Zbl. Bakt. Abt. II, Bd. 130, S.
41~44
(1975)
[University of Agricultural Sciences, Bangalore, India]
Effect of Seed Fumigation on the Symbiosis of Rhizobium sp. with Arachis hypogaea Linn. J. H. Kulkarni, J. S. Sardeshpande, and D. J. Bagyaraj Summary T o minimize the loss due to insect pests during storage, the fumigation of the seeds is advocated. However, no information is available on the subject how these fumigants, if such fumigated seeds are treated with Rhizobium, will interact with the Rhizobium legume symbiosis. Hence, groundnut seeds treated with ethylene dibromide, ethylene dichloride carbon tetrachloride mixture (EDCT), chloropicrin, and phosphine were treated with Rhizobium and sown. Observations were made on the number, fresh weight and leghaemoglobin content of the nodules, and on the yield, dry weight and nitrogen content of the plants. The results indicated that all the four fumigants adversely affected the nodulation status and the yield of groundnut treated with Rhizob'ium, and hence the use of fumigated seeds for Rhizobium treatment is not advisable. The insecticide malathion is also used in store houses to protect groundnut seeds from insect pests. When the malathion-treated seeds were inoc'ulated with Rhizobium and sown, no adverse effect on nodulation and yield was recorded.
Zusammenfassung Die Begasung von Saatgut wird haufig als Mittel gegen Verluste durch Insekten angewendet. Es sind keine Angaben vorhanden, auf welche Art und Weise diese Begasung die Symbiose von Rhizobium mit der Pflanze beeinflul.lt. Darum wurde Erdnul.l mit Athylenbl'omid, einem Athylendichlorid-Kohlenstofftetl'achlorid-Gemisch (EDCT), Chlorpikrin und Phosphin behandelt und angebaut. Die Zahl und die Masse del' frischen Knollchen, del' Gehalt an Leghamoglobin del' Knollchen sowie El'tl'ag und Stickstoffgehalt del' Pflanzen wurden el'mittelt. Aile angewandten Mittel beeinfluilten die Knollchenbildung und den El'tl'ag nachteilig. Es ist dahel' nicht cmpfehlenswert, sie anzuwenden. Dagegen hatte das Insektizid Melathion keine Nachwirkung, wenn es wahnmd del' Lagerung des Saatgutes angewandt wurde.
Arachis hypogaea Linn. (groundnut) is an important oil seed crop. Like other grains, ground nut seeds are subject to ravage of insects, rodents, and microorganisms during storage. To minimize the loss due to the insect pests and the microbial spoilage during storage the fumigation of seeds is advocated. MAJUMDAR et al. (1965) reported that fumigation of oil seeds with methyl bromide, ethylenc dibromide, chloropicrin, and ethylene dioxide have little effect on the spore-forming bacteria. Significant reduction in the fungal flora, but no marked effect on the bacterial flora of sorghum seeds, fumigated with ethylene dibromide, ethylene dichloride, and chloropicrin was reported by NARASIMHAN and RANGASWAMI (1968). Similar observations on groundnut, niger, and gingelli seeds were made by KULKARNI (1971), and on rice and finger millet by SRINIVASA et al. (1972). EBBELS (1967) studied the effect of four fumigants, viz., chloropicrin, dazomet, formaldehyde, and D. D. (1-3 dichloropropene dichloropropylene), applied to the soil on the nodulation of peas.
42
J. H. KULKARNI, J. S. SARDESHPANDE, and D. J. BAGYARAJ
Seed treatment of groundnut with Rhizobium for increasing the crop yields is becoming popular among the farmers in India. However, no information is available on the subject how these fumigants, if the fumigated seeds are treated with Rhizobium, will affect the Rhizobium-legume symbiosis. Hence, the present study was undertaken to investigate the effect of fumigation of groundnut seeds on the Rhizobium-legume symbiosis.
Materials and Methods 500 g. of TMV-2 groundnut seeds were takeI). into air-tight bottles of 600 m!. capacity. The fumigants, viz., ethylene dibromide, ethylene dibromide carbon tetrachloride mixture (EDCT), chloropicrin (each at the dosage of 3 mi. per quintal), and phosphine in the form of celphos (at the rate of 3 mg. per quintal) were introduced into the bottles. The lids were closed tightly, waxed, and stored for 7 days. Bottles containing seeds without fumigants served as control. Treatment of the groundnut seed!1 with the insecticide malathion (S-I, 2-di ethoxycarbonyl ethyl phosphoro dithioate) is also recommended for the control of the st orage pests. Hence, treatment of the groundnut seeds with malathion at the recommended rate (3 mg. per quintal) in bottles was also done for comparison. 7 days after fumigation, the groundnut seeds were treated with peat cultures, made from 5-day old Rhizobium sp. effective against groundnut, and sown in pots of the size of 45 x 30 em., filled with red sandy loam soil of pH 6.8. P in the form of superphosphate and K in the form of muriate of potash were applied to the pots at the rate of 60 kg. P and 15 kg. K per ha., respectively. Six replications were prepared for each treatment. Eight weeks after sowing, plants from 3 pots under each treatment were removed carefully and observations on the number of nodules and fresh weight of the nodules were recorded. The leghaemoglobin concentration of freshly excised nodules was determined by the method described by JORDAN and GARRARD (1951).
At harvest time, pod yield and dry matter weight of the plants were recorded. ThE' nitrogen content of the plants was also determined by the micro-Kjeldahl method.
Results and Discussion ~odulation status and leghaemoglobin content of the nodules of groundnut, as influenced by different fumigants, are given in Table I . The variations in the number and wet weight of the nodules, due to seed treatment with fumigants, were not statistically significant. However, treatment of seeds with fumigants was found to decrease considerably the number and wet weight of the root nodules. Maximum reduction in the number and fresh weight of the nodules was recorded due to fumigation with EDCT, followed by ethylene dibromide. The number and weight of the root nodules was found to have no correlation with the leghaemoglobin content of
Table 1. Effect of fumigation of seeds on number, weight, and leghaemoglobin content of the root nodules of groundnut, inoculated with Rhizobium Treatment
Nodule Number /plant
Fresh nodule weight mg./plant
Leghaemoglobin content mg. /g. nodule
Control Ethylene dibromide Phosphine Chloropicrin EDCT Malathion
53.80 26.40 33.20 32.00 19.60 39.00
140 80 87 103 45 99
1.80 2.32 1.80 2.66 2.32 2.66
Effect of Seed Fumigation on the Symbiosis of Rhizobium sp.
43
the nodules. Plants raised from seeds fumigated with chloropicrin, EDCT, and ethylene dibromide showed more leghaemoglobin content compared to the control. Fumigation with phosphine was found to have no effect on the leghaemoglobin content of the root nodules. Yield, dry matter weight, and nitrogen content of the plants at harvest time are presented in Table 2. All the fumigants tried were found to reduce significantly the yield and dry matter content of the groundnut plants. A slight reduction in the nitrogen content of groundnut plants, raised from the seeds treated with all the four fumigants used, was recorded. Table 2. Effect of fumigation of seeds on the yield, dry matter weight, and nitrogen content of groundnut plants, inoculated with Rhizobium Treatment
Yield of pods g./plant1 )
Dry matter weight g./plant 2 )
Nitrogen content %
Contre,l Ethylt.ne dibromide Phosphine Chloropicrin EDCT Malathion
2.95 1.83 1.71 2.33 2.71 3.12
7.76 5.65 5.82 6.4:3 5.85 7.85
2.71 2.65 2.23 2.32 2.14 2.82
1) Significant at level P = 0.05. 2) Significant at level P = 0.01.
These results clearly show that the fumigation of seeds with ethylene dibromide, EDCT, chloropicrin, and phosphine at the recommended rate adversely affect the numher and weight of nodules, yield, dry matter weight, and nitrogen content of plants which are taken as indices for the efficiency of nitrogen fixation by rhizobia. Hence, the use offumigated groundnut seeds for Rhizobium treatment is not advisable. Seed treatment with the insecticide malathion is also recommended to prevent damage due to insect pests during storage. It is interesting to observe that the plants raised from seeds treated with the insecticide malathion showed increased leghaemoglobin content of nodules, yield, dry weight, and nitrogen content compared to the control plants. There are a few reports that insecticides like DDT (Dichlorodiphenyl trichloroethane), BHC (Benzene hexachloride), lindane, dieldrin etc., applied to the soil, have no harmful effect on nodulation and yield of legumes treated with Rhizobium (PAREEK and GAUR 1970, SELIM et al. 1970). Some of the soil-applied insecticides were also reported to stimulate the crop growth of legumes treated with Rhizobium (SIMKOVER and SHENEFELT 1951, OSTWAL 1972). EBBELS (1967), studying the effect of 4 soil fumigants, viz., chloropicrin, dazomet, formaldehyde, and D-D on nodulation of peas, reported that all the four fumigants applied to the soil decreased the root nodulation. However, there is no information available on the effect of fumigants and insecticides used for sead treatment under storage on Rhizobium-legume symbiosis. It may be concluded that the use of groundnut seeds fumigated with ethylene dibromide, EDCT, chloropicrin, and phosphine for Rhizobium inoculation is not safe. Further, using the malathion-treated seeds for Rhizobium inoculation is not only safe,. but also beneficial as it slightly improved the crop growth of groundnut. Acknowledgement The authors are grateful to Dr. R. B. PATIL, Professor of Agricultural Microbiology, University of Agricultural Sciences, Bangalore. for his encouragement during the course of this study.
44
J. H. Kl:LKARNI, J. S. SARDESHPANDE, and D. J. BAGYARAJ, Effect of Seed Fumigation ..
References EBBELs, D. L.: Effect of soil fumigants on Fusarium wilt and nodulation of peas. Ann. appl. Biol. 60 (1967), 39l. JORDAX, D. C., and GARRARD, E. F.: Studies on the legume root nodule bacteria. r. Detection of effective and ineffective strains. Can. J. Bot. 29 (1951), 360. KULKARXI, J. H.: Studies on the microflora and biochemical changes in some stored oil seeds and pulses. :VI. Sc. (Agric.) Thesis, Univ, Agri. Sci., Bangalore (India) 1971. ::VIA,ruMDAR, S. K., NARASIMHAN, K. S., and PARPIA, H. A. B.: Micro.ecological factors of microbial spoilage and occurrence of mycotoxins in stored grains. Symp. Mycotoxins in Food Stuffs. Mass. lnst. Tech., Massachusetts, U.S.A. 1965. NAHASIMHAN, K. S., and RANGASWAlIi!, G.: Effeet of some fumigants on the microflora of sorghum seeds. Bull. Indian Phytopath. Soc. 4 (1968), 57. OS~'WAL, K. P.: Effect of nematocide Dasanite on nodulation and yield of Pha8eolu8 aureU8 L. In: Abstr. Symp. Legume Inoculants - Science and Technology, New Dehli 1972, p. 29. PAl~EEK, R. P., and GAUR, A. C.: Effect of Dichlor Diphenyl Trichlorethane (DDT) on symbiosis of Rhizobium sp. with Phaseolu8 (lureus (Green gram). Plant Soil 33 (1970), 297. SELIM, K. G., MAHMOUD, S. A. Z., and EL-MoKODAM, M. T.: Effect of Dieldrin and Lindane on the growth and nodulation of Vicia faba. Plant Soil 33 (1970), 325. SIll' KOVER, H. G., and SHENEFELT. R. D.: Effect of B.H.C. and chlordane on certain soil organisms. J. Econ. Entomol. 44 (1951), 426. SRlNIVASA, H. P., OBLISAlIII, G., and RANGASWAMI, G.: Microflora of finger millet and rice seeds. Mysore J. Agric. Sci. 6 (1972), 271. Authors' address:
J. H.KliLKARNI, J. S. SARDESHPANDE, and Dr. D. J. BAGYARAJ, University of Agricultural Science Hebbal, Bangalore-560024, South India.
41~44
(1975)
[University of Agricultural Sciences, Bangalore, India]
Effect of Seed Fumigation on the Symbiosis of Rhizobium sp. with Arachis hypogaea Linn. J. H. Kulkarni, J. S. Sardeshpande, and D. J. Bagyaraj Summary T o minimize the loss due to insect pests during storage, the fumigation of the seeds is advocated. However, no information is available on the subject how these fumigants, if such fumigated seeds are treated with Rhizobium, will interact with the Rhizobium legume symbiosis. Hence, groundnut seeds treated with ethylene dibromide, ethylene dichloride carbon tetrachloride mixture (EDCT), chloropicrin, and phosphine were treated with Rhizobium and sown. Observations were made on the number, fresh weight and leghaemoglobin content of the nodules, and on the yield, dry weight and nitrogen content of the plants. The results indicated that all the four fumigants adversely affected the nodulation status and the yield of groundnut treated with Rhizob'ium, and hence the use of fumigated seeds for Rhizobium treatment is not advisable. The insecticide malathion is also used in store houses to protect groundnut seeds from insect pests. When the malathion-treated seeds were inoc'ulated with Rhizobium and sown, no adverse effect on nodulation and yield was recorded.
Zusammenfassung Die Begasung von Saatgut wird haufig als Mittel gegen Verluste durch Insekten angewendet. Es sind keine Angaben vorhanden, auf welche Art und Weise diese Begasung die Symbiose von Rhizobium mit der Pflanze beeinflul.lt. Darum wurde Erdnul.l mit Athylenbl'omid, einem Athylendichlorid-Kohlenstofftetl'achlorid-Gemisch (EDCT), Chlorpikrin und Phosphin behandelt und angebaut. Die Zahl und die Masse del' frischen Knollchen, del' Gehalt an Leghamoglobin del' Knollchen sowie El'tl'ag und Stickstoffgehalt del' Pflanzen wurden el'mittelt. Aile angewandten Mittel beeinfluilten die Knollchenbildung und den El'tl'ag nachteilig. Es ist dahel' nicht cmpfehlenswert, sie anzuwenden. Dagegen hatte das Insektizid Melathion keine Nachwirkung, wenn es wahnmd del' Lagerung des Saatgutes angewandt wurde.
Arachis hypogaea Linn. (groundnut) is an important oil seed crop. Like other grains, ground nut seeds are subject to ravage of insects, rodents, and microorganisms during storage. To minimize the loss due to the insect pests and the microbial spoilage during storage the fumigation of seeds is advocated. MAJUMDAR et al. (1965) reported that fumigation of oil seeds with methyl bromide, ethylenc dibromide, chloropicrin, and ethylene dioxide have little effect on the spore-forming bacteria. Significant reduction in the fungal flora, but no marked effect on the bacterial flora of sorghum seeds, fumigated with ethylene dibromide, ethylene dichloride, and chloropicrin was reported by NARASIMHAN and RANGASWAMI (1968). Similar observations on groundnut, niger, and gingelli seeds were made by KULKARNI (1971), and on rice and finger millet by SRINIVASA et al. (1972). EBBELS (1967) studied the effect of four fumigants, viz., chloropicrin, dazomet, formaldehyde, and D. D. (1-3 dichloropropene dichloropropylene), applied to the soil on the nodulation of peas.
42
J. H. KULKARNI, J. S. SARDESHPANDE, and D. J. BAGYARAJ
Seed treatment of groundnut with Rhizobium for increasing the crop yields is becoming popular among the farmers in India. However, no information is available on the subject how these fumigants, if the fumigated seeds are treated with Rhizobium, will affect the Rhizobium-legume symbiosis. Hence, the present study was undertaken to investigate the effect of fumigation of groundnut seeds on the Rhizobium-legume symbiosis.
Materials and Methods 500 g. of TMV-2 groundnut seeds were takeI). into air-tight bottles of 600 m!. capacity. The fumigants, viz., ethylene dibromide, ethylene dibromide carbon tetrachloride mixture (EDCT), chloropicrin (each at the dosage of 3 mi. per quintal), and phosphine in the form of celphos (at the rate of 3 mg. per quintal) were introduced into the bottles. The lids were closed tightly, waxed, and stored for 7 days. Bottles containing seeds without fumigants served as control. Treatment of the groundnut seed!1 with the insecticide malathion (S-I, 2-di ethoxycarbonyl ethyl phosphoro dithioate) is also recommended for the control of the st orage pests. Hence, treatment of the groundnut seeds with malathion at the recommended rate (3 mg. per quintal) in bottles was also done for comparison. 7 days after fumigation, the groundnut seeds were treated with peat cultures, made from 5-day old Rhizobium sp. effective against groundnut, and sown in pots of the size of 45 x 30 em., filled with red sandy loam soil of pH 6.8. P in the form of superphosphate and K in the form of muriate of potash were applied to the pots at the rate of 60 kg. P and 15 kg. K per ha., respectively. Six replications were prepared for each treatment. Eight weeks after sowing, plants from 3 pots under each treatment were removed carefully and observations on the number of nodules and fresh weight of the nodules were recorded. The leghaemoglobin concentration of freshly excised nodules was determined by the method described by JORDAN and GARRARD (1951).
At harvest time, pod yield and dry matter weight of the plants were recorded. ThE' nitrogen content of the plants was also determined by the micro-Kjeldahl method.
Results and Discussion ~odulation status and leghaemoglobin content of the nodules of groundnut, as influenced by different fumigants, are given in Table I . The variations in the number and wet weight of the nodules, due to seed treatment with fumigants, were not statistically significant. However, treatment of seeds with fumigants was found to decrease considerably the number and wet weight of the root nodules. Maximum reduction in the number and fresh weight of the nodules was recorded due to fumigation with EDCT, followed by ethylene dibromide. The number and weight of the root nodules was found to have no correlation with the leghaemoglobin content of
Table 1. Effect of fumigation of seeds on number, weight, and leghaemoglobin content of the root nodules of groundnut, inoculated with Rhizobium Treatment
Nodule Number /plant
Fresh nodule weight mg./plant
Leghaemoglobin content mg. /g. nodule
Control Ethylene dibromide Phosphine Chloropicrin EDCT Malathion
53.80 26.40 33.20 32.00 19.60 39.00
140 80 87 103 45 99
1.80 2.32 1.80 2.66 2.32 2.66
Effect of Seed Fumigation on the Symbiosis of Rhizobium sp.
43
the nodules. Plants raised from seeds fumigated with chloropicrin, EDCT, and ethylene dibromide showed more leghaemoglobin content compared to the control. Fumigation with phosphine was found to have no effect on the leghaemoglobin content of the root nodules. Yield, dry matter weight, and nitrogen content of the plants at harvest time are presented in Table 2. All the fumigants tried were found to reduce significantly the yield and dry matter content of the groundnut plants. A slight reduction in the nitrogen content of groundnut plants, raised from the seeds treated with all the four fumigants used, was recorded. Table 2. Effect of fumigation of seeds on the yield, dry matter weight, and nitrogen content of groundnut plants, inoculated with Rhizobium Treatment
Yield of pods g./plant1 )
Dry matter weight g./plant 2 )
Nitrogen content %
Contre,l Ethylt.ne dibromide Phosphine Chloropicrin EDCT Malathion
2.95 1.83 1.71 2.33 2.71 3.12
7.76 5.65 5.82 6.4:3 5.85 7.85
2.71 2.65 2.23 2.32 2.14 2.82
1) Significant at level P = 0.05. 2) Significant at level P = 0.01.
These results clearly show that the fumigation of seeds with ethylene dibromide, EDCT, chloropicrin, and phosphine at the recommended rate adversely affect the numher and weight of nodules, yield, dry matter weight, and nitrogen content of plants which are taken as indices for the efficiency of nitrogen fixation by rhizobia. Hence, the use offumigated groundnut seeds for Rhizobium treatment is not advisable. Seed treatment with the insecticide malathion is also recommended to prevent damage due to insect pests during storage. It is interesting to observe that the plants raised from seeds treated with the insecticide malathion showed increased leghaemoglobin content of nodules, yield, dry weight, and nitrogen content compared to the control plants. There are a few reports that insecticides like DDT (Dichlorodiphenyl trichloroethane), BHC (Benzene hexachloride), lindane, dieldrin etc., applied to the soil, have no harmful effect on nodulation and yield of legumes treated with Rhizobium (PAREEK and GAUR 1970, SELIM et al. 1970). Some of the soil-applied insecticides were also reported to stimulate the crop growth of legumes treated with Rhizobium (SIMKOVER and SHENEFELT 1951, OSTWAL 1972). EBBELS (1967), studying the effect of 4 soil fumigants, viz., chloropicrin, dazomet, formaldehyde, and D-D on nodulation of peas, reported that all the four fumigants applied to the soil decreased the root nodulation. However, there is no information available on the effect of fumigants and insecticides used for sead treatment under storage on Rhizobium-legume symbiosis. It may be concluded that the use of groundnut seeds fumigated with ethylene dibromide, EDCT, chloropicrin, and phosphine for Rhizobium inoculation is not safe. Further, using the malathion-treated seeds for Rhizobium inoculation is not only safe,. but also beneficial as it slightly improved the crop growth of groundnut. Acknowledgement The authors are grateful to Dr. R. B. PATIL, Professor of Agricultural Microbiology, University of Agricultural Sciences, Bangalore. for his encouragement during the course of this study.
44
J. H. Kl:LKARNI, J. S. SARDESHPANDE, and D. J. BAGYARAJ, Effect of Seed Fumigation ..
References EBBELs, D. L.: Effect of soil fumigants on Fusarium wilt and nodulation of peas. Ann. appl. Biol. 60 (1967), 39l. JORDAX, D. C., and GARRARD, E. F.: Studies on the legume root nodule bacteria. r. Detection of effective and ineffective strains. Can. J. Bot. 29 (1951), 360. KULKARXI, J. H.: Studies on the microflora and biochemical changes in some stored oil seeds and pulses. :VI. Sc. (Agric.) Thesis, Univ, Agri. Sci., Bangalore (India) 1971. ::VIA,ruMDAR, S. K., NARASIMHAN, K. S., and PARPIA, H. A. B.: Micro.ecological factors of microbial spoilage and occurrence of mycotoxins in stored grains. Symp. Mycotoxins in Food Stuffs. Mass. lnst. Tech., Massachusetts, U.S.A. 1965. NAHASIMHAN, K. S., and RANGASWAlIi!, G.: Effeet of some fumigants on the microflora of sorghum seeds. Bull. Indian Phytopath. Soc. 4 (1968), 57. OS~'WAL, K. P.: Effect of nematocide Dasanite on nodulation and yield of Pha8eolu8 aureU8 L. In: Abstr. Symp. Legume Inoculants - Science and Technology, New Dehli 1972, p. 29. PAl~EEK, R. P., and GAUR, A. C.: Effect of Dichlor Diphenyl Trichlorethane (DDT) on symbiosis of Rhizobium sp. with Phaseolu8 (lureus (Green gram). Plant Soil 33 (1970), 297. SELIM, K. G., MAHMOUD, S. A. Z., and EL-MoKODAM, M. T.: Effect of Dieldrin and Lindane on the growth and nodulation of Vicia faba. Plant Soil 33 (1970), 325. SIll' KOVER, H. G., and SHENEFELT. R. D.: Effect of B.H.C. and chlordane on certain soil organisms. J. Econ. Entomol. 44 (1951), 426. SRlNIVASA, H. P., OBLISAlIII, G., and RANGASWAMI, G.: Microflora of finger millet and rice seeds. Mysore J. Agric. Sci. 6 (1972), 271. Authors' address:
J. H.KliLKARNI, J. S. SARDESHPANDE, and Dr. D. J. BAGYARAJ, University of Agricultural Science Hebbal, Bangalore-560024, South India.
