Response of Microbial Populations to Carbofuran in Soils Enhanced

1.4 (x). Plate and. CO2. 8. Carbofuran. INC*. 10-100 (x) Plate. 9. EPTC*. (6 mg) ..... t h e r e m a i n d e r , 6% was i n c o r p o r a t e d i n t ...
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Chapter 12

Response of Microbial Populations to Carbofuran in Soils Enhanced for Its Degradation 1

2

R. F. Turco and A. E. Konopka 1

Department of Agronomy, Purdue University, West Lafayette, IN 47907 Department of Biological Sciences, Purdue University, West Lafayette, IN 47907

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2

The size of the microbial populations able to rapidly degrade carbofuran in soils enhanced for its degradation were enumerated by means of substrate addition and fumigation. Use of these techniques followed unsuccessful attempts to enumerate the population using plate or direct counts in the enhanced soils. Overall biomass size declined following application of carbofuran. No biomass suppression was observed in the non-enhanced soils and implies this suppression may be related to the formation of metabolites such as carbofuran-phenol or methylamine. In the enhanced soils, 6% of applied pesticide was i n i t i a l l y incorporated into biomass carbon. This contrasts with 0.87% incorporation in the non-enhanced soils. After 15 days there was complete loss of the pesticide; at this time the biomass contained 2% of the applied material.

Enhanced

biodégradation

microbial

exploitation

soil

is

that

pesticide the

10

of

an

Fig. to

losses

days

reports

on

the

(5).

occur

soil

The a

the

to

the

multistage of

size

of

the

changes

degrading

microorganisms

occur work

during has

of

degradation

impact in

the

focused

period on

than

75% o f

of

the

the

the as

estimated

kinetics

2).

by

plate

from

shown

in

postulated efforts

centered or

to or on

pesticide-

counts

population

or

changes

degradation,

pesticide

within

population,

have

a

where

response

is is

Most

total

carbofuran of

typical

microbial

in

derived

carbofuran

of

the

was

A

biomass,

types

described

frame

pesticide

(Fig.

or

rapid

of

applied

enhanced

one

applied material

(1-4).

the

extreme recognize

c a r b o f u r a n as

time

application process

have

of the

This

c a r b o f u r a n on

of

We

carbofuran

numbers

Few e f f o r t s

an

material.

degradation

application.

following techniques.

the

loss

constitutes

soil-applied

greater

rapid

in

define

are

pesticides

a

for

following

enhanced 1

understand to

enhanced

of of

degradation

0097-6156/90/0426-0153$06.00/0 © 1990 American Chemical Society

Racke and Coats; Enhanced Biodegradation of Pesticides in the Environment ACS Symposium Series; American Chemical Society: Washington, DC, 1990.

MPN that

as

most

in

non-

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154

ENHANCED BIODEGRADATION O F PESTICIDES IN T H E ENVIRONMENT

Time (days) 1

Figure 1. Conversion of carbofuran (100 μg g" soil) in sterile ( ^ C O irradiated) and non-sterile soils. Each bar is one standard error. (Reprinted with permission from Ref. 5. Copyright 1990 Pergamon Press.)

Carbofuran Total C O 2 (sidechain)

? — —

Carbofuran-phenol

Soil A d s o r p t i o n

J

Subset

Biomass

( < 4 days )

Figure 2. Proposed pathway for the microbial conversion of carbofuran.

Racke and Coats; Enhanced Biodegradation of Pesticides in the Environment ACS Symposium Series; American Chemical Society: Washington, DC, 1990.

12.

Response of Microbial Populations to Carbofuran

T U R C O & KONOPKA

enhanced

soils

(Table

to

have

total

of

microbial

size

applications

work

that

(7)

but

carbofuran

to

In

a l l

cases

Table

the

cited

I.

c a r b o f u r a n to

above,

to

Attempts

not

were

made

1.4

Carbofuran

NE

Carbofuran

INC

Carbofuran

INC

Carbofuran

INC*

1.4

NE

. . .

enhanced

plate

and

counts.

organics Reference 31 32

^C-C02 L

Plate

and

33 18

Addition

Plate

7

Fumigation

6 34

Plate

10-100 (x) 10 1 1--1 10 0 (x)

(x)

(x)

(x)

repeat

showed

population.

8

Acids

=

non-target

(x)

NR

INC INC

or

earlier

that

Method

Slight

EPTC*

Phenolic

or

single

p o p u l a t i o n changes

Substrate

INC

INC

from

a p p l i c a t i o n of

(x)

Vedasan

mg)

on

(8)

considered

understand microbial

Carbofuran

(60

al.

actinomycetal

were

DEC

HCCH

effect

et

MPN

INC

mg)

and

to

showed types

work c o n f i r m e d

little

Mathur

10-1000 (x) 10-20

INC

Thiram

(6

(6) of

exposed

This

soils

from

Johnson

distribution

soils

have of

Magnitude

INC

2,4-D 2,4-D MCPA 2,4-D & Captan,

in

bacterial

the

and

the

estimates

soil

Effect

work

the

population

resulting Compound

on

pesticide.

contradicted stimulate

microbial

effect

populations

of

showed

organisms,

Duah-Yenturni

little

repeated

soil

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I).

carbofuran

application,

INC =

Plate Plate

1 4

Plate 1 4

9

C O 2

and

C-MPN 19

C-MPN

Substrate

17

Addition

16

Fumigation, Substrate

increase,

Addition

NE = no

effect,

DE =

decrease

Microbial We

are

Populations

developing

'enhanced' around has

two

Our

sets

agar,

with

initial media

Henrici

evident

of

changes

count

nitrogen

to

soils

following media

in

in

which We

these

we

two were

have

of

used

Use

failed

to

of

of

have

populations have

carbofuran

estimated

conducted soil

the

carbofuran

improve

microbial

in

centered

degradation

the

microbial

systems.

difference

treatment

Degradation

Our e f f o r t s

enhanced

carbofuran-agar Little

how

carbofuran.

(5).

studies

and

source).

carbofuran.

plating

understanding

respond

demonstrated

population plate

an

soils

been

and Enhanced C a r b o f u r a n

using

extract

(carbofuran in

total

enhanced as

recovery

sole of

plate

agar,

counts.

starch

as

carbon

microbial or

For

glycerol and/or

numbers

non-enhanced

carbon

degraders

source from

were soils

in

the

soil.

We

Racke and Coats; Enhanced Biodegradation of Pesticides in the Environment ACS Symposium Series; American Chemical Society: Washington, DC, 1990.

155

156

ENHANCED BIODEGRADATION O F PESTICIDES IN T H E ENVIRONMENT

have

isolated

containing have

been

period

unable

of

10

recoveries reported to

over

400

carbofuran. days

of

by

to

detect

or

longer.

pesticide

(9).

select We

cell in

adapted

numbers

following

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our

the

count

in

numbers

we

shown

to

side

application

soils

10

have

shown

by

that

C-CO2

different

for

10

weeks

the

enhanced

occurs

in

This easily

extract

Table

of

C O 2 the

II.Changes

used

have

time

the

poor

a

soil,

chemostat

isolated

is

four

5

this

not

days,

(5).

of

soils

4b).

labeled to

is

onto

was

followed

useable

is

five

days

of

incubation

of

not

earlier

soil

carbon

by

significantly as

indicated

minimal

the

during that

sidechain,

adsorption

surface soil

soil)

conclusion

of

as

soils

c a r b o f u r a n g"

removal

the

II),

previous

these

Degradation

our

shifts

unexpected

in

c a r b o f u r a n was

extended

total

or

our

within

μg

and

in

(Table

with

Subsequent

from

and

changes method

completely

occurs

(10)

changes

carbofuran

that

limited

Ramsey

the

consistent is

(Fig.

of

consistent with

of

evolved soils

(Fig.

of

the

3 and

5).

microorganisms

can

carbofuran.

in bacteria

^ g

this

3a,b)

This

amount

from

few

are

from r i n g

4a).

first

the

to

we

a

enhanced

have

study

4

(carbofuran phenol)

limits

similar

and

^ C - c a r b o f u r a n (100

mechanism

the

metabolite

The

to

observed

(Fig.

total

(Fig.

we

media

culture,

within

from

method

soil.

hydrolysis

evolution

the

are

work

counts

utilization

with

any

degradation

bacteria,

recovery

However,

chain

weeks

solid

liquid

bacteria

direct have

that

pesticide for

to

findings

recent

application,

findings.

has

limited

soil

orange

pesticide

work

in

different

strains.

directly

microbial

plate

Our

degrading

However,

acridine

from transfer

carbofuran

carbofuran-degrading

carbofuran-degrading utilized

colonies Following

numbers

in

soil

as

Carbofuran) g'

1

estimated Time

soil

0

AODC

(days) 5

log Soil

by

cell

10 g""^ s o i l



0

I

9, . 2 7

9.36

9 .41

II

9, . 2 8

9.60

9, . 7 7

III

9. .62

9.75

9, . 6 2

IV

9. . 4 4

9.42

9, . 1 4

9. . 4 6

100 I

9. .32

9.53

II

9. . 3 0

9.46

9, . 8 2

III

9. . 4 0

9.66

9, . 3 5

IV

8. ,97

9.25

8. .92

If

the

source,

a

6 μg of

N.

to

be

that

entire

100

However,

limited if

μ g g" to

the

carbofuran

structure

of

application the

nutrient

carbonyl is

c a r b o f u r a n was to

would

contribution

group

serving

soil

as

(11). a

usable supply of

as 65

nutrient

carbofuran

Our e s t i m a t e s

carbon

a

μg of

source,

have taking

Racke and Coats; Enhanced Biodegradation of Pesticides in the Environment ACS Symposium Series; American Chemical Society: Washington, DC, 1990.

C and

appears shown into

12.

TURCO & KONOPKA

Response of Microbial Populations to Carbofuran

100

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80+ 60 + 40 + 20 +

Time (days) 14

1

14

Figure 3. Evolution of C - C 0 from soils treated with 100 g" (a) C-carbonyl or (b) C-ring carbofuran. (Reprinted with permission from Ref. 5. Copyright 1990 Pergamon Press.) 2

14

Racke and Coats; Enhanced Biodegradation of Pesticides in the Environment ACS Symposium Series; American Chemical Society: Washington, DC, 1990.

157

ENHANCED BIODEGRADATION O F PESTICIDES IN T H E ENVIRONMENT

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158

carbofuran and incubated 10 weeks. (Adapted from Ref. 24.)

Racke and Coats; Enhanced Biodegradation of Pesticides in the Environment ACS Symposium Series; American Chemical Society: Washington, DC, 1990.

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12.

T U R C O & KONOPKA

Response of Microbial Populations to Carbofuran

100 CSS Unextractable

•i co

•1

ΓΛ

2

80 +



Extractable

60 + 40 +

^

!

20 +

JLn-l

1 II

Soil

III

14

IV

Figure 5. Percentage distribution of C-radioactivity in three fractions from four soils after treatment with C-ring carbofuran. (Reprinted with permission from Ref. 5. Copyright 1990 Pergamon Press.) 14

Racke and Coats; Enhanced Biodegradation of Pesticides in the Environment ACS Symposium Series; American Chemical Society: Washington, DC, 1990.

159

160

ENHANCED BIODEGRADATION O F PESTICIDES IN T H E ENVIRONMENT

account

loss

microbes

indicate

gr"

A

total the

5.8

The

10 that

of

can

crop,

has to

population. have

of

the

the

phenol

or

6c).

of

and

10

10^

cells

increase

in

elaborate is

to

in

the

estimate

the

system.

was but

needed

to

reported was

is

to

adsorption

Sparling

et

and

The

protects

the

where

been

an

substrate

addition

conducted

materials

applied

the

fungicides

was

respiratory on

microbial

application

rates

altered

population.

the

dominant

(5

μg

population

likely

a

of

day to

with

) The away

Ana Yeva

product

and

the

to

materials

suppression from

study

fungi

the

that

was

these

al.

(17) the

population.

the

effect

et

impact

coupled

towards

soil

addition

et

even

radically

of

to

utilized

the

Anderson

found

further

However,

formulated

They

degraded,

Following

in

method

soil

findings

had

as and

carbofuran

not

applied

changes

bipmass. g

were

They

to

by

importantly

the

the

response

twice

prevent

period.

differentiated.

been

/ig m l "

Kale

as

is

may

soil,

has

production

More

with

size.

the

20

be

a

(14)

inhibitory

phenol.

to

Using

of

to

diminishes and

the

within

carbofuran

carbofuran.

phenols

10

exposed

quantify

not

light

was

contrasts

prometrine

to

than

biomass

biomass

method

excess

the

uptake by

(carbofuran

methylamine

carbofuran-phenol

over

or in

of

biomass

to

that

respiration.

series

previously

increase

carrier

a

increased

hexachlorocyclohexane

was

most

soils.

in or

biomass

methylamine

phenol

microbial

biomass

nature.

in

soil

control

phosphoreum.

and

toxic

carbofuran

(16),

reported

work

less

repression

size

had not

50%

reported

This al.

biomass

suppress

soil

transient

adsorbed

its

that

and

phenol

the

g"

phenol

the

suppressive

trait.

the

metabolite

concentrations

they but

of

indicates

primary

Photobacterium

that

Moreover,

numbers

This a

of

the

to

/ig

coupled

carbofuran

response

the

carbofuran

reported

were

the

have

for

the

the

active

be

the

in

(13)

untreated

may

a

pesticide

maintain

the

by

that

for

A return

control

found.

may

of

a or

pesticide

changes

100

of

in

unit

however,

A

the

6a,b).

for

method

(5) .

size

this

period to

C

affected

to

lag

either

of

in of

less

clear,

crop

organic

enhanced

response

the

to (Fig.

one

is

standing

the

coupled

soils

untreated

developed

degradation.

the

x

Direct

x

Within

population

reduced

carbofuran phenol

effect

the

or

of

or

decreased

adsorption

soil

It

substrate

soils

carbofuran,

(15)

phenol

Their

to

approach

(12).

is

enhanced

the

toxic,

phosphoreum.

microbial

of

No d i f f e r e n c e

They

as

a

compared

methylamine)

carbofuran

of

to

chain

test

toxicity

soils

1.9

biomass

weight

more

system

the

non-enhanced

evaluated.

and

5.8

0.05%

failed

becomes

defined.

ability

carbofuran

not

microtox

but

is

than

2-3%

biomass

soil

in

in

the

are

a

been

as

as

that

results

the

non-enhanced

Raghu

have

Between

the

enhanced

soils

side

is

the

never

similar

(Fig.

toxic

size

less

microbial

defined

Why

applied

of

treated

itself

P.

is

about

mg""'" b i o m a s s ) .

A better

the

soil. with

biomass

enhanced size

is

members

an

and

We

application

soil

on

support

population

change

occurring.

I).

This

degradation

the

are

given

are

transition

of

a

(Table

degradation

a

would

bacteria

enumerations

associated

there

to

10**

average

carbofuran

in

be

pesticide

the

chain x

soil

biomass

cells

study

the

1

Estimations

microbial

soil

side

gr Simple

changes

living

the

that

x

influence

Biomass

the

(assumes

numbers.

subtle

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soil

counts .

CO2,

of

g"

(18)

of

three

very

low

suppressed

and

to

at

of

al.

a

shift

bacteria.

Racke and Coats; Enhanced Biodegradation of Pesticides in the Environment ACS Symposium Series; American Chemical Society: Washington, DC, 1990.

in

Duah-

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T U R C O & KONOPKA

Response of Microbial Populations to Carbofuran

80·60· 40·

Soil Soil Soil III Δ Soil IV A-

20-·

-Δ -A 100 /*g Carbofuran g ""1 9

Time (days)

10

Figure 6. Changes in total biomass size for Soils 11(a) and IV(b) following addition of 0, 10, or 100 μg carbofuran g" soil. Bar is on LSI (P) i s a l t e r e d to f o l l o w i n c o r p o r a t i o n o f radiolabel into the specialized portion of the population. F o l l o w i n g an i n c u b a t i o n w i t h the p e s t i c i d e , the s o i l s a r e f u m i g a t e d and the f r a c t i o n o f the biomass c o n t a i n i n g l a b e l i s d e t e r m i n e d . S o u l a s e t a l . (20) w o r k i n g w i t h 2,4-D r e p o r t e d t h a t 13.4% o f the a p p l i e d m a t e r i a l s were i n c o r p o r a t e d i n t o s o i l biomass. From an initial a p p l i c a t i o n r a t e o f 3.7 mg 2,4-D kg""'", the p e s t i c i d e s p e c i f i c biomass i n c r e a s e d i n s i z e to 0.372 mg kg" s o i l o r 3.7 x 10 microbe g soil. Kassim e t a l . (21) showed t h a t a f t e r 12 weeks, a r e a d i l y u s a b l e m a t e r i a l l i k e a c e t a t e would be t a k e n up i n t o as much as 70.4 % o f the biomass. However, a f t e r t h i s l e n g t h o f i n c u b a t i o n the l i k e l i h o o d t h a t o t h e r forms o f c a r b o n d e r i v e d from a c e t a t e are a c t u a l l y c y c l i n g i n the biomass i s g r e a t . We used a s p e c i f i c l a b e l l i n g approach to e s t i m a t e the s i z e o f the dégrader p o p u l a t i o n i n our enhanced soil that could use c a r b o f u r a n as a c a r b o n s o u r c e (24). Our system d i f f e r s somewhat from t h a t d e s c r i b e d by S o u l a s e t a l . (20) i n t h a t o n l y carbofuran s i d e - c h a i n c a r b o n appears to be a v a i l a b l e to the biomass. T h i s may r e f l e c t the f a c t t h a t c a r b o f u r a n i s s e r v i n g as b o t h a n i t r o g e n and c a r b o n source (11). Less than 3% o f the r i n g carbon, as compared to 74 % o f the s i d e c h a i n i s e v o l v e d as CO2 w i t h i n the 10 day time period. Because c a r b o f u r a n r i n g breakage i s so limited, only i n c o r p o r a t i o n o f c a r b o n y l - l a b e l e d C from c a r b o f u r a n was s t u d i e d . To i n s u r e maximal a c t i v i t y i n the p o p u l a t i o n , s o i l s I and I I were p r e t r e a t e d w i t h 25 μg c a r b o f u r a n g"^" s o i l and a d j u s t e d to -0.33 bar water c o n t e n t 10 days b e f o r e a p p l i c a t i o n o f the materials. A t o t a l o f 100 ug c a r b o f u r a n g soil was a p p l i e d and the soils i n c u b a t e d a t 25°C ( 2 4 ) . A t days 1, 5, 10 and 15, t h r e e s o i l samples were removed, f u m i g a t e d w i t h incubated f o r 24 h, and the f u m i g a n t removed w i t h e v a c u a t i o n . A t s a m p l i n g , the NaOH t r a p s i n a l l o t h e r samples were renewed. Fumigated samples were t r a n s f e r r e d

CHC3 I,

Racke and Coats; Enhanced Biodegradation of Pesticides in the Environment ACS Symposium Series; American Chemical Society: Washington, DC, 1990.

12. to

T U R C O & KONOPKA

Response ofMicrobial Populations to Carbofuran

2-1

the

glass

Traps

1

4

C O 2

using

evolved

C-C0

or

2

and

exchanged

calculated or

jars

were

the

in

the

Incorporation C

averaged was

recovered biomass

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by

the Cg""'"

day

addition chain. evolved

remainder,

of

1.02

x

been

lost

as

the

applied to

to

5.1

1,

x

likely

g"^

was

in

7).

Of

induced

(25) .

The

g"^

for

use

day

5,

65%

the

25

enzymes lag

of

the

the

of

Of

corresponds

the

pesticide only

This

5% C

of had

corresponds size

was

pre-application

was

needed

phase

from

biomass.

population

/ i g g"

significance

100

containing

initial

in

A

materials

contain

biomass

and

changes

This

to

In

5

(5).

C g"

the

size % of

to

applied

in

of

15.

1

the

/ig

the

declined

The

a

s

C-C

6.0

day

day

to

biomass.

the

to

0.87% of

was

% by

from

applied pesticide.

the

of

of

for

By

soil.

more

lack

2.0

10.28

17%

had

response

a

the

biomass

^" C-C

a d d i t i o n method

into

15,

the

to

the 4

of

dropped

supply

soil.

g"^

where

averaged

corresponds

/ig

day

2% o f

cells

elevated

population

By

NaOH.

w

^f

subtracting

Overall

substrate

incorporated

than

10

(Fig.

size

and biomass

in

C-CO2

s

Incorporation

diminishing

8.53

by

biomass

II.

approximately

cells

C O 2

carbofuran

the

This

the

leaving

10~*

less

1

(1984)

corrected

I.

soil

biomass

with

al.

Incorporation

day

materials.

declined

process

at

6% was

about

most

enhanced

et

trapped

Biomass

27.

carbofuran would

CO2,

as

is

to soil

(Fig. 7).

day

had

about

10

At

the

days

soil.

month.

Soulas

21

C-CO2

and

one

C-C into

total

observed

had to

the

mg

soils,

size

g

side

of

C-carbofuran

enhanced

14

non-enhanced

% in

0.77

applied

/ig

for

3.76

about

the

first

of

from day

2

applied

for

technique

C0 evolved

CC>2

evolved

weekly

the

fact

for

is

the

of

that

the

degradation

indicative

of

this

induction. Utilization first

5

days.

substrate loss

of

model

became the

the

the

five

limited.

predicted

short

Buildup

of

However,

the

the

estimated the

5.14

x

degradation

distinct

was 10^ of

have

gr

is

the

the

soil

not

a

a

the in

the the

found

at

widespread

the and

growth

on

predicted

conversion enhanced

activity (.26)

day

and

to

increase

curves

Soulas

the

theoretical

for

complete

case

the

as

response

rapid

to

in

in

adapted

decreased

considered

cells

and

systems

within

decline

developed

indicated

not

may

not

ability

subsection

was

dropped

conforms

model

This

metabolite

This

in

rapid

uptake

had

phase

size

Our data

applied pesticide.

population.

(26)

lag

biomass

pesticide.

model.

Soulas

was

and

The p o p u l a t i o n

a

in

pesticide

utilization

supply.

decline

particular

by

uptake

day

carbon

that

subsequent a

and

At

15

is

of

soils. of

the

model.

The

implies

harbored

that in

a

population.

Conclusions The

substrate

fact is

that

reflected

the

biomass in

conversion

both

use

al.

(U)

some

the

the

of

estimations

can

It

addition

the

it

is in

shown

organisms.

is

likely

degradation carbon.

on

to

biomass the

the

specifically clear the

that

glucose.

labeled the

size

for

bacterial

would

that

nitrogen use

of

of

serve

would

the

C-CO2.

to

as

is

the

relies

small.

nitrogen this

Ν

methylamine, overlook

methylamine

on

pesticide on From

m i c r o b i a l biomass

overlook

utilize

relies

of

Fumigation

biomass

side-chain

c a r b o f u r a n may

estimates

estimation

introduction

carbofuran

organisms

product,

Data

to

that Our

that

for

response

the

carbon

has

method response

is,

Karns source

that et for

utilization. the

this

primary

source

however,

Racke and Coats; Enhanced Biodegradation of Pesticides in the Environment ACS Symposium Series; American Chemical Society: Washington, DC, 1990.

163

of

limited.

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164

ENHANCED BIODEGRADATION O F PESTICIDES IN T H E ENVIRONMENT

0.5 10 CO CO

8+

σ

0

ε

*

.2 --5. CQ c

1c

o. ° Ο

f

«

Ο

6+

Soil I οεοιΙ II · -

4+

2+ 0

5 10 Time (days) 1 4

Figure 7. (a) Biomass size as indicated by fumigation; (b) incorporation of C into biomass. (Adapted from Ref. 24.)

Racke and Coats; Enhanced Biodegradation of Pesticides in the Environment ACS Symposium Series; American Chemical Society: Washington, DC, 1990.

12.

Results

from

process

that

follow

the

response

either is

changes

to

derived in

bacteria

the

two

the

from

become

about rely

insight

If

isolate

frequency be

of

into

are

at

direct

best

estimates

estimation

of

would

materials

regulating

a

possible

to

extract

for

a

gene

from

the

that

gene

sequence

This

growth

subculturing changes

probe

taken

established.

microbial

on

more

the

the

be

to

bacterial

application.

has

an

systems

A

genetic

DNA ( 2 7 - 2 9 ) .

could

not

direct

it

the

conclusions would

in

pesticide

restrictable changes

of

occurring.

Recently was

Response ofMicrobial Populations to Carbofuran

T U R C O & KONOPKA

in

and

from the

enhanced

soil

in

a

in

treated

soil

function then

(10),

population

information

function the

from

specific

would

soil. soil

The but

of

allow method provide

population.

Downloaded by UNIV LAVAL on April 10, 2016 | http://pubs.acs.org Publication Date: May 3, 1990 | doi: 10.1021/bk-1990-0426.ch012

Acknowledgment This

work

Pesticide University

was

supported

Impact

by

Assessment

a

Grant

Program.

A g r i c u l t u r a l Experiment

from Paper

Station

the No.

North

Central

12,353

of

the

Region Purdue

Series.

Literature Cited 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19.

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RECEIVED February 8, 1990

Racke and Coats; Enhanced Biodegradation of Pesticides in the Environment ACS Symposium Series; American Chemical Society: Washington, DC, 1990.