1990_____effect of water potential, temperature, and clay-coating on survival of beuveria bassiana...

8/17/2019 1990_____Effect of Water Potential, Temperature, And Clay-Coating on Survival of Beuveria Bassiana Conidia in a …

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JOURNAL OF INVERTEBRA TE PATHOLOGY s&417-427 (1990)

Effect of Water Potential, Temperature, and Clay-Coating on Survival

of Beauveria bassiana Conidia in a Loam and Peat Soil

JOHN

P.

STUDDERT'ANDHARRY

K.

KAYA

Departm ents of Entomology and Nem atology, University of Cal i fornia, Dav is, Cal i fornia 95616

AND

JOHN M. DUNIWAY

Departmen t of Plant Pathology, University of Cal i fornia, Dav is, Cal i fornia 95616

Received May 10, 1989; accepted September 11, 1989

Wh en Beauveria bassiana conidia w ere mixed in nonsterile Yolo tine sand y loam (YFSL ) or

Staten peaty m uck (peat) soil at water potentials ranging from 0 .0 bars (saturation) to - 1500 bars,

conidia ha lf-l ives were longes t at - 15 bars and decreased as the wate r potential approached either

0.0 or -200 bars. Conidia half-l ives increased as the wate r potential decreased fr om -200 to

- 1500 bars. Conidia half- lives were longest at a soi l temperature of 10°C and decreased both a t 2°C

and as the temperature approached 50°C where no conidia were recovered after 2 wee ks. T he

longest mean half- li fe value w as 44.4 week s for conidia in YFSL at - 10 bars and 10 °C; the shortest

half-l ife value w as 0.3 we eks in peat soil at 0 bars and 28°C. Clay-co ating lengthened conidia

survival in al l treatments in a factorial experiment involving the two soi l types and several co m-

binations of soi l temperature and water potential . When tw o strains of B. bassiana were compared,

colony co unts of strain’ABG-6178 alwa ys decreased relative to an initial b aseline co unt taken so on

after mixing conidia and soil; colony cou nts of strain IL-l 16 routinely increased after the baseline

count was taken before decreasing later. Conidia survival was often significantly longer in the low

organic Y FSL than in the high organic peat. The resu lts suggest that conidia survival is influenced

by the direct effect of physica l fact ors and soil microbial populations. Since B. bassiana conidia

survival is highly variable, its potential as a microbial insecticide is mu ch greater in som e soil

environments than in others.

6 1990 cademic press, Inc.

KEY W OR DS : Beauveria bassiana ; conidia surv ival; conidia half-l ife; soil wate r potential; soil

temperature; clay-coating; microbial insecticide.

INTRODUCTION

The entomopathogenic fungus, Beau-

veria bassiana, is a potentially important

microbial insecticide in the soil environ-

ment achieving partial control of the Colo-

rado potato beetle, Leptinotarsa decemlin-

eata (Watt and LeBrun, 1984), the pecan

weevil, Curculio caryae (Gottwald and

Tedders, 1983), and the curculionid, Sitona

lineatus (Mtiller-Kogler and Stein, 1970).

To be effective, B. bassiana conidia must

remain viable and infect insect hosts in the

soi l under a wide range of physical and bi-

ological conditions. Studies have deter-

’ Present address: Cooperative Extension, Univer-

si ty of Cal i fornia, 142A Garden H ighway, Yuba City,

California 95991.

mined the effect of temperature, relative

humidity, and light on the survival of ento-

mopathogenic Deuteromycetes conidia in a

nonsoil setting (Kawakami, 1960; Stein-

haus, 1960; Miiller-Kogler, 1964; Clerk and

Madelin, 1965; Walstad et al., 1970; Daoust

and Roberts, 1983). Soil-based studies with

B. bassiana have been limited to the lon-

gevity of conidia (Lingg and Donaldson,

1981; Fargues et al., 1983; Fargues and

Robert, 1985; Miil ler-Kogler and Zimmer-

mann, 1986). However, Lingg and Donald-

son (1981) determined the effect of physical

factors (soil moisture content, relative hu-

midity, temperature, and pH) on conidia

survival in the soil using moisture content

as a percentage of the saturation capacity of

the soil . Lingg and Donaldson (1981) and

417

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418

STUDDERT, KAYA , AND DUNIWAY

Fargues et al. (1983) also demonstrated the

importance of soil-biotic factors on conidia

survival.

Soil moisture has important effects on

microbial activity. Unfortunately, much of

the early research evaluating soi l moisture

effects on microorganisms cannot be inter-

preted adequately because soi l moisture

content was usually expressed in noncom-

parable terms (e.g., percentage of water-

holding capacity or percentage of soi l dry

weight). Furthermore, these soi l moisture

measurements are not related to the avail-

ability of water to soil microorganisms

(Griffin, 1963). Water potential is a more

meaningful measure biologically because

two soi ls with the same water potential

make water equally available to soil micro-

organisms even if their water, measured as

a percentage of soi l dry weight, differ (Grif-

fin, 1963; Sommers et al., 1981). Water po-

tential is defined as the chemical potential

of water per unit volume and has the dimen-

sions of pressure. The units are usually bars

or megapascals (Duniway, 1983; Griffin,

1963, 1981). In recent years water potential

has been used extensively to define soi l wa-

ter status in research conducted by plant

pathologists and soil microbiologists (e.g.,

Cook and Duniway, 1981; Duniway, 1976,

1983; Duniway and Gordon, 1986; Griff in,

1981; Sommers et al., 1981).

The objective of this study was to deter-

mine relationships between soi l water po-

tential and temperature and the survival of

B. bassiuna conidia in two nonsterile soils,

a low organic sandy loam and a high organic

peat. This was done for both clay-coated

and noncoated conidia and for two different

strains of B. bassiana.

MATERIALS AND METHODS

B. bassiana strains. Two strains of B.

bassiuna conidia, ABG-6178 and IL-l 16,

were obtained from Abbott Laboratories,

North Chicago, Illinois. The conidia were

stored in the dark at 10°C and 0% relative

humidity (RH). Unless otherwise stated,

only ABG-6178 was used because it was

being developed as a microbial insecticide.

Soil types. Two soil types, Yolo fine

sandy loam (YFSL) (cl% organic matter)

and Staten peaty muck (peat) (62% organic

matter), were used. The YFSL came from

the Botany Department (stored outdoors

and exposed to ambient conditions) at the

University of California at Davis; the peat

came from an abandoned field near Termi-

nous, California. The percentage sand, silt ,

andclaywas64.5,23.4, 11.6and12.1, 11.4,

14.4 in YFSL and peat, respectively. Water

release curves showing the relationship be-

tween soi l matric potential and percentage

water content were determined for both

soils (Fig. 1). Each soil was sieved (2-mm

mesh) to remove stones and large particles

and mixed in a cement mixer to achieve

uniformity throughout the soi l mass. Both

soils were stored air-dried at 24°C until ex-

perimental matric and water potentials

were established. The soils were not steril-

ized.

Water and matric potential determina-

tion. Saturated soi l has a water potential at

nearly 0 bars. As soil becomes drier, the

water potential becomes increasingly nega-

tive. Soil water potential has two major

components, matric potential and solute

potential. The capillary and absorption

forces associated with the soil matrix con-

stitute the matric potential. The matric po-

tential is essentially equal to the water po-

tential as long as a soil is low in salt. The

solute potential consists of the osmotic

forces caused by salt in the soil solution. In

our experiments, matric potentials were es-

tablished between 0 and - 15 bars; water

potentials were established for the -2OO-

and - 1500-bar soils.

Matric potentials of 0, -0.1, and -0.3

bars were established using Buchner fun-

nels of 9-cm diameter with fritted glass

plates of fine porosity (KIMBLE 28400-

9OF) as tension plates (Duniway, 1976). The

desired matric potential was obtained by

adjusting the height of a water column run-

ning between the surface of a water reser-


3/11

SURVIVAL OF Beauveria IN SOIL

160

pziq

ii?

xi 120-

E

8

ti EO-

2

w

0

Y

0

0 I I I I

I I

0 -3

- 6 - 9 - 12 - 15 - 18

Soi l Math Potential (bars)

FIG. 1. Soi l water content (percentage calculated as g water/100 g dry soi l) of Yolo f ine sandy loam

(YFSL) and Staten peaty m uck (peat) plotted as functions of decreasing soi l matric potential .

419

voir and the tension plate. Matric potential

was established in the Buchner funnels in

less than 24 hr. The -0.3-bar soi l was re-

moved from the Buchner funnels after ma-

tric potential was established and equili-

brated for 12

weeks

using the procedures

employed for the -2, - 10, and -U-bar

soils (see below). After this time period B.

bassiana conidia were mixed in the

-0.3-bar soil, and within l-3 hr, and the

first inoculated soi l samples were taken for

the baseline dilution series for the conidia

survival experiments.

It was not possible to follow the same

procedure with 0 and -0. l-bar soils be-

cause they were too wet to mix with conid-

ia. For these matric potentials, the conidia

were initially mixed in - IO-bar soil which

was placed in Buchner funnels where 0 and

-0.1 bar matric potentials were estab-

lished. Immediately thereafter the first soil

samples were taken from the Buchner fun-

nels for a baseline dilution series. Thus, the

0 and - 0. l-bar soi ls did not undergo a long

equilibration period before the mixing of

soi l with conidia as did the soi ls at the other

matric and water potentials.

A pressure plate apparatus (Soil Mois-

forma) was used for the determination of

soil water content at -2, - 10, and - 15

bars (Grifftn, 1963). These matric potentials

were subsequently established by first mix-

ing air-dried soi l (2000 g) with appropriate

amounts of water to create moist soil. This

moist soil, after equilibrating for 3 weeks,

was mixed with appropriate amounts of air-

dried soi l to adjust the net moisture content

of the soi l by weight to give the final matric

potential. For this work, water was added

to the soil with a hand-held mister. Al l mix-

ing of soil and water or moist soi l and air-

dried soil was done by hand inside polyeth-

ylene sacks to prevent moisture loss. The

soil was then allowed to equilibrate for 12

weeks inside polyethylene sacks wrapped

in cotton (to reduce condensation on the

inside of the soil sacks), covered with damp

paper towels (wetted weekly), and kept in

the dark in constant temperature chambers.

Soil samples taken monthly showed water

losses were no more than 5% of the original

soil moisture content in any soil sack over

the course of the experiments. The long

equilibration period was necessary to cre-

ate a uniform water potential throughout

the soi l mass. It may also have allowed dis-

ture Equipment Co., Santa Barbara, Cali-

tinct microbial populations, associated with


4/11

420


each soi l environment, a long time period to

develop prior to adding conidia to the soil .

Soil water potentials of - 200 and - 1500

bars were established by placing 200-g

quantities of air-dried soil to a depth of 2 cm

in open plastic containers inside desicca-

tors over saturated KC1 and MgCl, solu-

tions, respectively. These soils were al-

lowed to equilibrate for up to 20 weeks be-

fore the soi l water content became stable.

The KC1 and MgCl, solutions produce

RH values of 86 and 33%, respectively

(Greenspan, 1977). Because RH estab-

lished over saturated salt solutions varies

with temperature, the - 200- and - 1500-

bar values were approximate. For soils be-

tween -0.3 and - 1500 bars, conidia and

soil were mixed after the equilibration pe-

riod .

Soil inoculation with conidia and soil

storage. Al l soils were inoculated with dry

conidia by hand, using a tongue depressor

to mix the soi l and conidia, inside polyeth-

ylene sacks to prevent moisture loss. From

previous experience, 20 min of mixing were

sufficient to evenly distribute conidia in

1000 g of soil.

For the duration of the survival experi-

ments the 0- and - 0. l-bar-soils were left in

the Buchner funnels, -0.3-, -2-, and

- U-bar soils were left in the sacks where

soi l and conidia were mixed, and the - 200-

and - 1500-bar soi ls remained in the desic-

cators where water potential was estab-

lished. The soil sacks, desiccators, and

Buchner funnels were maintained inside

constant temperature chambers during both

the equilibration period and the survival ex-

periments (at either 16” or 28°C). For each

treatment, soi l was stored in four contain-

ers: either four Buchner funnels, each hold-

ing ca. 350 g of soil; or four polyethylene

sacks, each holding ca. 1000 g of soil; or

four open containers inside desiccators,

each holding ca. 200 g of soil.

Dilution series and colony counts. To

test the survival of B. bassiana conidia in

the soil, factorial experiments using differ-

ent soi l water content and temperature val-

ues were conducted. Five-gram soi l sam-

ples were taken, one from each of the four

soi l storage containers constituting a treat-

ment each time a dilution series was con-

ducted. Each soil sample was stirred in 250

ml of 0.1% agar water used to keep the soil

in suspension. After 3 hr, 5 ml of the orig-

inal suspension was pipetted into 200 ml of

0.1% agar water; 2 hr later, 1.25 ml was

taken from the second suspension and pi-

petted on each of three Petri plates contain-

ing a selective medium made with the fun-

gicide dodine (Chase et al., 1986). The Petri

plates were stored in the dark at 24°C for

6-7 days prior to counting all B. bassiana

colonies on each plate.

Except for the 0- and -0. l-bar matric

potentials, the baseline dilution series were

started within 1-3 hr of mixing conidia and

soil. For the 0- and -O.l-bar soils, the

baseline dilution series were begun 24 hr

after mixing soil and conidia, to allow ma-

tric potentials to establish in the Buchner

funnels. Later dilution series and colony

counts were conducted 2 and 5 weeks after

the baseline series and at 5-week intervals

thereafter until no conidia could be recov-

ered from a treatment or until 50 weeks had

passed. In the comparison of two fungal

strains, colony counts were also made 1

week after the baseline series. Sufftcient

numbers of conidia were mixed with the

soi l so that the baseline colony counts

ranged from 300 to 400 colonies per plate.

Clay-coating of conidia. B. bassiana co-

nidia were washed in distilled water, al-

lowed to dry, and coated by mixing conidia

with a bentonite clay (1:3 by weight) pro-

vided by Mycogen Corp., San Diego, Cali-

fornia. This mixture was spread out flat on

a plastic surface, sprayed lightly with ster-

ile distilled water from a hand-held mister,

and allowed to dry for 72 hr in the dark. The

dry mixture was used in experiments where

the survival of clay-coated and noncoated

conidia were compared.

Calculations of conidia half-lives and

statistical analysis. Conidia half-lives were

calculated using the procedures employed


5/11

SURVIV AL OF Beauveria IN SOIL 421

by Lingg and Donaldson (1981). A half-life

was calculated for each curve generated by

the decrease in B. bassiuna (strain ABG-

6178) colony counts over time. Four such

curves were generated per treatment, one

curve from the soi l samples taken from

each of the four soi l containers that consti-

tuted a treatment. These half-life values

were subjected to a logarithmic transforma-

tion prior to an analysis of variance and

Duncan’s multiple-range test. The 5% level

of significance was employed for all data.

Terminology.

During the discussion of

experimental procedures, care was taken to

maintain the distinction between matric and

water potential. Since our soils were low in

salt (peat has < 1.0 mmhos/cm and YFSL

~0.2 mmhos/cm), the difference between

water potential and matric potential was

small so the term water potential is used in

the remainder of the paper.

RESULTS

B. bassiana

strain and conidia survival.

Two strains of

B. bassiana

responded very

differently in a dilution series experiment

(Figs. 2A and 2B). The ABG-6178 colony

counts decreased over time in all trkat-

ments. With the IL-116 strain an initial in-

crease in colony counts, relative to the

baseline count, occurred for all combina-

tions of soi l type and water potential. This

increase was apparent at 1 week following

the baseline count. At 2 weeks counts were

more than twice the baseline count. From

that point on IL-l 16, colony counts de-

creased at a more rapid rate than the de-

crease shown by ABG-6178 counts so that

recovery of the two strains ceased at about

the same time. Since populations of ABG-

6178 always decreased in a manner approx-

imating first-order kinetics, survival of

conidia of this strain was expressed in half-

lives to compare treatments in the experi-

ments discussed below.

Soi l water potential and conidia survival.

When B. bassiana conidia were subjected

to water potentials ranging from 0 to - 15

bars, half-l ives were longest at - 15 bars

and decreased as the soil became wetter,

i.e., as water potential increased to 0 bars

(Table 1). At saturation (0 bars) and -0.1

bars, conidia were recovered in all treat-

ments at 2 weeks, but no recovery occurred

at 5 weeks. Al l conidia half-lives at these

high water potentials were less than 1

week. As water potential decreased from

-0.1 to - 15 bars, conidia half-lives in-

creased more rapidly at 16°C than at 28°C

(for YFSL and peat) and more rapidly for

YFSL than for peat (at 16” and 28°C). As a

result, conidia half-lives reached their high-

est values of 36.3 and 15.2 weeks at 16°C

and - 15 bars in YFSL and peat, respec-

tively. When soil water potential was de-

creased from - 15 to - 200 bars, there were

significant decreases in conidia half-lives in

all treatments. At -200 bars conidia half-

lives ranged from 1.1 to 3.5 weeks. How-

ever, as water potential decreased further

to - 1500 bars, conidia half-lives increased

again and were significantly higher than

were the corresponding values at -200

bars. At - 1500 bars, conidia half-lives

ranged from 5.3 to 7.9 weeks. At all water

potentials, conidia survival was longer in

YFSL than in peat (at both temperatures)

and longer at 16°C than in 28°C (for both

soil types). These differences were usually

significant in the middle range of water po-

tentials between -0.3 and - 15 bars and

usually not significant in the wetter and

drier soils.

Soi l temperature and conidia surviva l. B.

bassiuna conidia (strain ABG-6178), mixed

in soil and subjected to a range of temper-

atures from 2” to 50°C survived longest at

10°C (Tab e 2). Conidia half-lives then de-

creased steadily as temperatures increased

from 10” to 50” until no recovery occurred

at 2 weeks at 50°C. Conidia half-lives were

significantly less for all 2°C treatments

when compared to the corresponding 10°C

treatment. At 2”, lo”, and 20°C conidia sur-

vival was significantly longer in YFSL than

in peat (at -0.3 and - 10 bars) and signifi-

cantly longer at - 10 bars than at - 0.3 bars

(for YFSL and peat). At 30”, 40”, and 50” no


6/11

422

STUDD ERT, KAYA, AND DUNIWA Y

t

-

5

z

m

A

I

YFSL 28” C

I

,e -1OBars IL-116

- -0.3 Ban IL-1 16

- -10 Bars ABG-6176

e -0.3 Bars ABG-6176

100

50

0

0

5 10 15 20

25

z 250

B

‘;

iti

t

P

200

PEAT 28” C

-o- -1OBars IL-116

--)- -0.3 Bars IL-116

150

---(>- -1OBars ABG -617 6

--f- -0.3 Bars ABG-6178

100

50

0

0 5 10

15

Time (Weeks)

20 25

FIG. 2. Beauveriu bassiana colony counts made over t ime and expressed as a percentage of the

basel ine colony counts taken a t Wee k 0. The graphs show the results of di lut ion series made period-

ically after mixing strain ABG-6178 or IL-116 conidia in (A) Yolo tine sandy loam (YFSL) and (B)

Staten peaty muck (peat). The soils were maintained at 28°C with a water potential of -0.3 or - 10

bars. Each percentage is based on four dilution series, one of each of four soil containers making up

a treatment, and three colony counts for each di lut ion series.


7/11

SURVIVAL OF Beauveria IN SOIL

423

TABLE 1

SURVIV AL (HALF-LIVES IN WEEKS’) OF Beauveria bassiana CONIDIA IN NONSTERILE SOIL, EITHER YOLO

FINE SANDY LOAM (YFSL) OR STATEN PEATY MUCK (PEAT), AT DIFFERENT WATER POTENTIALS

AND TEMPERATURES

Temperature

Water potential (bars)

Soil (“Cl 0.0 -0.1 -0.3

-2.0 - 15.0 -200.0 - 1500.0

YFSL 16 0.6 Aa 0.9 Aa 15.0 Ad 26.7 Ae 36.3 Af 3.5 Ab 7.9 AC

28 0.4 Aa 0.5 Aa 4.9 Bb 5.1 Cb 5.4 Cb

1.2 Ba 6.3 Ab

Peat 16

0.5 Aa 0.1 Aa 5.1 Bb 13.8 Bc 15.2 Bc 1.3 Ba 6.9 Ab

28 0.3 Aa 0.4 Aa 2.1 Cbc 2.1 Dbc 3.1 Ccd

1.1 Bab 5.3 Ad

e Mean of four half-l ife values in we eks. Each half- l ife value was calculated from the conidia survival curve

generated from B. bassiana colony counts from a single soi l container. There were four containers per treatment.

Means fol lowed by different uppercase letters in a column and by different lowercase letters in a row are

significantly different (P < 0.05) according to Dun can’s m ultiple-range tes t.

significant differences in conidia half-lives,

related to water potential or soi l type, were

observed.

Clay-coating and conidia survival. In all

cases, regardless of soi l type, water po-

tential, or temperature, conidia half-lives

(strain ABG-6178) were significantly longer

in treatments using clay-coated conidia

than in the corresponding treatments with

noneoated conidia (Table 3). For both clay-

coated and noncoated conidia at 10°C sur-

vival was significantly longer in YFSL than

in peat, regardless of water potential, and

significantly longer at - 15 bars than at

- 0.3 bars, regardless of soil type. At 30°C

the spread between the shortest mean half-

life value (2.0 weeks for noncoated conidia

in peat at -0.3 bars) and the longest mean

half-life value (12.2 weeks for clay-coated

conidia in YFSL at - 15 bars) was much

less than the corresponding spread at 10°C

(11.8 to 64.4 weeks). As a result of the rel-

atively low half-life values at 30°C most of

the significant differences related to soil

type and water potential at 10°C did not oc-

cur at 30°C. Also for clay-coated and non-

coated conidia, half-lives were significantly

longer at 10°C than in the corresponding

30°C treatment, regardless of soi l type or

water potential.

DISCUSSION

B. bassiana strain and conidia survival.

We cannot adequately explain why strain

TABLE 2

SURVIV AL (HALF-LIVES IN WEEKS~) OF Beauveria bassiana CONIDIA IN NONSTERILE SOIL, EITHER YOLO

FINE SANDY LOAM (YFSL) OR STATEN PEATY MUCK (PEAT) , AT DIFFER ENT TEMPERATURES AND

WATER POTENTIALS

Soil

Water

potential

(bars) 2 10

Temperature (“C)

20 30 40 50

YFSL - 10.0 21.6 Ad 44.4 Ae

20.8 AC 4.3 Ab 1.8 Aa NR b

-0.3 12.2 cc 21.8 Bd 10.9 Bc 3.9 Ab 1.6 Aa NR

Peat - 10.0 16.9 Bd 23.4 Be 10.7 Bc 3.3 Ab

1.1 Aa NR

-0.3 8.3 DC 13.9 Cd 6.9 Cc 3.0 Ab

0.8 Aa NR

’ Mean o f four half-l ife values in we eks . Each half-l ife value wa s calculated from the conidia survival curve

generated from B. bassiana colony counts from a single soil container. There were four containers per treatment.

Means fol lowed by di ierent uppercase letters in a column and by different lowercase letters in a row are

significantly different (P < 0.05) according to Dun can’s multiple-range tes t.

b NR , no recovery at 2 wee ks after conidia were mixed in soil .


8/11

424

STUDDERT, KAYA, AND DUNI WAY

TABLE 3

SURVIVAL (HALF-LIVES IN WEEKSO) OF CLAY-COATED

(CC)

AND NON COAT ED (NC) Beauver ia bass iana

CONIDIA IN NONSTERILE SOIL , EITHER YOLO FINE SANDY LOAM (YFSL) OR STATEN PEATY MUC K (PEAT),

AT DIFFERENT WATER POTENTIALS AND TEMPERATURES

Temperature

cc ,

Conidia

YFSL Peat

-0.3 Bars

- 15.0 Bars

-0.3 Bars - 15.0 Bars

10

30

cc 31.4 Ab 64.4 AC 20.3 Aa 34.2 Ab

NC 26.1 Bb

44.2 Bc 11.8 Ba 26.8 Bb

cc 11.4 Cb 12.2 Cb 6.6 Ca 9.7 Cab

NC 4.6 Dab

6.6 Dbc 2.0 Da 4.4 Dab

LIMean of four half- l ife values in wee ks. Each half- l ife value was calculated from the conidia survival curve

generated from the B. bassiana colony counts from a single soi l container. There were four containers per

treatment. Means fol lowed by different uppercase letters in a column and by different lowercase letters in a row

are significantly different (P < 0.05) according to Dun can’s multiple-range tes t.

IL-116 colony counts increased to about

200% of their baseline values over a 2-week

period before declining. Baseline colony

counts, using ABG-6178 conidia, were usu-

ally within 15% of values expected when

extrapolating from the number of conidia

per gram in the storage containers. For the

IL-116 strain, the baseline colony counts

were about 25% of what would be expected

when extrapolating from the storage con-

tainer counts. Possibly, at the time of the

baseline dilution series, the IL-116 conidia

were clumped in the soil, each clump giving

rise to a single B. bassiuna colony. Later,

less clumping occurred and colony counts

increased. If such artifacts are operating,

care must be taken in interpreting colony

count numbers. Fargues and Robert (1985)

speculated that the survival of Metarhizium

unisopliue is influenced by microcycl ic

conidiation in the soil , but we have no evi-

dence that microcyclic conidiation could

account for the survival pattern of strain

IL- 116. Miiller-Ki igler and Zimmermann

(1986) also obtained an unexplained in-

crease in B. bussiunu colony counts from

soil dilution series over time.

Soi l water potential and conidiu survival.

In nonsoil experiments, Clerk and Madelin

(1965) and Daoust and Roberts (1983)

showed that the viabili ty of M. unisopliue

conidia first decreased and then later in-

creased as equilibrium RH decreased from

98 to 92% (- 25 to - 110 bars water poten-

tial) to 0% (0 bars). The reason for this phe-

nomenon is unknown. The survival of B.

bussiunu conidia in the soil follows a similar

pattern. In our study, B. bussiunu conidia

half-life values decreased from a high point

at - 15 bars (98.9% RH) to significantly

lower values at - 200 bars (86% RH). How-

ever, as the water potential decreased fur-

ther to - 1500 bars (33% RH), conidia half-

lives increased again. Since there is little

microbial activity in soils drier than -50

bars (Wilson and Griff in, 1975), the expla-

nation for this phenomenon is probably

physiological in nature.

Lingg and Donaldson (1981) pointed out

that soil RH often is not drier than 98.9%

( - 15 bars). This is certainly true in agricul-

tural soils where moisture levels are kept

high enough to support crop growth. Since

many plant species reach their permanent

wilting point at ca.

- 15 bars, RH values

below 98.9% usually do not occur in these

soils except at the soil-air interface. How-

ever, in soi l environments where there are

long periods without precipitation, water

potential can reach values at least as low as

- 100 bars (92.9% RH) (e.g., Cook and

Duniway, 1981). The survival of

B. bussi-

unu conidia will be influenced to the extent

that natural soi ls in dry areas reach these

low water potentials.

A review by Sommers et al. (1981)

showed that the most extensive microbial

(mostly bacterial) decomposition of organic


9/11

SURVIV AL OF Beauveria IN SOIL

425

material occurs in soils at water potentials

ranging from -0.1 to -0.3 bars. Extensive

bacterial decomposition can also occur in

wetter soils at or approaching saturation (0

bars). Bacterial movement, the ability to

lyse fungal mycelium, and bacterial respi-

ration become negligible in all soi ls by the

time water potential decreases to - 15 bars

Wilson and Griff in, 1975; Grifftn, 1981).

Wilson and GrifIin (1975) also observed a

rapid decline in total soi l microbial respira-

tion between -3 and -8 bars, mostly due

to reduced bacterial activity. Between -8

and -30 bars total microbial respiration

(mostly fungal respiration) declined slowly

before decreasing rapidly and becoming

negligible at - 50 bars. Thus, it seems likely

that the low conidia half- lives in our study

at 0 and -0.1 bars were at least partly due

to the destruction of B. bassiana conidia by

bacteria. The increase in conidia viabil ity

as water potential decreased from - 0.1 to

- 15 bars may represent decreased destruc-

tion of conidia by soil microorganisms as

moisture conditions became drier and less

favorable to their activity. However, be-

cause the direct effects of high water poten-

tials on

B. bassiana

conidia are unknown,

there may also be physiological limitations

on conidia survival in very wet soil .

Fargues et al. (1983) demonstrated that

clay-coating increases B. bassiana blas-

tospore survival by protecting against

bacterial lysis in soils at 80% of mois-

ture-holding capacity. Unless clay-coating

changes the water potential experienced by

blastospores, this shows that they can sur-

vive the direct effect of water potentials

close to saturation for longer time periods

when the effect of bacterial lysis is reduced.

We do not have similar data for B. bassiana

conidia, but clay-coating significantly en-

hanced B. bassiuna conidia survival in soil

at -0.3 and - 15 bars (Table 3).

Since B. bassiana conidia did not survive

well in our soil at 0 and -0.1 bars, the ef-

fectiveness of B. bassiana, as a microbial

insecticide, will likely be reduced in very

wet soils. On the other hand, since conidia

survival significantly increased at water po-

tentials between -0.3 bars (field capacity

for many soi ls) and - 15 bars (permanent

wilting point for many plants), it appears

that

B. bassiuna

conidia survive relatively

well over the range of water potentials oc-

curring most frequently in agricultural

soils. Also, because clay-coating of B. bas-

siuna conidia increases half-lives over a

wide range of water potentials, we con-

clude that conidia survival will often be suf-

ficient to use

B. bassiuna

as an insecticide

in the soil.

Soi l temperature and conidia survival.

The importance of temperature to the sur-

vival of B. bassiana conidia outside of soil

has been demonstrated in several studies

showing sharp decreases in conidia survival

as temperature increased from ca. 8°C to

the thermal death point at 50°C (Clerk and

Madelin, 1965; Steinhaus, 1960; Walstad et

al., 1970).

Soil temperature probably also had an

important indirect effect on conidia sur-

vival in our experiments. Microbial decom-

position of soi l organic matter is greatest at

temperatures from ca. 18” to 35°C when wa-

ter potential is between saturation and - 15

bars (Nyhan, 1976; Wildung et al., 1975).

Accordingly, the decrease in B. bassiana

conidia survival associated with increasing

temperature in our experiments, especially

at temperatures within the optimum range

for microbial activity, may have been due

to increased microbial degradation of these

conidia.

To the best of our knowledge, the sur-

vival of

B. bassiana

conidia has not been

tested previously at 2°C. Our finding that

conidia half-lives were less at 2°C than at

10°C is contrary to several nonsoil studies

where conidia survival has been shown to

increase as the temperature approaches

freezing (Muller-Kogler, 1964). For M. an-

isopliue, at most temperatures, conidia sur-

vival was longest at 0% and 97-98% RH

and shorter at intermediate RH values; but,

at 4°C viabili ty was less at 98% than at

some intermediate RH values (Daoust and


10/11

426


Roberts, 1983).As a result, conidia survival

at 98% RH was shorterat 4°C than at higher

temperatures. f the same relationship be-

tweenmoisture evel and temperature olds

for B. bassianaconidia, t could explain the

observed decrease n conidia half-lives in

our experiment at 99.9% RH (-0.3 bars)

and 99.3% RH (- 10bars) as the tempera-

ture approached reezing.

Conidia half-lives were ess than 1 month

at temperaturesof 30°C or higher for all

combinationsof soil type and water poten-

tial. This rapid loss of conidia viability at

higher temperatureswould make t difficult

to use B. bassiana as a microbial insecti-

cide n the soil in warm climates. This prob-

lem may be partly overcomeby using clay-

coated conidia since our results demon-

strate the usefulnessof this technique for

prolonging he life of

B. bassiana

conidia n

the soil.

Soil type and conidia survival. Because

we measured the moisture levels in our

soils n terms of water potential, it was pos-

sible to compareconidia survival in two dif-

ferent soils over a wide range of moisture

and temperature conditions. In all of our

experiments,conidia half-lives were signif-

icantly longer in YFSL than in the corre-

sponding eat reatmentat the middle range

of water potentials (-0.3 to - 15bars)and

at temperaturesup to and including 20°C.

At the more extreme water potentials and

at the higher temperatures, these differ-

enceswere no longer significant. The peat

soil had a much higherorganiccontent han

the YFSL and high organicsoils often have

high populations of bacteria, fungi, and

actinomycetes which show significant an-

tagonisticactivity againstsome ungal plant

pathogens (Weste and Vithanage, 1978;

Malajczuk, 1983), and an entomopatho-

genie fungus(Lingg and Donaldson, 1981).

Thus, high populations of microorganisms

may account, in part, for the relatively

short conidiahalf-lives observed n our peat

soil.

Our results indicate that

B. bassiana co-

nidia will survive better in some soil types

than in others. For this reason, t may be

necessary to initially determine conidia

half-lives in soils where ong-term survival

is important, so that the proper quantities

of conidia can be added to soil where

B.

bassiana is to be used as an insecticide.

ACKNOWLEDGMENTS

We thank Dr. M . Stimmann and Dr. E. Butler for

reviewing initial dra fts of the ma nusc ript. The senior

author w as supported by grants from the IR-4 program

and scholarships from the Cal i fornia Nurseryme n’s

Association.

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1990_____effect of water potential, temperature, and clay-coating on survival of beuveria bassiana...

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