-

open

arrowheads

-RRB-

In

flowers

the

basal

transcript

levels

of

five

genes

and

were

at

a

high

level

-LRB-

gray

arrowheads

-RRB-

These

results

indicate

that

the

organ-specific

expression

profiles

of

genes

varied

considerably

The

and

which

are

located

on

chromosome

were

highly

expressed

in

healthy

roots

In

healthy

flowers

the

expression

levels

of

five

genes

OsPR

and

were

also

higher

Responses

of

genes

to

defense

signal

compounds

in

young

rice

plants

The

responses

of

genes

to

defense

signal

compounds

were

studied

Intact

young

rice

plants

in

soil

pots

were

dipped

in

water

SA

or

ACC

solutions

or

exposed

to

the

vapor

of

MeJA

RNA

from

the

fourth

leaves

was

subjected

to

qPCR

-LRB-

Fig.

-RRB-

and

were

upregulated

by

all

three

signal

compounds

tested

while

and

were

upregulated

by

JA

but

not

by

ACC

or

SA

and

which

were

early

responsive

genes

to

blast

fungus

-LRB-

Fig.

-RRB-

were

SA-inducible

was

upregulated

by

SA

but

suppressed

by

ACC

Among

the

six

genes

that

were

inducible

by

the

wound

signal

compound

JA

and

were

also

upregulated

by

wounding

-LRB-

Fig.

-RRB-

and

were

not

particularly

sensitive

to

any

of

these

compounds

and

were

downregulated

by

ACC

and

SA

respectively

and

was

downregulated

by

both

ACC

and

SA

Fig.

of

genes

to

defense-signal

compounds

ACC

SA

and

JA

Expression

levels

of

each

gene

in

fourth

leaves

at

h

after

treatment

with

defense-signal

compounds

were

analyzed

by

qPCR

Relative

expression

levels

compared

with

that

of

the

control

actin

gene

are

shown

The

ACC

SA

and

JA-inducible

genes

are

marked

with

gray

dark

gray

and

black

arrowheads

respectively

For

ACC

or

SA

treatment

plastic

pots

with

rice

seedlings

were

dipped

in

solutions

containing

mM

ACC

or

mM

SA

solution

and

incubated

for

h

in

the

greenhouse

control

for

ACC

and

SA-treatment

-LRB-

leaves

of

water

treated

plant

-RRB-

For

JA

treatment

pots

with

seedlings

each

was

put

in

an

air-tight

clear

plastic

box

and

a

cotton

pad

with

volatile

methyl

jasmonate

-LRB-

MeJA

-RRB-

dissolved

in

EtOH

was

put

at

the

corner

of

the

box

to

give

a

final

concentration

of

μM

and

incubated

for

h.

control

for

JA-treatment

-LRB-

leaves

of

plant

treated

with

μM

of

JA

-RRB-

bars

indicate

mean

SD

Comparison

of

the

defense-signaling

pathways

conferred

by

ACC

SA

and

JA

in

rice

with

those

in

tobacco

and

Arabidopsis

Using

the

genes

as

probes

we

analyzed

the

relationship

of

responsiveness

to

three

defense

signal

compounds

in

rice

In

tobacco

SA

signaling

was

antagonistic

to

JA

signaling

on

PR

gene

expression

Niki

et

al.

-LRB-

-RRB-

showed

that

the

expression

of

acidic

and

genes

was

upregulated

by

SA-treatment

and

it

was

suppressed

in

the

presence

of

JA

in

a

dose-dependent

manner

while

the

expression

of

basic

and

genes

was

upregulated

by

JA

and

suppressed

by

SA

Thomma

et

al.

-LRB-

-RRB-

reported

that

the

JA-signaling

pathway

is

different

from

the

SA-signaling

pathway

in

Arabidopsis

Glazebrook

et

al.

-LRB-

-RRB-

reported

that

the

expression

patterns

of

some

genes

revealed

mutual

inhibition

between

SA

and

JA-dependent

signaling

using

global

expression

phenotyping

analysis

of

mutant

Arabidopsis

plants

Synergistic

induction

by

ET

and

JA

was

reported

in

basic

tobacco

PR

genes

-LRB-

Xu

et

al.

-RRB-

However

the

expression

patterns

of

genes

in

Fig.

showed

no

clear

mutual

inhibition

between

SA

and

JA-dependent

signaling

except

for

We

further

analyzed

the

relationship

by

means

of

Pearson

's

correlation

coefficient

-LRB-

Fig.

-RRB-

Correlation

coefficients

for

SA

versus

ACC

ACC

versus

JA

and

SA

versus

JA

were

and

respectively

The

linear

correlation

coefficient

r

represents

the

strength

and

the

direction

of

the

linear

relationship

between

two

variables

A

correlation

of

means

an

increasing

linear

relationship

and

complete

correlation

while

means

a

decreasing

linear

relationship

Cohen

-LRB-

-RRB-

proposed

that

r

below

is

regarded

as

insubstantial

r

as

weak

r

as

medium

and

r

as

strong

correlation

While

levels

of

statistical

significance

at

and

-LRB-

n

-RRB-

are

and

respectively

The

r

values

in

Fig.

suggest

synergism

between

ACC

versus

SA

and

ACC

versus

JA

and

low

or

no

correlation

between

SA

versus

JA

rather

than

antagonism

Fig.

of

the

three

signaling

pathways

in

genes

Based

on

the

data

in

Fig.

the

response

of

young

rice

plants

to

ACC

SA

or

JA-treatments

was

compared

with

other

treatments

and

the

correlation

coefficients

between

the

responses

to

pairs

of

signals

were

calculated

See

the

text

for

details

Tissue

specific

expression

of

a

representative

gene

in

rice

plants

Analysis

of

the

tissue

specific

expression

of

rice

gene

by

means

of

histological

studies

has

not

been

performed

previously

To

study

the

tissue

specific

expression

profiles

of

a

representative

gene

we

selected

because

it

clearly

responded

to

both

compatible

and

incompatible

fungal

interactions

bacterial

bright

infection

wounding

and

the

defense

signal

compounds

The

promoter

sequence

of

was

obtained

by

PCR

amplification

from

the

Nipponbare

genome

The

promoter

sequence

containing

bp

was

inserted

into

the

vector

-LRB-

Kawahigashi

et

al.

-RRB-

to

construct

the

plasmid

GUS

which

contains

the

promoter

and

the

coding

region

for

amino

acids

of

the

N-terminus

of

the

protein

to

express

the

GUS

protein

effectively

as

a

translational

fusion

-LRB-

Fig.

-RRB-

The

GUS

gene

was

introduced

into

rice

plants

-LRB-

Oryza

sativa

cv

Nipponbare

-RRB-

Among

hygromycin-resistant

plants

regenerated

three

independent

lines

were

selected

as

representatives

exhibiting

a

negligible

level

of

GUS

activity

in

healthy

leaves

and

more

than

higher

levels

after

wounding

or

inoculation

with

a

conidia

suspension

of

M.

grisea

The

GUS

plants

of

the

second

or

third

generation

were

subjected

to

histological

GUS

analysis

using

X-gluc

as

the

substrate

Fig.

analysis

of

gene

expression

in

transgenic

rice

plants

a

Schematic

diagram

showing

the

construct

of

the

introduced

gene

The

promoter

sequence

of

was

used

b

Localized

GUS

staining

in

healthy

day-old

transgenic

rice

seedlings

-LRB-

a

e

-RRB-

Because

similar

expression

profiles

were

obtained

in

three

independent

transgenic

lines

the

data

from

a

representative

line

is

shown

f

unhulled

rice

c

GUS

staining

of

month-old

transgenic

rice

plants

a

Healthy

roots

b

roots

at

days

after

wounding

c

e

leaves

at

days

after

wounding

d

GUS

staining

assay

of

leaves

at

-LRB-

a

c

d

-RRB-

or

-LRB-

b

-RRB-

dpi

Black

arrows

indicate

stained

mesophyll

tissue

Brown

arrows

indicate

the

necrotic

regions

Figure

shows

the

localization

of

GUS

activity

in

healthy

transgenic

rice

seedlings

A

high

level

of

GUS

activity

was

detected

in

the

primary

root

-LRB-

arrows

Fig.

a

c

-RRB-

and

the

root

tip

-LRB-

arrowheads

-RRB-

but

it

was

absent

or

considerably

reduced

in

the

elongation

zone

-LRB-

white

arrowheads

-RRB-

In

cross

sections

of

the

middle

part

of

the

root

μm

in

thickness

-LRB-

Fig.

e

-RRB-

a

strong

GUS

activity

was

detected

in

the

vascular

cylinder

-LRB-

vc

-RRB-

and

sclerenchyma

-LRB-

sc

-RRB-

and

a

weak

activity

in

the

cortex

-LRB-

co

-RRB-

GUS

staining

was

found

in

the

slit

surface

of

the

coleoptile

-LRB-

yellow

arrowheads

Fig.

a

b

-RRB-

and

on

the

surface

of

unhulled

rice

-LRB-

white

arrow

Fig.

a

f

-RRB-

In

the

anther

a

weak

blue

staining

was

found

in

the

matrix

of

pollen

loculus

-LRB-

data

not

shown

-RRB-

In

the

vector

control

plants

no

GUS

activity

was

detected

in

roots

-LRB-

Fig.

d

-RRB-

or

any

other

part

analyzed

under

these

in

condition

-LRB-

data

not

shown

-RRB-

In

month-old

GUS

plants

no

GUS

activity

was

found

in

healthy

roots

-LRB-

Fig.

a

-RRB-

while

high

levels

of

GUS

activity

were

found

in

healthy

day-old

-LRB-

Fig.

b

and

c

-RRB-

and

week-old

-LRB-

data

not

shown

-RRB-

plants

Thus

expression

in

roots

is

likely

to

be

regulated

developmentally

Wounding

induced

a

high

level

of

GUS

activity

in

the

roots

of

plants

When

the

aerial

parts

were

removed

by

cross

cut

from

of

the

plants

a

strong

GUS

activity

was

induced

in

the

roots

of

cut

plants

at

days

after

cutting

-LRB-

Fig.

b

-RRB-

In

cross

sections

of

the

root

in

the

middle

region

GUS

staining

was

found

in

all

tissues

including

the

vascular

cylinder

cortex

and

sclerenchyma

No

detectable

GUS

activity

was

found

in

healthy

leaves

of

transgenic

rice

plants

-LRB-

data

not

shown

-RRB-

however

wounding

induced

a

strong

GUS

activity

in

the

neighbor

region

of

the

cut

surface

especially

in

the

vascular

systems

including

xylem

parenchyma

-LRB-

xy

-RRB-

and

phloem

-LRB-

ph

-RRB-

-LRB-

Fig.

c

e

-RRB-

Next

GUS

activity

in

rice

leaves

infected

with

blast

fungus

was

analyzed

In

the

compatible

cultivar

-LRB-

WT

-RRB-

lacking

the

Pi-iresistance

gene

enlarged

lesions

were

found

days

after

inoculation

developing

in

size

with

time

and

with

conidia

formation

on

the

infected

leaf

surface

After

GUS

staining

many

blue

spots

were

found

at

or

around

the

lesions

in

the

leaves

days

after

inoculation

-LRB-

Fig.

a

-RRB-

When

the

leaf

of

Fig.

a

was

cross

cut

at

the

marked

line

and

shown

in

Fig.

c

the

blue

staining

was

found

to

localize

in

the

mesophyll

cells

-LRB-

black

arrows

-RRB-

around

the

necrotic

regions

-LRB-

brown

arrows

Fig.

d

-RRB-

At

days

after

inoculation

GUS

activity

was

found

around

the

necrotic

lesions

-LRB-

Fig.

b

-RRB-

Discussion

We

studied

here

the

response

of

genes

to

pathogens

wounding

and

defense

signal

compounds

and

their

constitutive

expression

in

organs

This

is

the

first

example

of

a

comparative

study

on

the

characterization

of

a

set

of

PR

family

genes

by

qPCR

a

high

throughput

method

The

genes

include

and

b

which

have

been

reported

as

pathogen-inducible

genes

by

Agrawal

et

al.

-LRB-

-RRB-

and

ten

of

the

genes

are

contained

in

the

listed

rice

genes

including

at

least

pseudogenes

whose

expression

profiles

were

not

obvious

-LRB-

van

Loon

et

al.

-RRB-

We

searched

rice

genes

from

full-length

cDNA

-LRB-

-RRB-

and

EST

-LRB-

-RRB-

Because

strongly

expressed

genes

should

be

easily

extracted

from

the

databases

for

expressed

genes

a

majority

of

highly

expressed

rice

genes

are

thought

to

be

contained

in

the

genes

The

sequences

of

the

proteins

were

well

conserved

with

complete

matches

in

amino

acid

residues

in

the

C-terminal

half

-LRB-

Fig.

-RRB-

Because

the

residues

were

also

very

well

conserved

in

all

proteins

selected

from

different

plant

species

including

Brassica

napus

Hordeum

vulgare

Lycopersicon

esculentum

Medicago

trunculata

Triticum

aestivum

and

Zea

mays

-LRB-

van

Loon

and

van

Strien

-RRB-

these

regions

in

the

C-terminus

should

be

important

for

the

putative

functions

of

proteins

in

plants

All

genes

selected

here

were

clearly

upregulated

in

a

compatible

rice-blast

fungus

interaction

indicating

that

they

really

are

pathogen

related

Notable

is

that

the

induction

pattern

of

each

gene

by

different

pathogen

host

interactions

was

characteristic

-LRB-

Fig.

-RRB-

Four

genes

and

responded

positively

to

blast

fungus

infection

in

both

compatible

and

incompatible

interactions

and

also

to

Xoo

infection

Among

these

and

were

responsive

to

blast

fungus

in

the

early

infection

period

in

both

interactions

Conversely

the

expression

of

and

was

reduced

by

Xoo

infection

compared

with

the

control

after

mock-inoculation

which

is

a

wound

treatment

Assuming

that

Xoo

infection

may

inhibit

wound-induced

expression

of

these

genes

it

is

consistent

that

they

are

wound-inducible

-LRB-

Fig.

-RRB-

The

above

result

that

all

are

characteristically

pathogen

inducible

is

apparently

different

from

the

data

of

Arabidopsis

where

only

one

gene

was

pathogen-inducible

-LRB-

-RRB-

among

predicted

genes

-LRB-

van

Loon

et

al.

-RRB-

We

studied

this

result

using

an

AtGenExpress

Visualization

Tool

from

and

confirmed

that

only

the

gene

was

upregulated

by

infection

with

either

Pseudomonas

syringae

pv

tomato

or

Pytophythora

infestans

and

the

other

genes

were

not

The

reason

for

different

responses

of

to

pathogen

infection

in

rice

and

Arabidopsis

is

not

clear

but

we

could

speculate

that

have

evolved

in

the

rice

genome

to

provide

resistance

to

a

broad

range

of

pathogens

but

a

limited

set

of

has

evolved

to

respond

to

pathogens

In

rice

the

level

of

free

SA

in

young

healthy

rice

leaves

was

as

high

as

about

μg

per

g

fresh

leaf

-LRB-

Silverman

et

al.

-RRB-

but

it

was

only

about

ng

-LRB-

Seo

et

al.

-RRB-

or

ng

-LRB-

Silverman

et

al.

-RRB-

in

tobacco

leaves

and

about

ng

-LRB-

Park

et

al.

-RRB-

or

ng

-LRB-

our

unpublished

data

-RRB-

in

Arabidopsis

leaves

Thus

the

SA

content

in

rice

leaves

corresponds

to

to

of

that

in

tobacco

or

Arabidopsis

leaves

which

is

comparable

to

the

induced

SA

level

after

HR

lesion

formation

by

TMV-infection

Such

a

high

SA

level

in

rice

plants

could

affect

expression

after

pathogen

infection

treatment

with

SA

JA

or

ACC

and

wounding

Actually

the

responses

of

the

genes

to

defense

signal

compounds

indicate

that

the

signaling

pathways

in

rice

conferred

by

SA

and

JA

were

synergistic

as

well

as

that

by

ET

and

JA

and

SA

and

ET

-LRB-

Fig.

-RRB-

This

finding

is

considerably

different

from

the

antagonistic

relationship

of

SA

and

JA

signaling

found

in

tobacco

-LRB-

Niki

et

al.

-RRB-

and

Arabidopsis

-LRB-

Gupta

et

al.

-RRB-

In

tobacco

acidic

proteins

such

as

b

and

c

were

inducible

by

SA

-LRB-

Ohashi

and

Matsuoka

-RRB-

which

is

a

defense

signal

compound

for

systemic

acquired

resistance

The

-LRB-

Eyal

et

al.

Niki

et

al.

-RRB-

and

-LRB-

Santamaria

et

al.

-RRB-

genes

which

encode

basic

PR

proteins

in

tobacco

and

Arabidopsis

respectively

were

reported

to

respond

positively

to

JA

and

ET

and

negatively

to

SA

The

gene

-LRB-

-RRB-

encodes

a

basic

protein

that

is

inducible

by

SA

To

compare

the

natures

of

the

genes

the

results

of

the

expression

of

genes

are

summarized

in

Fig.

It

is

again

impressive

that

the

expression

upregulated

by

various

treatments

differed

considerably

depending

on

the

gene

and

the

transcript

levels

constitutively

found

in

leaf

root

and

flower

also

varied

according

to

the

genes

When

comparing

the

four

genes

and

which

are

clustered

in

a

locus

on

chromosome

we

found

the

following

common

characteristics

all

four

genes

-LRB-

-RRB-

encode

acidic

proteins

-LRB-

-RRB-

are

constitutively

expressed

in

healthy

roots

at

a

high

level

and

-LRB-

-RRB-

are

upregulated

by

blast

fungus-infection

In

spite

of

these

similarities

their

expression

profiles

after

certain

treatments

were

significantly

different

from

each

other

For

example

-LRB-

-RRB-

the

transcript

of

accumulated

at

an

earlier

time

period

than

those

of

or

in

fungus-infected

resistant

cultivar

-LRB-

-RRB-

but

not

or

was

upregulated

by

Xoo-infection

-LRB-

-RRB-

but

not

or

was

wound-inducible

-LRB-

-RRB-

was

ACC

SA

and

JA-inducible

was

JA-inducible

was

not

inducible

by

any

signal

compounds

used

here

and

appeared

to

be

suppressed

by

ACC

-LRB-

-RRB-

was

expressed

in

flowers

at

a

high

level

but

and

were

not

Fig.

of

expression

characteristics

of

genes

Induction

levels

of

genes

by

pathogens

wounding

and

defense

signal

compounds

and

their

constitutive

expression

levels

in

organs

were

compared

The

transcript

levels

in

healthy

leaves

are

shown

in

the

right

most

column

very

highly

inducible

or

expressed

gene

highly

inducible

or

expressed

gene

gene

whose

induction

or

expression

was

not

clear

not

inducible

or

expressed

gene

From

the

expression

characteristics

of

the

genes

obtained

here

the

possible

contribution

of

all

genes

for

plant

self-defense

against

pathogen

attack

was

indicated

Although

the

functions

of

genes

are

not

fully

understood

proteins

in

some

plant

species

have

been

reported

to

have

antifungal

activity

including

tobacco

acidic

overexpression

that

induced

increased

tolerance

to

two

oomycete

pathogens

in

tobacco

-LRB-

Alexander

et

al.

-RRB-

and

basic

proteins

of

tobacco

and

tomato

which

have

an

antimicrobial

activity

against

Phytophythora

infestans

-LRB-

Niederman

et

al.

-RRB-

Thus

the

redundant

expression

of

the

genes

may

effectively

contribute

to

defend

the

attack

of

various

pathogens

The

expression

levels

of

the

genes

in

healthy

tissues

and

wounded

leaves

were

also

very

different

according

to

each

particular

gene

The

data

in

Fig.

shows

a

representative

example

of

tissue

and

organ-specific

expression

of

The

level

of

GUS

activity

in

transgenic

rice

plants

carrying

the

GUS

gene

was

confirmed

to

be

similar

to

the

transcript

level

as

shown

in

tobacco

GUS

plants

-LRB-

Yamakawa

et

al.

-RRB-

in

which

the

GUS

gene

was

expressed

as

a

similar

translational

fusion

gene

described

in

Fig.

GUS

analysis

using

independent

transgenic

lines

showed

expression

was

developmentally

regulated

in

roots

and

constitutively

expressed

in

young

rice

plants

but

not

in

old

plants

in

which

the

expression

was

strongly

upregulated

by

wounding

-LRB-

Fig.

B

-RRB-

The

wound-induced

expression

in

adult

plants

was

localized

mainly

in

the

vascular

system

which

is

important

as

the

sensing

tissue

of

the

water

pressure

change

after

wounding

and

as

the

site

of

bacterial

pathogen

propagation

The

information

on

the

genes

obtained

here

will

be

useful

for

further

studies

on

self-defense

mechanisms

in

rice

plants

and

for

the

usage

of

each

gene

as

a

characteristic

marker

gene

O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O B O O O O O O O O B O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O O O O O B O O O O O O O O O B I O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B I O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O

bacterial blight
histological analysis
pathogen resistance
rice


The Implications of Maternal Overweight and Obesity on the Course of Pregnancy and Birth Outcomes The increasing prevalence of overweight and obesity among women of childbearing age is a growing public health concern in the United States . The average body mass index -LRB- BMI -RRB- is increasing among all age categories and women enter pregnancy at higher weights . Women are also more likely to retain gestational weight with each pregnancy . Women who are overweight -LRB- BMI 25 -- 30 -RRB- and obese -LRB- BMI ≥ 30 -RRB- are at greater risk of adverse reproductive health outcomes compared to women of normal weight status -LRB- BMI 19.8 -- 25 -RRB- . This article provides an overview of the complications associated with maternal overweight and obesity including diabetes , pre-eclampsia , c-sections , and birth defects . We present updated information on the weight trends among women . Finally , we present an overview of the prevention studies aimed at adolescents and women prior to pregnancy . Introduction The effect of maternal nutritional status prior to pregnancy on birth outcomes is of great public health importance . Epidemiological studies have shown a clear association between maternal pregravid weight and birth outcomes . As a marker of nutritional status , a woman 's pregravid body mass index -LRB- BMI = kg/m2 -RRB- , if low -LRB- < 19.8 BMI -RRB- , may reflect chronic nutritional deficiency whereas a high BMI -LRB- > 26.1 BMI -RRB- reflects an imbalance between energy intake and expenditure , and thus varying degrees of adiposity -LSB- 1 -RSB- . The effects of each on birth outcomes differ , with low BMI being associated with intrauterine growth retardation -LRB- IUGR -RRB- , preterm birth , and iron deficiency anemia -LSB- 2 -- 5 -RSB- . Whereas , BMI above the normal range of 19.8 to 26.1 -LRB- IOM 1990 -RRB- is associated with a number of adverse reproductive health outcomes . For example , infertility -LSB- 6 -RSB- gestational diabetes -LSB- 7 -RSB- , pregnancy induced hypertension and pre-eclampsia -LSB- 8 -RSB- , birth defects -LSB- 9 -RSB- , large for gestational age -LRB- LGA -RRB- or macrosomia -LRB- > 4500 g -RRB- -LSB- 10 -RSB- , cesarean sections -LSB- 11 -- 13 -RSB- , prolonged labor -LSB- 14 -RSB- , and recently postpartum anemia -LSB- 15 -- 17 -RSB- have all been associated with maternal overweight yet the exact mechanisms have not been identified . The purpose of this paper is to describe an overview of the complications associated with maternal overweight and obesity ; present updated information on the weight trends among women ; and review prevention studies aimed at adolescents and women prior to pregnancy . Prevalence of overweight and obesity among women of childbearing age Until recently , most studies on the topic of maternal pregravid weight focused on the lower end of the weight spectrum ; however , with the shift in body weight that has occurred globally , more recent research has examined the effect of overweight and obesity on birth outcomes . Obesity world wide -LRB- > 30 BMI -RRB- now exists at a prevalence of 15 -- 20 % and accounts for 2 -- 7 % of the total health care costs -LSB- 18 -RSB- . In the US , the latest NHANES survey -LRB- 1999 -- 2002 -RRB- indicate that 26 % of non-pregnant women 20 -- 39 years of age are overweight -LRB- 25 -- 29.9 BMI -RRB- , and 29 % are obese -LRB- > 29.9 BMI -RRB- -LSB- 19 -RSB- . On average , obesity among all women appears to have peaked at 33 % with no appreciable increase between 1999 -- 2000 and 2003 -- 2004 -LSB- 20 -RSB- . Determinants of overweight among women A higher proportion of women of child bearing age are overweight or obese compared to men , and women of younger or older ages . In addition , upwards of 80 % of African American women are either overweight or obese -LSB- 19 -RSB- . Using 2003 -- 2004 data from NHANES , non-Hispanic black women are twice as likely to be obese than their non-Hispanic white counterparts -LRB- OR 2.01 , 95 % CI , 1.76 , 2.29 -RRB- -LSB- 20 -RSB- . Among individuals with less than a high school education , the prevalence of obesity was roughly twice that of college graduates -LSB- 21 -RSB- . Paradoxically , low-income households appear to have a higher risk of obesity -LSB- 22 -RSB- , although , at the same time they are more likely to be at risk of food insecurity . Household food insecurity has been associated with increased BMI -LSB- 23 -RSB- and an increased risk of overweight -LSB- 24 -RSB- in women , but not among men or children -LSB- 25 -- 27 -RSB- . Influences of adolescent overweight on reproductive health Moreover , thirty percent of non-pregnant adolescent girls 12 -- 19 years of age are considered overweight or at risk , based on a BMI for age at the 85th percentile or higher -LSB- 19 -RSB- . Excessive weight gain at younger ages is associated with earlier menarche , especially among black and Hispanic girls . Younger and more severely obese girls reaching reproductive capacity perpetuate obesity if left untreated , directly through the influence of maternal weight on fetal origins and indirectly through maternal to child social and developmental interactions -LSB- 28 -RSB- . Pregravid BMI is a predictor of gestational weight gain Among women who become pregnant , the shift towards higher pregravid weight also appears evident -LSB- 29 -RSB- . One study using a perinatal data base to ascertain weight before pregnancy for a 19 year period showed that the mean maternal weight of women at their first prenatal visit increased by 20 % between 1980 and 1999 , and the percentage of women weighing > 200 lbs at that visit increased from 7.3 to 24 % and for those weighing > 300 lbs the percentage increased from 2 to 11 % -LSB- 30 -RSB- . Another study using a perinatal data system of all live births from 8 contiguous counties in New York , an 11 % increase in pregravid overweight and an 8 % increase in pregravid obesity between 1999 and 2003 -LSB- 29 -RSB- . Not only are more women beginning pregnancy at a higher BMI , but women are also gaining in excess of the 1990 Institute of Medicine recommendation for gestational weight gain -LSB- 1 -RSB- . Specifically , and overweight and obese women are more likely to gain excessive gestational weight and keep it on after delivery -LSB- 31 -- 33 -RSB- . In affluent countries , women retain some weight with each successive pregnancy , gaining more weight than their non-pregnant counterparts -LSB- 34 , 35 -RSB- . These observations beg the far larger and more important question of how and when to intervene in order to optimize reproductive and individual health ? Weight intervention studies Many research studies of dietary , physical activity and weight interventions focus on individuals who are already obese or experiencing co-morbidities such as heart disease , diabetes or cancer . In contrast , only a limited number of interventions have focused on weight loss and/or maintenance of optimal weight in the context of reproductive health . On one end of the spectrum , surgical interventions such as gastric bypass and lap-band surgeries have been successful among obese women of child-bearing age with a decrease of reported rates of gestational diabetes , macrosomia , cesarean section while achieving adequate weight gain -LSB- 36 , 37 -RSB- . Some interventions have been targeted to address obesity during adolescents , pregnancy , and the postpartum or inter-conceptional periods . These commonly have been school - or clinic-based in nature . School-based randomized intervention trails such as the Trial of Activity in Adolescent Girls -LRB- TAAG -RRB- are presently underway to determine if increased opportunities for moderate and vigorous activity may decelerate the age related decline in physical activity -LSB- 38 -RSB- . Clinic-based interventions Clinic-based intervention usually begin with obesity management training for physicians because of the existence of a combination of a paucity of nutrition , physical and weight educational materials , lack of knowledge of how to counsel women , and lack of resources and personnel to assist with obesity management . For example , results from a survey of pediatricians found that only 12 % of pediatricians reported high self-efficacy in obesity management , although 39 % stated that physicians were important health care providers that could be effective with obesity management . Lack of non-MD staff reimbursement , of an on-site dietitian , and of patient educational materials were each highly associated with low self-efficacy in obesity management -LSB- 39 -RSB- . Weight loss interventions Tsai and Wadden -LSB- 40 -RSB- conducted a systematic review of major commercial weight loss programs undertaken to provide physicians with information on components , cost , and efficacy of these programs . eDiets.com , Health Management Resources , Take off Pounds Sensibly , OPTIFAST , and Weight Watchers were represented in the academic literature . The review identified Weight Watchers as the least costly with maintenance of 3.2 % loss of initial weight at two years . Medically supervised very-low-calorie diet programs had the greatest weight loss -LRB- approximately 15 to 25 % of initial weight -RRB- , but were associated with high costs and high attrition rates . Finally , Internet -LRB- i.e. , eDiets.com -RRB- and organized self-help programs -LRB- i.e. , Overeaters Anonymous -RRB- produced minimal weight loss . In a clinical intervention conducted among pregnant women , physicians were trained to chart and monitor gestational weight gain adequacy based on the Institute of Medicine weight gain recommendations . Women received five patient education newsletters by-mail with action-oriented messages of how to gain adequate weight in pregnancy . Findings indicated that low-income women who received the intervention were less likely to gain excessive gestational weight . Overweight low-income women were at a reduced risk to retain more than 2.7 kg of weight at one year -LSB- 41 -RSB- . A hospital based randomized control trail with an intensive stepped-care , behavioral intervention found that the intervention significantly decreased the percentage of normal weight women gaining in excess of the IOM recommendations -LSB- 42 -RSB- . Significance of weight status during postpartum period The postpartum period is hypothesized to significantly contribute to overweight and obesity through pregnancy weight retention . Olson et al. -LSB- 31 -RSB- found that over 25 % of women participating in a cohort study experienced major weight gain , defined as 4.55 kg or more , at one year postpartum . Gestational weight gain , postpartum exercise frequency , and food intake were all significantly associated with weight change from early pregnancy to one year postpartum . Lower income women who gained more than the IOM recommendation were at increased risk for major weight gain at one year postpartum . In a randomized trial of 40 overweight postpartum women receiving either a 12 week structured or self-directed diet and physical activity intervention , the structured intervention group had significant weight loss -LRB- 7.3 kg -RRB- , decrease in percent body fat -LRB- 6 % -RRB- and no change in fat-free mass , whereas the self-directed group had no significant change at one year postpartum -LSB- 43 -RSB- . Randomized control trials are currently underway in low-income WIC populations receiving usual care or a multi-component intervention including home visits , group classes and monthly telephone counseling -LSB- 44 -RSB- . Conclusion Given that the costs associated with treating obesity are so high and their limited success rates , the best strategy is to prevent obesity from occurring in the first place . However , prevention first requires a clear understanding of its etiology . Obesity is a chronic disease thought to develop from a complex interaction of genotype and environmental factors -LSB- 45 -RSB- . Our knowledge of how and why this disease develops is far from complete , but currently funded research projects are targeting the integration of social , behavioral , cultural , and biological factors . In addition , best practices at obesity prevention , treatment and optimal weight maintenance must be identified to provide practitioners with an array of strategies to help curb the ensuing epidemic .

O O O O O O B O O O O B O O O O O O O O O B O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O B O O O O B O O O O O O O O O O B O B O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O B O O B O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O B O O O O O O O O O B O O B O O O O O B O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O B O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O B O B O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O B O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O B O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O B O O O O O B O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O B O O B O O O O O O O O O B O B O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O B O O O O O B O O O O O O O O O O O O O O O B O O O B O O O B O O O O O O O O O O O O O O O O O O B O O O O O O O O B B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B O O O O O O O O O O O O O O O O O O O O O O O O O

obesity nutrition women pregnancy postpartum