- Email: [email protected]
Sensitivity of Rhizobia to Different Concentrations of Carbamates at Different Temperatures of Incubation
Zentralbl. :\Iikrobiol. 142 (1987), 4(\5 -4'jO VEB Gustav Fischer Verhg Jena [Departlllent of
~lier()biology,
Panjah University, Chandigal'h, India]
Sensitivity of Rhizobia to Different Concentrations of Carbamates at Different Temperatures of Incubation KRISHKA
SHARMA, NEERl' NARI-LA and K. C.
(iUPTA
Summary f-lensitivity of various RhizoiJium strains to different eoncentmtions (0.05, 0.5, 4.5/lg/llli) of carbamates namely '['!\lTD, Sevin and Dyrnid at three incubation temperatures (25, 28 and 37°C) were studied. The studies indicate that although no correlation exists between incubation temperature and pesticide concentmtion on the sensitivity of rhizobia, yet it strongly supports the view that incubation telllperature influences the sensitivity of a partieular' RhizoiJium strain in the presence of a particular concentration of a p8sticide.
Zusammenfassung Bei (lrei Inkubationstelliperaturen (25, 28 unrl 37 °e) wurde die Ellipfindlichkeit versehiedener Rhizobium-Stiitnme gegeniiber steigenden Konzentrationen (0,05, 0,5 lind 4,5/lg/111I) del' Karballlate TMTD, Sevin lIn,1 Dymid untersucht. Es bestand zwar keine Korrelation zwischen Tnkubationstemperatnr und Pestizidkonzentration, jedoch bestiitigte sich in hohem Maf3e die Ansicht, daf3 die Inkubationstemperatul' die EllIpfindlichkeit eines bestilllmten Rh£zob'ium-Stalllllles in Anwesenheit einer bestillltnten Pestizidkonzentration beeinflllssen kann.
Pesticides are known to influence soil biological processes like nitrification (Ft:NRE et al. 1970; AONIIIOTRI 1974), ammonification (BALICRA et al. 1969; WAINWRIGHT and PGGH 19n) and dinitrogen fixation (PAlmEK 1968; HAMDl et al. 1968; CASSEMAN et al. 1981). Treatment of soils with pesticides has shown to alter population of microorganisms related to soil fertility (BARTHA et al. 1967; BOLI,HN 19fH ; Tn 1970; Te and BOLLEN 1968). Since a pestieide in soil is not the only factor influencing the Roil popnlation, other factors like pH and temperature may be interacting with the pesticide for itR activity. The interaction of temperature with peRticides for their aetivity on soil microorganiRl1ls haR been studied in few easeR. For example WOLCOT'f ct a!. (1960) demonstrated this interaction by showing that fall application of fumigants reduces the nitrate level in soil until .J uly of the next year. A temperat lire dependent effect of residual nematoeides on the activitiefl of Roil mief'oorganistllR has been studied by Tc (19n). Since pesticides are applied to floils and plants at different periods of the year during which temperature variefl, it ifl desirable to know the effeet of pesticideR 011 soil microorganisms at different temperatures. Thl'refore, ill thc present investigation, the effect of different concentrations of Tetramethyl thiuram disulfide (TMTD), Sevin and Dymid on rhizobia nt different telllpemtUl'eR \mfl fltudied.
Materials and Methods Organisms and Culture Conditions HtandaJ'd isolatpR of rhizohia, t lH--"il' Rl1'
listed below:
lllllllbpl',
('ross inol'uhttioll gl'Olip and BOlU'('e are
466
K. SHARMA, N. NARl:LA and K. G. GUPTA
Strain No.
Name
Origin
D-228 Su-391 D-338 D-4(j7 D-232 Cp-20,5 & 10 U-4
R. legumino8arum R. legumino8arum R. japonicum L. corniculatu8 R. meliloti R. spec. from Vigna 8inen8i8 R. spec. from Pha8eolu8 mungo
Czechoslovakia Czechoslovakia Czechoslovakia Czechoslovakia Czechoslovakia Chandigarh Chandigarh
Pesticides The pesticides used are listed below along with chemical structure, percent active ingredient and water solubility: Pesticides
Chemical Structure
S Tetramethyl thiuram disulfide
Water solubility
99%
Insoluble
0.1% Very less
S
II
II
(CH 3hNC-S-CN( CH 3h
o
Purity
H
I
I
Sevin(l-naphthyl-N-methylcarbamate)
C-O-N--CH3
98.6%
Dymid (N-N-dimethyl 2,2 diphenyl acetamide)
H
99%
0
I I
C-C-N
/
eH 3
~
CH 3
These organism were maintained on asparagine-mannitol agar at 4 DC. The incubation temperature was 28 ± 1°C.
Sensitivity of rhizo bia and growth measurements for various crops Rhizobium strains were grown in asparagine-mannitol broth for 72 h and the optical density of the culture was determined and adjusted to 0.3 at 620 nm by using SPEKOL. 50 ml of asparaginemannitol broth was dispensed in conical flasks and sterilized. Pesticides were added in alcoholic solution to the broth so as to achieve final concentrations of 0.0, 0.05, 0.5, 5.0, 25.0, 125.0 and 250.0 ftg/ml of TMTD, 0.0, 0.01, 0.1, 1.0, 10.0, 50.0 and 100.0 ftg/ml of Dymid and 0.0, 0.01, 0.1, 1.0,10.0,100.0 and 1,000.0 ftg/ml of Sevin. The control contained only the same volume of solvent (which was used for dissolving the pesticides). The pesticides were separately sterilized, using G-5 sintered glass filter (and added under aseptic conditions). The sensitivity of rhizobia to pesticides was determined in terms of growth. To each flask 0.1 ml of the culture (optical density 0.3) was added. The flasks were incubated at 25, 28 and 37 DC. After 24 h, samples were withdrawn aseptically and optical density was determined at 620 nm· Readings were taken up to 5 d. The results expressed are mean values of triplicates.
Results and Discussion In literature a number of references are available, dealing with the sensitivity of rhizobia to pesticides or a physical factor like pH and temperature. The effect of one factor on a living system may be influenced by the presence of another factor. Since this type of information, i.e., the influence of two factors together on rhizobia, is not available, studies were made to see the sensitivity of rhizobia to different concentrations of pesticides at different incubation temperatures.
Different Concentrations of Carbamates
467
Effect of different concentrations of TMTD at different temperatures on sensitivity of Rhizobium strains The results (Table 1) show that in case of TMTD complete inhibition of the Rhizobium strains studied was obtained at 25,ug/ml and the change in incubation temperatures did not influence the sensitivity of rhizobia. At concentrations between 5 and 25,ug/ml, some growth was observed in certain strains but it was not an appreciable amount of growth and again change in incubation temperature had not influenced the sensitivity. 0.5,ug/ml of TMTD at 28°C decreased the growth in all the strains except D-232. Table 1. Effect of different concentrations of TMTD on the per cent increase (+) or decrease (-) of growth of rhizobia at different incubation temperatures Concentration of TMTD Wl/ml)
Strain number CP-5
D-338
CP-I0
U-4
D-232
Su-391
CP-20
-12.83 - 8.57 -20.00
-15.H5 -12.00 - 8.19
0.05
a) b) c)
- 3.fH\ -22.70 -10.70
+ 2.98 - l,(\9 -19.30
- 8.60 -20.80 -20.37
+l(i,HO - 3.44 + 11.53
0.5
a) b) c)
+ 12.20 -13.fiO -15.30
- 7.4H - 8.45 -22.50
+18.20 - 4.1(\ -11.10
+ 2.28 - 1.72 - 5.7fi
+ 1.20 + 3.60 -17.60
-22.85 -38.56 -30.00
-13.04 - 8.00 - 3.20
5.0
a) b) c)
+14.70 + 9.09 -29.90
-22.50 -21.10
+29.00 + 9.1H -54.28
- 7.14 -27.58 -38.40
+12.19 - H.09 -24.40
-50.00 -51.41 -30.00
+ 1.73 + U\O -29.50
-HUO
Note: At TMTD concentrations above 25pg/llll 100% decrease in growth was observed. a) = Incubation temperature 25°C, b) "' Incubation temperature 28°C, c) = Incubation temperature 37 °C. Level of significance at 0.01.
The increase in growth at higher concentrations of TMTD may he explained on the basis of earlier observations (SUD and GUPTA 1972) in which carbamates have been shown to calise the accumulation of pyruvic acid which in turn interacts with the carbamate and inactivates it. From this data, it may be suggested that strains differ in their sensitivity to TMTD at different incubation temperatures. Effect of Sevin at different temperatures on the sensitivity of different Rhizobium strains Sensitivity of rhizobia to Sevin at different incubation temperatures was also studied. The results (Table 2) show that, with increase in Sevin concentration, i.e., up to 1.0,ug/ml, temperature 37°C, a large number of strains showed increase in growth, compared to lower concentration at 25 and 28°C. The increase in growth at higher concentrations may again be explained on the basis of pyruvic acid accumulation, but in this case temperature of incubation Reems to playa role, too. Effect of different concentrations of Dymid at different temperatures on sensitivity of various Rhizobium strains In case of Dymid (Table 3) all the strains of Rhizobium, except lJ-4 and CP-1O, show 100 % decrease in growth in presence of 50,ug/ml at 25 °C, but at 28 DC, strains D-232, D-228, Su-:~91, CP-20, CP-5 and U-4 show 100 %, decrease in growth. At 37 °C, all strainR, except strains D-467 and CP-1O, show 100 % decrease in growth. 1n pres-
a)
~
10.0
1.0
0.1
0.01
8.47
--100
--100
-100
-
--lH.l
-18.0
+37.28
+17.6
+44.0
7.75
-38.7
-22.6
-19.3
-15.0
+38.0 -36.1
+31.0
-31.9
-
-23.25
°c, c)
-100
-100
-100
-4!J.5
-34.8
Incubation temperature 28
-100
-100
Incubation temperature 25°C, b)
--100
-54.2
-40.5
-100
-54.8
-55.7
h)
c)
-100
-31.H
-:H.2
a)
-100
-49.1
+
c)
-23.1
5.4
+
+11.5
a)
b)
-22.7
1.70
-
-30.5
3.2
e)
:l.07
4.41
+ -
>L )
-21.9
8.47
1.1
-
b)
5.06 +24.0
c)
+10.0 - 5.88
-25.0
-
-13.5
b)
4.06 8.50
2.3
+ -
+ 1.02 -23.0
a)
D·338
D·232
8.16
4.34
3.29
3.06
8.6
5.66
1.77
6.50
1.18
2.75
--100
--100
-100 ---100
--100
-75.88
-73.75
--71.98
+
+
+
-59.48
--61.95
--61.\)5
-69.30
-76.70 --76.11
Su-391
+40.0 -100
-38.4
-- 20.7
+24.4
+ 7.69 -42.:3
6.66
-30.7
+
+
5.76
0.0
D-467
9.6
~
--75.0 --74.6
-n.8
-100 --100
--11.1
--42.5
---:36.5
:>-
f-3
'"
q
<0
P -22.2
(i -1\1.6
~
;:l
t-< :>-
;:i
:>-
Z
~
.?'
~
~
:>-
;:q
C'lJ
-12.5
-I- 8.:1
+22.5 +1[).1
--100
--33.3
-
-10.8
+27.7
+ 18.0
0.0
4.0
+16.0
+
0.0
+36.1
+:16.0
-20.0 -19.44
+60.0
CP-I0
-24.0
CP-20
Incubation temperature 37°C. Lovel of significance at 0.01.
-100
--100
--100
-13.04
-18.6
--18.3
+
-10.9
+
-
-
D·228
r
CP·5
U·4
Concentration of Sevin (flg/ ml )
Strain Number
~
~
00
Tahle 2. Effect of different concentration8 of Scvin upon the per cent increase (+) or decrease (-) of growth of rhizobia at different incubation temperatures
--20.0
I- 5.:3
-:30.0 --24.2
b)
c)
a)
b)
c)
-41.6
+-1 :3.:3 -25.0
---38.4 -36.9 -52.5
--:32.7
-41.6
-45.0
7.69 -11.5
-27.2
-a1.0 -18.1
-40.0
-a:3.a
a)
8.08
-
:3.8
-2:3.07
5.0
-
c)
-·22.4
+
+46.1 +44.6
D-232
-12.7
---22.8 --17.5
a)
+11.1
c)
D-338
h)
+40.0 ---15.0
b)
a)
CP-5
Strain number
+:38.6
-17.8
--33.3
+84.6
-:3a.3 -17.8
+46.1
-14.7
-:31.6
D-228
+25.8 -i-12.9
+29.0
+16.5
+29.0
+35.4
+ 25.4
+:39.0
+45.1
+:39.0
+77.4 +68.0
8u-391
---37.5
+17.3 --19.6
0.0
0.0 --19.6
2.:3 ---10.7 -4:3,7
D-467
:3.82 8.84 5.00 -18.08
+ +
+ 12.24 1.21
+10.aO -- 4.20
+22.40
+ 17.02
+:36.05 + 19.:37
CP-20
+23.8 --15.7
-12.17
--26.8 ----4:3.6
--- :3.2
-23.9
0.0 -27.:3
+1:3.3
-;-16.6
--17.58
+8:3.9
+
+20.0 +42.7 --10.5
->-- 58.5
-1- 56.4
5')
+
+n.2
+:30.0
CP-I0
+52.6
+9:3.5 -I- 56.8
U-4
-"ote: At Dymid concentration;; above 50/!g/mI, 100% decrease in growth was observed except in strain D-338 where -47.2 decrease in growth wa;; ob""l'v('d at 28 0('. In ])-467 growt.h was there at 28 and 37°C. Some growth wa;; ob"erve<1 at 25°C in ease of U-4. CP-I0 showed 60.0, 66.6 and 6:3.1 % dt'en'a,,' in grO\yth at l(lO/lg/ml of DYIllid. a) - Incubation tOlllp('l'atul'o 25 "C, b) 1t1('ubation ttnnpol'aturo 28 "c, c) ~ Incubationtempnrat-ul'o :37 O( '. L('vol of ;;ignificanco at 0.01.
1O.U
1.0
0.1
0.01
l'ollcentrat ion of Dymid (flg/mI)
Table :3. Effect of different concentmtion8 of Dymid upon the pel' cent increa;;e (+) or decrem,e (-) in growt.h of rhizobia at different incubation t emperat ure;;
co
C'>
>I"'-
§. '"'"
~
or<
on o,..,
S
~
r'-
::J
'"
Q
::J
o
\~
[
'-'
;:;; CO' ...
470
K. SHARMA, N. NARULA and K. G. GUPTA, Different Concentrations of Carbamates
ence of O.I,ug/ml of Dymid per cent decrease or increase in growth varied with the incubation temperatures. From the results no generalization with regard to the relation between temperature and pesticide concentration on the sensitivity of rhizobia can be made, but the results strongly support the view that incubation temperature influences the sensitivity cf a strain to a particular concentration of a pesticide.
References AGNIHOTRI, V. P.: ThiraDl induced changes in soil microflora their physiological activity and control of damping-off in chillies (Capsicum annuum). Indian J. Expt. Biology 12 (1974), 85-89. BALICKA, N., and SOBIESZCANSKI, J.: The effect of herbicides on soil microflora. III. The effect of herbicides on ammonification and nitrification in the soil. Acta Microbial. Pol. Ser. B 18 (1969), 7-10. BARTHA, R., LANZILOTTA, R. P., and PRAMER, D.: Stability and effects of some pesticides in soil. Appl. Microbial. 15 (19Ii7), 67-75. BOLLEN, E. B.: Interactions between pesticides and soil microorganisms. Annu. Rev. Microbial. 15 (1961), 69-72. CASSEMANS, J., and VAN ASSCHE, C.: Comparative study of growth-promoting capacity of SODle insecticidesfnematicides. Plant and Soil 63 (1981), 251-259. FUNKE, B. R., LIN, SHIN CHSIANG, SCHULZ, J. T.: Effect ofsoDle organophosphorus and carbamate insecticides on nitrification and legume growth. Bact. Proc. (Abstract) (1970), A62, p.9. HAMDI, YOUSEF A., and TEWFIK, M. S.: Effect of the herbicides trifluralin on nitrogen-fixation in Rhizobium and Azotobacter and on nitrification. Acta Microbiol. Pol. Ser. B 1 (1969), 53-58. MARTIN, J. P.: Influence of pesticides on soil microbes and soil properties. In: Pesticides and their effect on soil and water. Soil Sci. Soc. Amer. Inc., Madison, Wisconsin, 1966, 95-108. PAREEK, R. P.: Effect of DDT on symbiotic nitrogen fixation and nitrification in soil. M.Sc. Thesis, Department of Microbiology, LA.R.I., Delhi (1968). SUD, R. K., and GUPTA, K. G.: On the sensitivity of isolates of Rhizobium spp. and Azotobacter chroococcum to TMTD and its degradation product NaDDC. Arch. Mikrobiol. 85 (1972),19-22. Tu, C. M.: Effect of four organophosphorus insecticides on microbial activities in soil. App!. Micro· bioI. 19 (1970), 479-484. Effect of four nematocides on activities of microorganisms in soil. Appl. Microbial. 23 (1973), 398-401and BOLLEN, W. B.: Effect of paraquat on microbial activities in soils. Weed Res. 8 (1968), 28-37. ~. AINWRIGHT, M., and PUGH, G. J. F.: The effects of three fungicides on nitrification and ammonification in soil. S
~lier()biology,
Panjah University, Chandigal'h, India]
Sensitivity of Rhizobia to Different Concentrations of Carbamates at Different Temperatures of Incubation KRISHKA
SHARMA, NEERl' NARI-LA and K. C.
(iUPTA
Summary f-lensitivity of various RhizoiJium strains to different eoncentmtions (0.05, 0.5, 4.5/lg/llli) of carbamates namely '['!\lTD, Sevin and Dyrnid at three incubation temperatures (25, 28 and 37°C) were studied. The studies indicate that although no correlation exists between incubation temperature and pesticide concentmtion on the sensitivity of rhizobia, yet it strongly supports the view that incubation telllperature influences the sensitivity of a partieular' RhizoiJium strain in the presence of a particular concentration of a p8sticide.
Zusammenfassung Bei (lrei Inkubationstelliperaturen (25, 28 unrl 37 °e) wurde die Ellipfindlichkeit versehiedener Rhizobium-Stiitnme gegeniiber steigenden Konzentrationen (0,05, 0,5 lind 4,5/lg/111I) del' Karballlate TMTD, Sevin lIn,1 Dymid untersucht. Es bestand zwar keine Korrelation zwischen Tnkubationstemperatnr und Pestizidkonzentration, jedoch bestiitigte sich in hohem Maf3e die Ansicht, daf3 die Inkubationstemperatul' die EllIpfindlichkeit eines bestilllmten Rh£zob'ium-Stalllllles in Anwesenheit einer bestillltnten Pestizidkonzentration beeinflllssen kann.
Pesticides are known to influence soil biological processes like nitrification (Ft:NRE et al. 1970; AONIIIOTRI 1974), ammonification (BALICRA et al. 1969; WAINWRIGHT and PGGH 19n) and dinitrogen fixation (PAlmEK 1968; HAMDl et al. 1968; CASSEMAN et al. 1981). Treatment of soils with pesticides has shown to alter population of microorganisms related to soil fertility (BARTHA et al. 1967; BOLI,HN 19fH ; Tn 1970; Te and BOLLEN 1968). Since a pestieide in soil is not the only factor influencing the Roil popnlation, other factors like pH and temperature may be interacting with the pesticide for itR activity. The interaction of temperature with peRticides for their aetivity on soil microorganiRl1ls haR been studied in few easeR. For example WOLCOT'f ct a!. (1960) demonstrated this interaction by showing that fall application of fumigants reduces the nitrate level in soil until .J uly of the next year. A temperat lire dependent effect of residual nematoeides on the activitiefl of Roil mief'oorganistllR has been studied by Tc (19n). Since pesticides are applied to floils and plants at different periods of the year during which temperature variefl, it ifl desirable to know the effeet of pesticideR 011 soil microorganisms at different temperatures. Thl'refore, ill thc present investigation, the effect of different concentrations of Tetramethyl thiuram disulfide (TMTD), Sevin and Dymid on rhizobia nt different telllpemtUl'eR \mfl fltudied.
Materials and Methods Organisms and Culture Conditions HtandaJ'd isolatpR of rhizohia, t lH--"il' Rl1'
listed below:
lllllllbpl',
('ross inol'uhttioll gl'Olip and BOlU'('e are
466
K. SHARMA, N. NARl:LA and K. G. GUPTA
Strain No.
Name
Origin
D-228 Su-391 D-338 D-4(j7 D-232 Cp-20,5 & 10 U-4
R. legumino8arum R. legumino8arum R. japonicum L. corniculatu8 R. meliloti R. spec. from Vigna 8inen8i8 R. spec. from Pha8eolu8 mungo
Czechoslovakia Czechoslovakia Czechoslovakia Czechoslovakia Czechoslovakia Chandigarh Chandigarh
Pesticides The pesticides used are listed below along with chemical structure, percent active ingredient and water solubility: Pesticides
Chemical Structure
S Tetramethyl thiuram disulfide
Water solubility
99%
Insoluble
0.1% Very less
S
II
II
(CH 3hNC-S-CN( CH 3h
o
Purity
H
I
I
Sevin(l-naphthyl-N-methylcarbamate)
C-O-N--CH3
98.6%
Dymid (N-N-dimethyl 2,2 diphenyl acetamide)
H
99%
0
I I
C-C-N
/
eH 3
~
CH 3
These organism were maintained on asparagine-mannitol agar at 4 DC. The incubation temperature was 28 ± 1°C.
Sensitivity of rhizo bia and growth measurements for various crops Rhizobium strains were grown in asparagine-mannitol broth for 72 h and the optical density of the culture was determined and adjusted to 0.3 at 620 nm by using SPEKOL. 50 ml of asparaginemannitol broth was dispensed in conical flasks and sterilized. Pesticides were added in alcoholic solution to the broth so as to achieve final concentrations of 0.0, 0.05, 0.5, 5.0, 25.0, 125.0 and 250.0 ftg/ml of TMTD, 0.0, 0.01, 0.1, 1.0, 10.0, 50.0 and 100.0 ftg/ml of Dymid and 0.0, 0.01, 0.1, 1.0,10.0,100.0 and 1,000.0 ftg/ml of Sevin. The control contained only the same volume of solvent (which was used for dissolving the pesticides). The pesticides were separately sterilized, using G-5 sintered glass filter (and added under aseptic conditions). The sensitivity of rhizobia to pesticides was determined in terms of growth. To each flask 0.1 ml of the culture (optical density 0.3) was added. The flasks were incubated at 25, 28 and 37 DC. After 24 h, samples were withdrawn aseptically and optical density was determined at 620 nm· Readings were taken up to 5 d. The results expressed are mean values of triplicates.
Results and Discussion In literature a number of references are available, dealing with the sensitivity of rhizobia to pesticides or a physical factor like pH and temperature. The effect of one factor on a living system may be influenced by the presence of another factor. Since this type of information, i.e., the influence of two factors together on rhizobia, is not available, studies were made to see the sensitivity of rhizobia to different concentrations of pesticides at different incubation temperatures.
Different Concentrations of Carbamates
467
Effect of different concentrations of TMTD at different temperatures on sensitivity of Rhizobium strains The results (Table 1) show that in case of TMTD complete inhibition of the Rhizobium strains studied was obtained at 25,ug/ml and the change in incubation temperatures did not influence the sensitivity of rhizobia. At concentrations between 5 and 25,ug/ml, some growth was observed in certain strains but it was not an appreciable amount of growth and again change in incubation temperature had not influenced the sensitivity. 0.5,ug/ml of TMTD at 28°C decreased the growth in all the strains except D-232. Table 1. Effect of different concentrations of TMTD on the per cent increase (+) or decrease (-) of growth of rhizobia at different incubation temperatures Concentration of TMTD Wl/ml)
Strain number CP-5
D-338
CP-I0
U-4
D-232
Su-391
CP-20
-12.83 - 8.57 -20.00
-15.H5 -12.00 - 8.19
0.05
a) b) c)
- 3.fH\ -22.70 -10.70
+ 2.98 - l,(\9 -19.30
- 8.60 -20.80 -20.37
+l(i,HO - 3.44 + 11.53
0.5
a) b) c)
+ 12.20 -13.fiO -15.30
- 7.4H - 8.45 -22.50
+18.20 - 4.1(\ -11.10
+ 2.28 - 1.72 - 5.7fi
+ 1.20 + 3.60 -17.60
-22.85 -38.56 -30.00
-13.04 - 8.00 - 3.20
5.0
a) b) c)
+14.70 + 9.09 -29.90
-22.50 -21.10
+29.00 + 9.1H -54.28
- 7.14 -27.58 -38.40
+12.19 - H.09 -24.40
-50.00 -51.41 -30.00
+ 1.73 + U\O -29.50
-HUO
Note: At TMTD concentrations above 25pg/llll 100% decrease in growth was observed. a) = Incubation temperature 25°C, b) "' Incubation temperature 28°C, c) = Incubation temperature 37 °C. Level of significance at 0.01.
The increase in growth at higher concentrations of TMTD may he explained on the basis of earlier observations (SUD and GUPTA 1972) in which carbamates have been shown to calise the accumulation of pyruvic acid which in turn interacts with the carbamate and inactivates it. From this data, it may be suggested that strains differ in their sensitivity to TMTD at different incubation temperatures. Effect of Sevin at different temperatures on the sensitivity of different Rhizobium strains Sensitivity of rhizobia to Sevin at different incubation temperatures was also studied. The results (Table 2) show that, with increase in Sevin concentration, i.e., up to 1.0,ug/ml, temperature 37°C, a large number of strains showed increase in growth, compared to lower concentration at 25 and 28°C. The increase in growth at higher concentrations may again be explained on the basis of pyruvic acid accumulation, but in this case temperature of incubation Reems to playa role, too. Effect of different concentrations of Dymid at different temperatures on sensitivity of various Rhizobium strains In case of Dymid (Table 3) all the strains of Rhizobium, except lJ-4 and CP-1O, show 100 % decrease in growth in presence of 50,ug/ml at 25 °C, but at 28 DC, strains D-232, D-228, Su-:~91, CP-20, CP-5 and U-4 show 100 %, decrease in growth. At 37 °C, all strainR, except strains D-467 and CP-1O, show 100 % decrease in growth. 1n pres-
a)
~
10.0
1.0
0.1
0.01
8.47
--100
--100
-100
-
--lH.l
-18.0
+37.28
+17.6
+44.0
7.75
-38.7
-22.6
-19.3
-15.0
+38.0 -36.1
+31.0
-31.9
-
-23.25
°c, c)
-100
-100
-100
-4!J.5
-34.8
Incubation temperature 28
-100
-100
Incubation temperature 25°C, b)
--100
-54.2
-40.5
-100
-54.8
-55.7
h)
c)
-100
-31.H
-:H.2
a)
-100
-49.1
+
c)
-23.1
5.4
+
+11.5
a)
b)
-22.7
1.70
-
-30.5
3.2
e)
:l.07
4.41
+ -
>L )
-21.9
8.47
1.1
-
b)
5.06 +24.0
c)
+10.0 - 5.88
-25.0
-
-13.5
b)
4.06 8.50
2.3
+ -
+ 1.02 -23.0
a)
D·338
D·232
8.16
4.34
3.29
3.06
8.6
5.66
1.77
6.50
1.18
2.75
--100
--100
-100 ---100
--100
-75.88
-73.75
--71.98
+
+
+
-59.48
--61.95
--61.\)5
-69.30
-76.70 --76.11
Su-391
+40.0 -100
-38.4
-- 20.7
+24.4
+ 7.69 -42.:3
6.66
-30.7
+
+
5.76
0.0
D-467
9.6
~
--75.0 --74.6
-n.8
-100 --100
--11.1
--42.5
---:36.5
:>-
f-3
'"
q
<0
P -22.2
(i -1\1.6
~
;:l
t-< :>-
;:i
:>-
Z
~
.?'
~
~
:>-
;:q
C'lJ
-12.5
-I- 8.:1
+22.5 +1[).1
--100
--33.3
-
-10.8
+27.7
+ 18.0
0.0
4.0
+16.0
+
0.0
+36.1
+:16.0
-20.0 -19.44
+60.0
CP-I0
-24.0
CP-20
Incubation temperature 37°C. Lovel of significance at 0.01.
-100
--100
--100
-13.04
-18.6
--18.3
+
-10.9
+
-
-
D·228
r
CP·5
U·4
Concentration of Sevin (flg/ ml )
Strain Number
~
~
00
Tahle 2. Effect of different concentration8 of Scvin upon the per cent increase (+) or decrease (-) of growth of rhizobia at different incubation temperatures
--20.0
I- 5.:3
-:30.0 --24.2
b)
c)
a)
b)
c)
-41.6
+-1 :3.:3 -25.0
---38.4 -36.9 -52.5
--:32.7
-41.6
-45.0
7.69 -11.5
-27.2
-a1.0 -18.1
-40.0
-a:3.a
a)
8.08
-
:3.8
-2:3.07
5.0
-
c)
-·22.4
+
+46.1 +44.6
D-232
-12.7
---22.8 --17.5
a)
+11.1
c)
D-338
h)
+40.0 ---15.0
b)
a)
CP-5
Strain number
+:38.6
-17.8
--33.3
+84.6
-:3a.3 -17.8
+46.1
-14.7
-:31.6
D-228
+25.8 -i-12.9
+29.0
+16.5
+29.0
+35.4
+ 25.4
+:39.0
+45.1
+:39.0
+77.4 +68.0
8u-391
---37.5
+17.3 --19.6
0.0
0.0 --19.6
2.:3 ---10.7 -4:3,7
D-467
:3.82 8.84 5.00 -18.08
+ +
+ 12.24 1.21
+10.aO -- 4.20
+22.40
+ 17.02
+:36.05 + 19.:37
CP-20
+23.8 --15.7
-12.17
--26.8 ----4:3.6
--- :3.2
-23.9
0.0 -27.:3
+1:3.3
-;-16.6
--17.58
+8:3.9
+
+20.0 +42.7 --10.5
->-- 58.5
-1- 56.4
5')
+
+n.2
+:30.0
CP-I0
+52.6
+9:3.5 -I- 56.8
U-4
-"ote: At Dymid concentration;; above 50/!g/mI, 100% decrease in growth was observed except in strain D-338 where -47.2 decrease in growth wa;; ob""l'v('d at 28 0('. In ])-467 growt.h was there at 28 and 37°C. Some growth wa;; ob"erve<1 at 25°C in ease of U-4. CP-I0 showed 60.0, 66.6 and 6:3.1 % dt'en'a,,' in grO\yth at l(lO/lg/ml of DYIllid. a) - Incubation tOlllp('l'atul'o 25 "C, b) 1t1('ubation ttnnpol'aturo 28 "c, c) ~ Incubationtempnrat-ul'o :37 O( '. L('vol of ;;ignificanco at 0.01.
1O.U
1.0
0.1
0.01
l'ollcentrat ion of Dymid (flg/mI)
Table :3. Effect of different concentmtion8 of Dymid upon the pel' cent increa;;e (+) or decrem,e (-) in growt.h of rhizobia at different incubation t emperat ure;;
co
C'>
>I"'-
§. '"'"
~
or<
on o,..,
S
~
r'-
::J
'"
Q
::J
o
\~
[
'-'
;:;; CO' ...
470
K. SHARMA, N. NARULA and K. G. GUPTA, Different Concentrations of Carbamates
ence of O.I,ug/ml of Dymid per cent decrease or increase in growth varied with the incubation temperatures. From the results no generalization with regard to the relation between temperature and pesticide concentration on the sensitivity of rhizobia can be made, but the results strongly support the view that incubation temperature influences the sensitivity cf a strain to a particular concentration of a pesticide.
References AGNIHOTRI, V. P.: ThiraDl induced changes in soil microflora their physiological activity and control of damping-off in chillies (Capsicum annuum). Indian J. Expt. Biology 12 (1974), 85-89. BALICKA, N., and SOBIESZCANSKI, J.: The effect of herbicides on soil microflora. III. The effect of herbicides on ammonification and nitrification in the soil. Acta Microbial. Pol. Ser. B 18 (1969), 7-10. BARTHA, R., LANZILOTTA, R. P., and PRAMER, D.: Stability and effects of some pesticides in soil. Appl. Microbial. 15 (19Ii7), 67-75. BOLLEN, E. B.: Interactions between pesticides and soil microorganisms. Annu. Rev. Microbial. 15 (1961), 69-72. CASSEMANS, J., and VAN ASSCHE, C.: Comparative study of growth-promoting capacity of SODle insecticidesfnematicides. Plant and Soil 63 (1981), 251-259. FUNKE, B. R., LIN, SHIN CHSIANG, SCHULZ, J. T.: Effect ofsoDle organophosphorus and carbamate insecticides on nitrification and legume growth. Bact. Proc. (Abstract) (1970), A62, p.9. HAMDI, YOUSEF A., and TEWFIK, M. S.: Effect of the herbicides trifluralin on nitrogen-fixation in Rhizobium and Azotobacter and on nitrification. Acta Microbiol. Pol. Ser. B 1 (1969), 53-58. MARTIN, J. P.: Influence of pesticides on soil microbes and soil properties. In: Pesticides and their effect on soil and water. Soil Sci. Soc. Amer. Inc., Madison, Wisconsin, 1966, 95-108. PAREEK, R. P.: Effect of DDT on symbiotic nitrogen fixation and nitrification in soil. M.Sc. Thesis, Department of Microbiology, LA.R.I., Delhi (1968). SUD, R. K., and GUPTA, K. G.: On the sensitivity of isolates of Rhizobium spp. and Azotobacter chroococcum to TMTD and its degradation product NaDDC. Arch. Mikrobiol. 85 (1972),19-22. Tu, C. M.: Effect of four organophosphorus insecticides on microbial activities in soil. App!. Micro· bioI. 19 (1970), 479-484. Effect of four nematocides on activities of microorganisms in soil. Appl. Microbial. 23 (1973), 398-401and BOLLEN, W. B.: Effect of paraquat on microbial activities in soils. Weed Res. 8 (1968), 28-37. ~. AINWRIGHT, M., and PUGH, G. J. F.: The effects of three fungicides on nitrification and ammonification in soil. S