Butt.Fae. o6 Ag~e., Univ.o6 C~o, Vol . 43, No. 3(July 1992 ; 1055-1012.

MICROBIOLOGICAL INTERACTION IN DAMPING-OFF DISEASE OF CASUARINA

(received: 18.04.1992) By -

(1). • (1) M.S.M1khail ; K.K. Sabet

M.E. Abd El- Massih( 2 ) and Fatma M. Radwan( 2 )

1) Plant Pathologhy Dept., Faculty of Agriculture, Cairo University, Giza, Egypt.

2) Plant Pathology Institute, Agriculture Research Research Center, Giza, Egypt.

ABSTRACT

Fu-6a!vW.m OX!f6poltum and Rhi.zoc-tonia 60lani were found--- to. be the rrain caus~ of pre- and post-emergence darning-off of casuarina in four nursery locations. Inter-action between these two fungi increased the severity of the disease specially when R. 60lani was added to the soi 1 one week before SOl'ling. The presence of root-knot nematode increased the disease severity • R/S ratio in the case of the less susceptible causarina species was higher than that of the highly susceptible species.

INTRODUCTION

Casuarina species are known as the most extensively grown woody trees in Egypt. Their seedl ings are seriously affecteri by damping-off and root-rot caused by several fungi. The presenc e of root-knot nematode in combination with these fungi may cause greater decay and losses. In addition, infected seedlings could be an imRortant source of contamination when transplanted to the newly reclaimed lands (Farahat, 1976). The present investigation was carried out to study the causal agents of Casuarina damping-off and root-rot and the _interaction between the fungal pathogens . and root-knot nematode.

-1055--

-105 6-

MATERIALS AND METHODS

Diseased casuarina seedlings were collected from four nurseries, Bragil (Giza), Kom Oshim (Fayoum), Qanater (Qalubia) and Qorashia (Gharbia).

Infected roots were cut into small pieces, washed and surface sterilized with 4% Sodium hypochlorite solution for two minutes, rinsed in s teri 1 e water and plated on potato dextrose agar (PDA) . Pure cultures of isolated fungi were obtained by single spore or hyphal tip techniques.

Identification of the isolated fungi was made according to Barnett (1960) and Domsch et ~., (1980) .

Inoucla of fungi tested for pathogenicity were grown in sand-barley medium (1:3 w/w), in glass bottles ( 500 ml) and incubated at 28 °C for two weeks. Soil infestation was achieved by mixing the inoculum of each fungus with sterile soil at the rate of 5'/o (W/H). One week later, surface sterilized seeds of Ca6ua~na giauca were planted. Fu6aJUum oxy6poltum and RIUzoc.-ton..i .. a 6oian--i .. were selected for the other experiments by virtue of their wide spread and high virulence.

Susceptibility of C. giauca, C. cunninghamiana and the natural hybrid (c. giauce. c. cumunghamiana) as we 11 as some other woody trees s ho,.,n in Tab le (2,and 3) to the isolated pathogens was studied. All tree seedlings were ob La ined from Agriculture Research Center

Individual egg-masses of Meio~ogyne. ~ncognita isolated from infected casuarina roots by Radwan (1992) were used to i noculate " Pritchard" tomato seedlings previously planted in 15 cm pots filled with sterile sand Infected tomato roots were used as a source of inoculum .

-1057-

To study the interaction between nematode and the fungal pathogens, 20 casuarina seedlings were transplanted to soi 1. Nematode sus pens ion was added at of 6000 larvae/pot. After 60 days, preand emergence darning-off were determined

root-knot week-old infested the rate post-

Total microbial flora in the rhizosphere of C. giauca and the hybrid were determined in different nursery soils. Ninety days old seedlings were uprooted and one gram of rhizosphere (R) soil as well as non-rhizosphere soil (S) each was transfer­red to a flask with 99 ml steril water, Barakat et al-., (1986).

Dilution plate method was used to determi~g. (i) total bacterial count at the dilution of 10 on soil extract yeast agar medium (Skinner et ~· · . 1952); (ii) actinomycetes at the dilutionof 10-~ on Jensen 's agar medium ( Jens_~, 1930) and ( iii) fungi at the dilution of 10 on peptone dextrose agar medium (Martin, 1950). One ml of the above dilutions . was plated on each medium. All plates were incubated at 28°C • Total bacterial count was determined after 4 days, actinomycetes after 7 days and fungi after 5 days. Ratio of rhizosphere/soil counts (R/S) was also calculated for each group.

RESULTS

Fu6alt.ium ·axy6pcJJwm and RfUzoc.:torU.a 6oiarU. were the most frequently isolated fungi from diseased casuarina seedlings in the four nursery locations followed by F. mon)ti6okme, F. eq~~ and Phoma sp. ~~en these fungi were tested for their pathoge­nicity to ~C.giauca, they caused damping-off and decreased emergence in different degrees.F. oxy6pok­un and R. 6oia~ caused the highest percentage of pre- and post-emergence darning-off (Table 1) and were considered the main cause of the disease. C. gtauc.a and C. cwtrU.nghamiana were more susceptible

-10

58

-

Tsb

l e ( 1

) :

Av

erage

perc

en

tat:e

of p

re-,

po

st-em

....... .:·~enee

dam

pin

g­

oft an

d h

eal tt:y

surv

ival·· p

l8n

ta o

n C

esual·in

_!! e

leuc~

wi.t h

i so

l s t ed !u

ng

i •

?u

r.gi

% D

amp

ing-

off

Pre-

Po

et-ece.:-ge!lce

e::Ierger.ce

Fu

sarium

eg

uie

eti

J.4

51

.6

!• mo

nilifo

r:ne

6.9

4

4.8

P.

0X

Y9

C0

l"u::1

o.o

69

.0

Pb.oma

sp.

lJ .a

24

.1

Rh

! zo

cto

nia

so

lan

i J7

.9

24

.2

Co

ntro

l 0

.0

0.0.

~

Su

rviv

als

44

.8

4S

.3

31

.0

62

.l .

37

.9

10

0.0

----------~-~-------~------------------------------------------

L.S

.D.

at 5 ~

1

0.4

9

.8

15

.2

-1059-

than t he natural hybrid (Table 2 ) . Infection was man ifested as basal fjtell rot, and reduction of root system with dark brown lesions. Other tested tree species were infected with t he two pathogens. LPu~a­e.na tcu~o~e.pha.ea gave the highest percentage of pre­and postemergencc damping-off , A~eia 6atigna show­ad res i r.tance to both. Other trees varied in the­ir s usceptibility to both fungi (Table 3).

Combined inocula of the two pathogens increas­sed disease incider.ce and signHicantly decreased t he percentage of sur~i val plants than e:ither pathogen o l~ne. Pre-cmergence damping- off was very severe .in soil infested with F. oxy6po!Lwn plus R. -6o.f.ani nt inoculum ·levels 1. 75i'.: ·2.25%, respectively, and vice versa. Also, addition of R. 6otani to the soil, one week before sowing reduced plant s urvival significantly (Table 4).

Presence of M. ·..t.n~ognLta in combinction with e i t her F • (l>:y6pO!tu.m or R. 60tani has incre:e·­scd the perccnt3~c of infected plants, when compared with the pethogen :;lone in the case of C. gta!..l.~a and C. c.untU.ngltan'..iana,~hile no significant diff<.>rencc was found in the case of _ the hybrid (Table 5 ) .

E&ch fungus c9mbined with t he nematode reduced plant height, more than each fungus alone. The ~ame trend was observed in shoot weight. All treatment including r oot knot nematode increased r oot weight of C. giau~a and C. cunnlnghamiana secdlingo (Table 6) .

R/S ratio in the case of the hybrid "less susc­eptible" was higher than thst of C. gtauc.a "highly susceptible" (Table 7) .

DISCUSSION

Isolation from diseased seed~ings and pa t hogen­icity tests proved that Fuoaaltium oxy6poltum and

-1060-

·rable

(1.) Pcrcentae~ ot

p~e-

, poet-cmcrr.en~c dam

pins-ot! ond

o~1~ival

pla

nts of

three

casuar1

na

ape~ico

f~Own ir.

uo

il ln

feQtcd

with~~ oY

.yooorum

and Rh1~octonia aol~ni.

Sp

ed

eo

CaaugrinQ

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Sle

b e~.

Sp

ring

~u a

rt na c u nn1 n:3 h8:!1i ur.t\

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.

The hybrid (2_.

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L.~.D.

ui

5 ~

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ped

a11

x :!'u

t~i

:G inl~cti

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1 th :

---------------------------------------------------------· !llB

t\1•:!.. 1101 ~Xy~ p

Ol"U

t'J R

hh:o

cton

ia

oo

lani -

r~.-o-

Poot-·

Su!'vivblo

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ao p

crcentu

ec o! ::cnni::!lt1

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lOO

-1061-

Table

(3} :.

Su

sceptib

ility o

t five tree seedlings to pathogenic fungi.

%in

fection

in so

il infested with

:

'l'ree epecie!e

Fusarium ox~s2orum

Rhi zoctonie eoleni

Pre-P

oet-S

urvival a Pre-

Poet-

Survivalo

emergence

emergence

eme~gence

emergence

Ace cl e ael1:,K!!,!! o.o

o.o 100.0

o.o o.o

100.0

Cuoreesue

~npervirena

s.J o.o

94.7 52.6

·0.0

47.4

Eucelyotu

·~ c

i triodora 10·. 7

)2. 7

57.1 28.6

21.4 50.0

Leucaena

~ueoeephele

44.4 22.)

JJ,J 90.6

0.0

9.4

Pi nu e pi nes1:!! o.o

0.0

100.0

42.9 22.8

34 .)

All values w

e.re calculated as percentage of germ

ination of the control treatment =

100.

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("ti/'11

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re-

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22

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+ g~~~·

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63

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Tab

le ( 5

) :

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ct

of M

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og

yn

e _!.r:cop;ni ta,

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end

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gn

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(co

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r :

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ecie

s

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ent x

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ecie

s

c. g

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25

.)

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83

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o.o

17

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12

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Jl.O

Percen

taGe

of w

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p

lan

ts

:

C. cunni~hruniene

The

hy

brid

0.0

0

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25

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16

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8.)

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)).)

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tlnO-:otl.t!l

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rhizo~phere o

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-1066-

(a961). Similar results were reported Khan ( 1969 , a) and Sa bet and Khan Sabet ~ ~., (1966).

by Sabet and (1969,b) and

Radwan (1992) reported that Metoidogyn~ ~ncog­nita was the most common root-knot nemato de on collected samples on C. gtauea, C.eun~nghamiana and the hybrid, and that C. gtauea showed more root gal ling a nd egg-masses t han the others.

Interaction between M. ~neognlta and either F. oxy~pokum or R. ~ota~ increased fungal root infection more than each fungus alone, thus the· n ema tode predisposed causarina seedlings to be attacked by the tested fungi. Powe ll (1971) reporte d that root-kont infections induced morpholo­gica l and physiol ogical changes which render the host more s usceptibl e to fungus invasion.

Plant hight as well as shoot weight were v ery much affected and r educe d, when the fu ngi were combined with nematode more than the fungi ~lone. These changes were also observed b~

. Pow.,ll (1963) and Ibrahim et al. , ( 1982). On the other hand' root weight was increased in combined trea tment . This cG uld be attributed to the presence of root galls.

The rh i zosphere of the hybrid (Less s u sceptib­l e) gave higher densties of total bacteria and actinomyce fe s than tllat of C, gtcuca ( s usceptible ) when grown in the same soi 1 type. However, there was n o significant differen ce in the fungal density. This d i fference in rhizo ,.; pher e microflora is attributable to the dif ference in root exudates (Schro t h and lliidebrand 1964, Neal et al., (1973) were t h e first to sh0\-1 that the g;notype of t he host gov e rns the magni tude and composi tion of bac terial populations in the rhizosphere with high spec ificity. Benson and Baker (1970) advoca ted that specific nutrients in the rhizos phere become limiting to microbial growth and crucial for

-1067-

RfUzoc;torU.a &otarU. we:re tJU! .mo.st dominant and the main cause of casuarina damping-off in four nurseries. Rkizoc;torU.a &oiarU. only was previous ly isolated from casuarina roots in Egypt by Abdallah (1985) • The frequency of these fungi varied from nursery to another, evidently due to differences in environ­mental conditions .

It was found that C. gfruca was the rrost . susceptible followed by C. cunrU.nghamiana, while the hybrid was the least.

Other than casuarina, Leucaena teucocephatal!as the most -susceptible species. This result confirms the orndings of Hsieh (1982) and perez-Guerrero (1982) who reported that R. ~otarU. and F. oxy&pOII.um cause damping-off of L. ~ in Taiwan. Acac-ia 6a­Ugna was found to be resistant to the disease caused by the two fungi, but P~nu6 plna&teA~was res­~stant to F. oxy6pO~only. Other reports indicated that both species \'lere susceptible to R. 6·otarU. (Darvas !! al., 1978 and Saxena and Saksena, 1981).

Mixing the two pathogens in equal amounts increased the percentage of pre-emcrgence damping­off. This reveals a synergistic effect between the tested fl!ngi on disease severity. When they were added to the soil in unequal rates, the severity of the disease increased more than the previous case. This may be du~ to the combined effect of the predisposition brought about by the low rate pathogen and the hightened pathoginici ty due to the increased inoculum potential of the other.

The disease rate was higher when R. 60iarU. was added to the soil one week before sowing and F. oxy-6pOkum at the time of sowing than when the opposite was done. This may be due to the tendency of F.ox­y6pokum to act £s an active saprophyte competing with other pathogens in the absence of a s uitable host, as mentioned by Garrett (1956) and wastie

-1068-

prepentration growth.

llir results may explain the relative resistance of the hybrid. Similar results were obtained by Barakat et al., (1986). and Saied (1986). - -

REFERENCES

Abdallah, M.A. (1985): The rhizosphere fungi of some Egyptian desert plants. Ph.D. Thesis, Fac. of Sci., Tanta Univ., 209 PP.

Barakat, F.M.: , ·sabet K.A. and Halib W.F. (1986): Varietal r~action of faba bean and lentil to specific strain of ru6a~m 60iani in relation to anatagonistic rhizosphere microflora. Agric. Res. Rev., 64; 245-256.

Barnett, H.L. (1960): Illustrated Genera of Imperfe­ct Fungi Butgess publishing ComFany, 218 PP.

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