Zbl. ::\Iikrobiol. 139 (1984), 97-107

[Division of Microbiology, Indian Agricultural R esearch Institute, New D elhi , India]

P ressmud: A P ot ent ial Carrier for Rhizobium and Azotobacter

II. Influence of Storage Temperatureon the Survival of Rhizobium and Azotobacterin Different Carriers and Nodulation Efficiencyof Rhizobial Inoculants

K. S. JAUHRI and KOSHY PHILIP

With 3 F igures

Summary

Factors a ffec t in g the proliferation and sur viv a l of t wo s t rains of rhizobia an d one st rain ofA zotobacter (Rhizobium j ap onicum, SB-16 ; Rhizobium sp ., M.IO; Azotobacter chroococcu m , A.41)in suga rcane pressmud were exam ine d . W hile pressmud as suc h was not a perfect ca rrier forRhizobium nor A zotobacter, p ossibilities of a mending it with various a dd itives for its enhancedeffec tivene ss wer e evaluated . The survival of test -organisms was stu d ied under d iffere nt s t oragetemperatures .

The nodula ti on effic iency of v arious ca rr ie r b a se inoculants, st ore d under d iffe re nt t empera­tures , was ev a lua ted in a p ot trial in Glycine max val' . Bragg an d Vigna radiata val'. P S-16. Them aximum n umber of rhizob ia on seed a nd nodules pel' pl an t were obtaine d wh en inoc ula nts wereprepa red fr om pressmud plus charc oal (75 : 25) an d s t ore d at 32 DC.

Zusammenfassung

Es wurden F aktoren, welch e Proliferation und Uberleb en zweier Rhieobium- und eines A zoto­bacter-Stammes in dem be i del' Zuck errohrverarbeitung anfa lle n de n Abpro du kt "Pressmud",ei ne r m iigl ich en Triigersubstanz fur Impfpriipa ra te, beeinflussen. Pressmud a lle in war kein e idea leTriigersubs tanz, do ch wurde n Verbesserungsmiiglichkeiten in F orm v erschied ener Zusatee ge­priift. D as U be rleben del' Test organismen wurde bei un terschiedlichen L agerungstemperaturenuntersuch t.

Die F iih igk ei t del' bei unterschiedlich en Temperaturen ge lagerten P ra p ara te , h ergestell t aufdel' B a sis vo n Presamud, zur Kniillch enbildung , wurde in Gefiil3vers uchen m it Glycine max val'.Bragg und Vigna rad iata val'. PS-16 ge p riift. Maxrmal e Rhizobium-Keim zahlen an Samen undKniillch en pro Pflanze wurden erzie lt, we nn das Impfpr iipa rat aus P ressmud und H olz k ohl e(75: 25) h ergestellt und b oi 32 DC gelagert wurde,

P ressmud has great pot ential as a carrier for microbial inoculants. Though it svalue as a nutrient source and as a soil amendment is well known, t here are other useswhich are worth st udying . The pot ential use of pressmud as a carrier mat eri al forAzotobacter (SANORIA 1965) and for Rhizobium iaponicuo: (TILAK and SUBBA RAO1978) has been pointed out earlier. DUBE et al. (1980) have observed that the rela tiveindex of performance of different carriers like pr essmud, coal, and lignite were identi­cal,but it s possible use in commercial production of inoculants is yet to be explored .

Aft er st udying the physical and che mical properties of various carrier materialsand the effect on the growt h of t est organisms, attempts are being made to amendpressmud with charcoal or soil to enhance its efficien cy as a carrier . The survival of

7 ZbI. l\I1krobi oI., Bd. 139

98 K. S. JAUHRI and K. PHILIP

the test organisms under different temperatures and periods of storage was studiedwith emphasis on standardization of some physical factors, responsible for regulatingthe growth and survival of the test organisms.

The quality of commercially produced inoculants vary widely with the nature ofpreparation and storage (GERNELL and ROUGHLEY 1975; SKIPPER et al. 1975). It isimperative, therefore, to test the efficiency of rhizobial inoculants, which have beenprepared in different carrier materials and stored under different temperatures, beforereleasing them for use.

Materials and Methods

Maintenance of test organisms, preparation of media, carrier materials, and inocula, assessmentof population of the test-organisms, and statistical analysis of the data were the same as describedearlier (PHILIP and JAUHRI 1984).

Preparation of inoculants: Triplicate sets of 200 g each of the carrier material were steam­sterilized at 121 DC (15Ibs.Jsq. inch pressure) for 4 h. The cooled material was transferred asep­tically into sterilized polythene bags and mixed with the required quantity of young culture. Themoisture level was adjusted to 40 %of the water-holding capacity ofthe carrier material by mixingwith sterile water (inclusive of the water carried by the inoculum) and the bags were sealed.

Storage of inoculants: The inoculants were divided into three lots and stored at three differenttemperatures, i.e., 5-8 DC (refrigerator), 32 DC (B.O.D. incubator), and 42 DC (E.O.D. incubator).

Pot trial: The efficiency of inoculants, prepared in different carrier materials and stored atdifferent temperatures for three months, was assessed by testing the effect of inoculation onnodulation of the homologous hosts. Earthen pots of 4 kg capacity were filled with alluvial soilfrom the farm of Indian Agricultural Research Institute, New Delhi. Soil in each pot was mixedwith superphosphate per 180 mg P 205 ; zinc sulphate, 12 mg; manganese sulphate, 6 mg; ammo­nium molybdate, 6 mg. Seeds of Vigna radiata var. PS-16 and Glycine max var. Bragg were inoculat­ed with their respective carrier-based inoculants at the normal rate, using 10 % sucrose solutionas adhesive, and sown in triplicates. After germination, 3 plants per pot were maintained. N odula­tion was examined after 35 days of sowing.

Results and Discussion

The results (Tables 1-3; Figs. 1-3) of the experiments on growth and survival ofRhizobium japonicum, Rhizobium cowpea sp., and Azotobacter chroococcusn. in variouscarriers at different temperatures show that the four major factors, viz., type of car­rier, additive, period, and temperature of storage, influence the growth and survivalof the test organisms.

ROUGHLEY and VINCENT (1967), while working with four types of peat, showedthat the quality of peat affected the growth and survival of rhizobia. PRAMANIK andISWARAN (1973) and TILAK and SUBBA RAO (1978) have shown the variability in thegrowth of the organisms, when grown in different substrates. The results of thepresent investigation (Fig. 1; Table 1) also show variation in growth of R. japonicum,Rhizobium sp. and A. chroococousn, when grown in various carriers used in this study.Maximum growth of the test organisms was observed in pressmud plus charcoal(75: 25), while minimum in pressmud alone (100 %). In combination other than theformer, the growth of organisms was lowered by increasing addition of charcoal. Theseobservations led to the presumption that addition of 25 % charcoal to pressmud issufficient to bring down its repressive effect. Addition of charcoal to pressmud beyondthis quality progressively reduces the nutrient level in the carrier mixture. Theresults of physical and chemical analyses (PHILIP and JAUHRI 1984) also showed lowercontents ofN, P, K and less availability ofN and P with other combinations of press­mud and charcoal, namely (50: 50) and (25: 75). The water-holding capacity of thesemixtures was lower than the combination of pressmud plus charcoal in the ratio 75: 25.

-> *

36

0R

hiz

ob

ium

jap

on

icu

mIS

8-1

6J

Rh

izo

biu

msp

IM

-\O

JA

zo

tob

acte

rch

roo

cocc

um

lA-4

1l

32

0

r-, S?

12

0xL. '"

28

0L. L. o U ....2

40

o Ol

<, 02

00

:0 o N &1

60

>-d & ~ iO P0­ t;» >-d o et­

a> E: :::.. Q ~ (p'

:-'

H ~

1210

86

Pe

rio

d(w

ee

ks)

4~=~=

~e\

~===-

.~

x

2

o ZL. Q)

L. L. o U ... o Ol

<, L. Q) U o .0 o (; N «

lDo x

120

108

(50

:'0

)

12

5:7

51

6P

eri

od

(we

eks)

42

->.

b'

~~~

•~

+~~=-~u-t~.

~P

EA

T

•A

DD

ITIV

E1

10

0"I

.I

•P

ftE

SS

MU

D1

10

0'I

.)

o+

AO

DIT

IVE

(7~.251

+ x

o ZL. ."! t o o .... o ~ o :0 o N :c 0: r-

, S? x 120

104

68

Pe

rio

dIw

ee

ksJ

2o

80

40

o Z

Fig

.1.

Infl

uen

ceo

fty

pe

of

carr

ier'

.mat

eria

lsat

vary

ing

per

iod

so

fst

ora

ge

on

gro

wth

an

dsu

rviv

al

of

Rh

izo

biu

mja

po

ni­

cum

,R

ho

zob

ium

sp.,

an

dA

zoto

bact

erch

rooc

occu

m,

(l:)

(l:)

100 K. S. JAUHRI and K. PHILIP

Table 1. Influence of type of carrier materials and period of incubation on the growth and survivalof Rhizobium japonicum (SB-16), Rhizobium sp. (M-10), and Azotobacter chroococcum (A.41)

Type of carrier Log No. of viable bacteria/g of carrier

Period (Weeks)

o 1 2 4 8 12 Mean

Peat (100%)Additive (100 %)Pressmud (100 %)Pressmud + additive (75: 25)Pressmnd + additive (50: 50)Pressmud + additive (25: 75)Mean

Rhizobium [aponicum (SB.16)

5.89 7.21 8.04 8.56 7.90 7.225.70 7.23 7.93 8.63 8.00 7.095.76 5.99 7.71 8.17 7.83 6.806.07 7.98 8.95 9.51 8.92 8.125.88 7.69 8.31 8.97 8.43 7.625.67 7.14 7.92 8.27 8.08 7.295.82 7.20 8.14 6.68 8.19 7.35

7.477.437.048.257.817.39

Peat (100%)Additive (100%)Pressmud (100%)Pressmud + additive (75: 25)Pressmud + additive (50: 50)Pressmud + additive (25: 75)Mean

Rhizobium sp. (M.IO)

6.48 7.18 7.55 7.945.85 7.95 7.38 7.635.33 7.46 7.53 7.926.46 7.60 8.12 8.546.74 7.50 7.87 8.265.74 7.17 7.57 7.766.10 7.47 7.67 8.00

7.997.798.289.008.628.308.33

7.127.227.188.288.137.767.61

7.377.307.288.007.857.38

Peat (100%)Additive (100 %)Pressmud (100 %)Pressmud + additive (75: 25)Pressmud + additive (50: 50)Pressmud + additive (25: 75)Mean

Azotobacter chroococcum (A.41)

5.39 6.61 7.36 7.66 7.77 7.775.55 6.48 7.26 7.69 7.77 7.525.81 6.29 7.12 7.35 7.35 7.185.48 6.60 7.46 8.32 8.41 7.845.56 6.51 7.39 8.07 8.10 7.505.45 6.36 7.15 7.81 7.87 7.335.54 6.47 7.29 7.81 7.87 7.52

7.097.046.857.357.186.99

R. japonicum Rhizobium sp. A. chroococcum

S.Em. C.D.at S.Em. C.D.at S.Em. C.D.at

5% 5% 5%

Carrier/period 0.06 0.17 0.05 0.15 0.04 0.12

Carrier X period 0.15 0.42 0.13 0.35 0.10 0.29

The poor growth and survival of the test organisms in pressmud alone (100 %)may be attributed to certain repressive effects, natural to its composition. Pressmud,being a by-product of the sugar industry, contains many organic and inorganic, metal­lic and non-metallic residues, left in it during the processing of sugar. It is quite likelythat these substances may have a repressive effect on the growth of the test organ­isms. Charcoal, being a good adsorbent of extraneous substances, might have beeneffective in adsorbing such substances when added to pressmud, which otherwisesuppress the growth of test organisms.

The variation in the population of test organisms at different intervals of storage(Table 1, Fig. 1) might be due to the differential availability of nutrients in the carriermaterials, provided for the growth of these organisms. Obviously the limited supplyof nutrients at later stages causes a decline in the growth of these organisms. The

H t-<Q "q (;;' :'>-d ::6 gj § p.. > ~ <

+C1

> ::l et- E

1210

86

Pe

rio

dI

we

eks

)

4

Azo

tob

acte

rch

roo

co

ccu

mI

A-4

1I

2

L '"L L o U ..... o Ql

<,

L '" U o .0 o (; N «

120

108..

6

Pe

rio

dIw

ee

ks)

4

.6S

OIL

..C

H...

RC

O...

.L

oa-

8'c

oJ

loC

•",Z

"c

Rh

izo

biu

msp

.(

M-1

01

2

."?

L L o U o Z..... o Ql

<, o ii o N

120

s: 0::

r-, o X

108

6

Pe

rio

d(w

ee

ks)

42

360~

Rh

izo

biu

mjo

po

nic

urn

15

6-1

6I

32

0

L ~2

80

L c, 0 o .....2

40

0 Ql

..... .s2

00

.0 0 N :.c1

60

0::

r-, 9

12

0X - 0

80

z

40 0

Fig

,2.

Infl

uen

ceo

fty

pe

of

carr

ier

mat

eria

lsan

dte

mp

era

ture

of

sto

rag

eo

ng

row

than

dsu

rviv

alo

fR

hiz

ob

ium

jap

on

i­cu

m,

Rh

izo

biu

msp

.,an

dA

zoto

bact

erch

rooc

occu

m.

......

o ......

102 K. S. JAUHRI and K. PHILIP

Table 2. Influence of storage temperature/additive on the growth and survival of Rhizobium [aponi­cum (SB-16), Rhizobium sp. (M-I0), and Azotobacter chroococcum (A-41)

Period (Weeks)

o1248

12

o1248

12

o1248

12

TemperatureAdditivePeriodPeriod X temperaturePeriod X additive

Log No. of viable bacteriajg of carrier

Storage Temp. (0C) Additive

5-8 32 42 Charcoal Soil

Rhizobium japonicum (SB-16)

5.83 5.83 5.83 5.88 5.787.16 7.08 7.37 7.45 6.977.64 7.89 8.90 8.37 7.927.82 8.88 8.36 8.99 8.388.03 9.18 7.36 8.47 7.917.86 8.11 6.11 7.68 7.03

Rhizobium sp. (M-1O)

6.08 6.08 6.15 6.28 5.927.20 7.15 7.08 7.37 6.917.49 7.62 7.89 7.90 7.437.74 8.36 7.92 8.17 7.857.92 9.20 7.90 8.56 8.107.90 8.76 6.19 7.82 7.41

Azotobacter chroococcum (A-4l)

5.59 5.51 5.51 5.57 3.516.64 6.16 6.62 6.51 6.446.69 7.35 7.83 7.44 7.146.89 8.14 8.42 8.01 7.627.06 8.52 8.02 8.06 7.687.19 8.15 7.03 7.68 7.24

R. [aponicum Rhizobium sp. A. chroococcum

S.Em. C.D. at S.Em. C.D.at S.Em. C.D.at

5% 5% 5%

0.04 0.12 0.04 0.10 0.03 0.080.04 0.10 0.03 0.08 0.02 0.070.06 0.17 0.05 0.15 0.04 0.110.11 0.30 0.09 0.75 0.07 0.210.09 0.25 0.07 0.20 0.06 0.17

results are in agreement with the findings by PRAMANIK and ISWARAN (1973) andTILAK and SUBBA RAO (1978), who also reported a decline in the growth of R. japoni­cum after a period of four months in a mixture of pressmud and charcoal (50: 50).

The results (Table 2, Fig. 2) of the present investigations show a marked influenceofthe storage temperature on the growth and survival of R. japonicum, Rhizobium sp.,and A. chroococcum-in-the various carriers tried. The test organisms differed widelyin their ability to multiply and flourish in various carriers, depending on the tempe­rature of storage. It is because the optimum temperature for each strain is knownto depend on the climatic origin of the plant from which it was isolated (GUKOVA1945, ISHIzAwA 1953, BOWEN and KENNEDY 1959).

Pressmud : A P otential Carrier. II. 103

Rhizobium sp ( M - ta l

jJJJdIIJI JI ~Jd~

5 6 7 8 2 4 5 6

Add it iv e Ca r r ier

Azotobacter chr oococcum ( A - 4\ I

Rh izobium j aponicum (58 t6)

t . Pea t2 . A d di t ive3. Pr es smud (100%)4 . + Addit ive (75: 2 515. (50:5016 . ( 2 5 : 7517. Charcoal8 . Soil

40

<,L.ClJ

0U0

.IJ

BO ]0"0N L.

« ClJ'C 4 0

cd<0 L.a 0_ u

dillLx .... dIIiII-: 0 00z ci>2 3 4

Carrier

5201 o 5 - 8 · C

lIIll 3 2 ·C

2BO1• 42 ·C

l.@ Soil

240 ~ Charcoal

160

120

8 00>

.....0".0 L. 40o ClJN . _

. - L.

odr. L.0:: 0 0u

10 ....

2~ 1a 0

x

ci240

Z

200 18 0

Fig. 3. Influence of carrier /a dditive/temperature on the growth and survival of Rhizobium [ap o­nicum, Rhizobium sp., an d A zotobacter chroococcum .

A study of t he interact ion of various carriers wit h va rying tempera t ures of st ora ge(Table 3, Fi g. 3) reveals that the maximum growth of organisms is in pressmud pluschar coal (75 : 25) at 32 °C. Th e organisms failed t o proliferate at 5-8 °C and no sta­tis t icaIly sign ificant ga in in their growth was noticeable till the end of 12 weeks (Fig . 2).This may be du e to the retardation wit h multiplication and metabolic act ivi ty of thebacter ial cells (VAN SCHREVEN 1970) . This is the reason why storage of ino oulants at

104 K. S. J A UIIRI and K. PIIILIP

Table 3. Influen ce of ca rrier materialsjstorage temperature/additives on growth and surviva l ofRhizobiumjaponicum (SB-16), Rhizobium sp . (M.IO), and Azotob acter chroococcum (A-41)

Type of carrier Log No. of v iable bacteriajg of carrier

Storage temperature (OC) Additive

5-8 32 42 Charcoal Soil

Rhieobium j aponicuni (SB -16)

P eat (100 %) 7.28 7.68 7.47 7.47 7.41Additive (100 % ) 6.95 8.00 7.34 8.12 6.73Pressmud (100 % ) 6.99 6.97 7.17 7.05 7.05Pressmud + a ddit ive (75 : 25) 8.02 8.72 8.04 8.41 8.10P ressmud + additive (50 : 50) 7.68 8.08 7.69 8.16 7.47Pressmud + a ddit ive (25: 75) 7.42 7.53 7.23 7.62 7.18l\lean 7.39 7.83 7.49 7.81 7.33

AdditiveCharcoal 7.69 8.10 7.63Soil 7.09 7.57 7.35

Rhizobium sp. (M-IO)

Peat (100 %) 7.47 7.73 6.93 7.38 7.37Additive (100%) 6.93 7.57 6.91 7.79 6.49Pressmud (100 %) 7.15 7.35 6.85 7.12 7.1 2Pressmud + additive (75 : 25) 7.86 8.38 7.76 7.13 7.87Pressmud + additive (50 : 50) 7.69 8.30 7.57 7.97 7.73Pressmud + additive (25 : 75) 7.25 7.83 7.07 7.72 7.05Mean 7.39 7.86 7.18 7.68 7.27

Ad d itiveCharcoal 7.60 8.13 7.32Soi l 7.18 7.59 7.05

A zotobacter chroococcum (A-41)

Peat (100%) 6.59 7.30 7.04 7.03 7.03Additive (100 %) 6.61 7.37 7.12 7.24 6.83Pressmud (100 % ) 6.45 7.10 6.97 6.85 6.84Pressmud + additive (75: 25) 6.95 7.49 7.61 7.56 7.14Pressmud + additive (50: 50) 6.85 7.45 7.41 7.34 7.04Pressmud + additive (25: 75) 6.62 7.13 7.27 7.26 6.73Mean 6.68 7.31 7.24 7.21 6.94

Ad d itiveCharcoal 6.7 1 7.40 7.53Soil 6.65 7.21 6.94

R. japonicum Rhizobium sp. A. chroococcum

S.Em. C.D.at S.Em. C.D.at S.Em . C.D.at

5 % 5% at 5 %

Carrier 0.06 0.17 0.05 0.15 0.04 0.12Storage t emperature 0.04 0. 12 0.04 0.10 0.02 0.09Additive 0.04 0.10 0.03 0.08 0.02 0.07Carrier Xtemperature 0.11 0.30 0.09 0.25 0,07 0.21Carrier Xadditive 0.09 0.25 0.07 0.20 0.06 0.17Temprature Xadditive 0.06 0.17 0.05 0. 15 0.04 0.12

Pressmud: A Potential Carrier. II. 105

refrigeration temperatures is considered to be the best method for the preservationof inoculants.

The storage of inoculants at 42°C in the present study (Fig. 2) caused a rapiddecline in growth of R. japonicum. This may be due to the detrimental effect of hightemperature (ISWARAN et al. 1970). Contrary to this MARSHALL (1964), however,reported that R. japonicum was relatively resistant to drying and high temperature.In the case of Rhizobium sp. and A. chroococcum. the multiplication of cells continuedup to 4 weeks of storage, which probably indicates the resistance of these organismsto higher temperatures of storage.

It is apparent from the results (Table 4) that the maximum number of rhizobiaon seed and nodules in plants in both the crops was observed with inoculants, preparedfrom pressmud plus charcoal (75: 25) and stored at 32°C. TILAK and SUBBA RAO1978) also reported highest number of nodules per plant of Glycine max, where theseeds were inoculated with pressmud plus charcoal (50: 50) base inoculants and storedat 30°C. There is, however, no report available on the efficiency of pressmud pluscharcoal base inoculants with Vigna radiata.

Table 4. Efficiency of different carrier-based inoculants, stored at different temperatures on thenodulation of soybean var. Bragg and Moong var. PS·16

Type of carrier

Peat (100%Charcoal (100%)Pressmud (100 %)Soil (100 %)Pressmud + charcoal (75: 25)Pressmud + charcoal (50: 50)Pressmud + charcoal (25: 75)Pressmud + soil (75: 25)Pressmud + soil (50: 50)Pressmud + soil (25: 75)

Peat (100%)Charcoal (100 %)Pressmud (100 %)Soil (100%)Pressmud + charcoal (75: 25)Pressmud + charcoal (50: 50)Pressmud + charcoal (25: 75)Pressmud + soil (75: 25)Pressmud + soil (50: 50)Pressmud + soil (25: 75)

Temperature (0G) of storage for inoculants

5-8 32 42

No. of No. of No. of No. of No. of No. ofviable nodules viable nodules viable nodulesrhizo- per rhizo- per rhizo- perbiaJseed plant biaJseed plant bia/seed plantX104 X 104 X 104

(Av, of three replicates)

Soybean (Glycine max) var. Bragg

2.50 26.00 47.50 50.00 2.00 0.004.50 36.00 87.50 79.00 11.50 0.007.50 19.00 57.50 77.00 11.50 0.003.00 13.00 32.00 20.00 2.50 0.00

24.00 41.00 135.50 96.00 22.00 0.0017.50 40.00 65.00 60.00 15.50 0.0016.50 39.00 53.50 49.00 13.00 0.0017.50 38.00 117.50 80.00 13.50 0.0017.50 34.00 63.00 80.00 12.00 0.0016.50 26.00 33.00 70.00 10.50 0.00

Moong (Vigna radiata) var. PS-16

1.55 10.00 13.00 19.00 4.00 17.000.50 11.00 18.00 12.00 5.00 11.001.75 17.00 18.50 28.00 5.00 20.001.50 7.00 14.50 9.00 7.00 7.008.75 26.00 37.50 47.00 18.50 25.008.35 21.00 30.00 36.00 8.50 15.003.70 13.00 20.00 19.00 4.50 13.006.00 21.00 36.50 29.00 11.00 17.005.15 20.00 27.50 21.00 5.00 11.004.25 13.00 12.50 19.00 4.00 9.00

106 K. S. JAUHRI and K. PHILIP

Better nodulation with inooulants, stored at 32 DC over those stored at 5-8 and42 DC, may be attributed to the higher population of rhizobia (Table 3). One notabledeviation from the normal trend of correlation between number of rhizobia per seedand nodule per plant was observed in the case of soybean, where the inoculantsstored at 42 DC showed high number of rhizobia on seed, but failed to nodulate theplants (Table 1).

Although the successful nodulation depends on many factors, the failure of in­oculants to nodulate soybean in the present study may be attributed to the soybeanrhizobia, possibly being inffective due to high storage temperature (42 DC) of prolongedduration. Although there is no report on this effect, DART et al. (1976) showed thatsoybean rhizobia becomes ineffective at 33 DC for nitrogen fixation. The inoculantsstored at higher temperature (42 DC) not only adversely affected the population ofrhi­zobia on seed, but also reduced the efficiency of the organisms for nodulation. Lowertemperature (5-8 DC) was found suitable for storage, but suppressed multiplicationof rhizobia in inoculants.

Conclusions

It can be concluded from the above study that pressmud can be advantageouslyused as carrier material for commercial production of inoculants only after amendmentwith charcoal. Pressmud with charcoal (75: 25) proved effective as carrier not onlyfor maintaining higher population of test organisms, but also by improving the shelf­life for a longer period of storage. Maximum growth of the test-organisms was 0 btainedat 32 DC. Higher temperature of storage (42 DC) adversely effected the growth andsurvival of the test organisms. Refrigeration temperature (5-8 DC) was the mostsuitable for the storage of inoculants for a longer period.

References

BOWEN, G. D., and KENNEDY, M. M.: Effect of high temperature on Rhizobium. Qd. J. Agric.Sci. 16 (1959), 177.

DART, P. J., DAY, J. M., ISLAM, R., and DOBEREINER, J.: Symbiosis in tropical grain legumes:Some effects of temperature and the composition of the rooting medium. In: Symbiotic Nitro­gen Fixation in Plants (P. S. NUTMAN, ed.). Cambridge 1976, p. 361.

DUBE, J. N., MAHERE, D. P., and RAWAT, A. K.: Development of coal as a carrier for rhizobialinoculants. Sci. and Cult. 46 (8) (1980), 304.

GUKOVA, M. M.: The effect of soil temperature on nitrogen fixation by nodule bacteria. Dokl.s.-kh. Akad. Timiryazeva 30 (1945), 33.

ISHIZAWA, S.: Studies on root nodule bacteria of leguminous plants. 1. Characters in artificialmedia. II. Effect of temperature on the growth of Rhizobium. J. Sci. Soil Manure (Tokyo) 24(1953), 227.

ISWARAN, V., SUNDRA RAo, W. V. B., and JAUHRI, K. S.: Nodulation under unfavourable soilconditions. Curro Sci. 39 (4) (1970), 93.

MARSHALL, K. C.: Survival of root nodule bacteria in dry soils. Austr. J. Agric. Res. 15 (1964),273.

PHILIP, KOSHY, and J AUHRI, K. S.: Pressmud: A potential carrier for Rhizobium and Azotobacter.1. Comparative analytical studies of various carrier materials. Zbl. Mikrobiol. 139 (1984),35-41.

PRAMANIK, M., and ISWARAN, V.: Survival of Rhizobium japonicum in various carriers. Zbl.Bakt. II 128 (1973), 232.

ROUGHLEY, R. J., and VINCENT, J. M.: Growth and survival of Rhizobium spp. in peat culture.J. appl. Bact. 30 (2) (1967), 362.

Pressmud: A Potential Carrier. II. 107

SANORIA, C. L.: Seed bacterization and synthesis of growth-promoting substances by differentstrains of Azotobacter chroococcum. Ph.D. Thesis (Microbiology), Indian Agricultural ResearchInstitute, New Delhi (India) (1965), 68.

TILAK, K. V. B. R., and SUBBA RAo, N. S.: Carriers for legume (Rhizobium) inoculants. Fert.News 23 (2) (1978), 25.

VAN SCHREVEN, D. A.: Some factors affecting growth and survival of Rhizobium spp. in soil-peatcultures. Plant and Soil 32 (1970), 113.

Eingegangen am 22. 10. 1982.

Authors' address:

Dr. K. S. JAUHRI and Mr. KOSHY PHILIP, Division of Microbiology, Indian Agricultural ResearchInstitute, New Delhi· 110012, India.