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Chaudhary Charan Singh Haryana Agricultural University, Hisar
Chaudhary Charan Singh Haryana Agricultural University popularly known as HAU, is one of Asia's biggest agricultural universities, located at Hisar in the Indian state of Haryana. It is named after India's seventh Prime Minister, Chaudhary Charan Singh. It is a leader in agricultural research in India and contributed significantly to Green Revolution and White Revolution in India in the 1960s and 70s. It has a very large campus and has several research centres throughout the state. It won the Indian Council of Agricultural Research's Award for the Best Institute in 1997. HAU was initially a campus of Punjab Agricultural University, Ludhiana. After the formation of Haryana in 1966, it became an autonomous institution on February 2, 1970 through a Presidential Ordinance, later ratified as Haryana and Punjab Agricultural Universities Act, 1970, passed by the Lok Sabha on March 29, 1970. A. L. Fletcher, the first Vice-Chancellor of the university, was instrumental in its initial growth.
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ThesisItem Open Access Development of biocontrol agent against phytopathogenic fungi of tomato (Lycopersicon esculentum Mill.) crop.(CCSHAU, Hisar, 2020-07) Kapoor, Divya; Rakesh KumarIn the present study, seventy-eight bacterial isolates were retrieved from the tomato rhizosphere soil and screened for their antagonistic activity against Fusarium oxysporum f. sp. lycopersici, Alternaria solani and Rhizoctonia solani under in vitro conditions. Thirteen isolates, NAU5, NAU10, NAU13, NAU20, NAU21, NAU23, NAU29, NAU32, NAU36, NAA59, NAA60, KBA43 and KBA44 showed broad inhibition zones against A. solani measuring more than 20 mm. Six isolates, NAU19, NAU20, NAU23, NAU24, NAU27 and NAU30 resulted in broad inhibition zones against F. oxysporum f. sp. lycopersici measuring 10.1-20 mm. Six isolates, NAU20, NAU30, NAU35, LBV58, NAV66 and NAV73 resulted in broad inhibition zones against R. solani measuring 15.1-25 mm. Among the retrieved 78 isolates, 28% of rhizobacteria produced high amount of IAA and seven isolates, NAU4, NAU5, NAU10, NAU20, NAU28, KBA43 andKBV47 produced maximum IAA. The amount of IAA production by different isolates varied from 3.62μg/ml (NAV69) to 26.96μg/ml (NAU5). Among 78 isolates tested for chitinase assay, two isolates, NAU13 and NAV68 showed maximum activity. Out of 78 isolates, siderophore production was observed in 42% of retrieved bacteria and five isolates, NAU19, KBV45, KBV46, LBV56 and NAV75 produced maximum orange halo zone. HCN was produced by 10% of isolates and highest HCN was observed by 6 isolates, NAU4, NAU6, NAU20, NAU29, NAU41 and NAA59, qualitatively. Seven isolates, NAU5, NAU20, NAU30, NAU35, NAA59, NAV68 and NAV75 were selected and total of 27 treatments were prepared on the basis of antagonistic and plant growth promoting attributes tested in vitro for further evaluating under pot house conditions. Isolate NAU5 showed maximum shoot length, shoot weight, root weight, fruit yield and disease control (100%) against F. oxysporum f. sp. lycopesici. Isolate NAV75 showed maximum shoot length, shoot weight, root weight, fruit yield and disease control (87%) against A. solani. Isolate NAU35 showed maximum shoot length, shoot weight, root weight, fruit yield and disease control (100%) against R. solani. Irrespective of the fungi, isolate NAU5 showed biocontrol activity against all the three phytopathogens. The seven promising isolates showed the PGPR activities in the order NAV75> NAV68> NAA59> NAU5> NAU35> NAU20> NAU30 irrespective of the fungi in pot house studies. The isolates NAU5 against all the three fungi, NAU35 against R. solani and NAV75 against A. solani can be further tested on the tomato fields as potent biocontrol agents. On the basis of biochemical characterization isolate NAU5 was found to belong to Alcaligenes, NAU35 to Bacillus and NAV75 to Burkholderia genus.ThesisItem Open Access Impact of co-inoculation of plant growth promoting actinomycetes and biofertilizers on mustard (Brassica juncea L.) crop(CCSHAU, Hisar, 2020) Umang; Leela WatiMustard (Brassica juncea L.) a major oilseed crop and the second largest agriculture commodity after cereals in India, plays a significant role in Indian economy. Due to inconsistent climatic changes, occurrence of diseases and implementation of unsuitable dosage of chemical fertilizers and pesticides, overall production of mustard has been comparatively static for last two decades regardless of the use of numerous traditional and breeding approaches. Actinomycetes, a class of actinobacteria are Gram-positive bacteria found in soil and possessing various plant growth traits can play significant role in improving growth and yield of mustard crop. In the present study, two biofertilizer cultures viz., Azotobacter chroococcum and phosphate solubilizing bacterium (PSB) along with seven available actinomycetes isolates AK1, AK3, AK6, AK11, AK17, AK20 and AK37 were screened for different plant growth promoting traits including IAA production, phosphate solubilization, potassium solubilization, zinc solubilization, siderophore production, HCN production, cellulose and chitin degradation under in vitro conditions. The actinomycetes isolates exhibited three or more plant growth promoting traits and were compatible with standard biofertilizer cultures. Actinomycete isolates exhibiting multiple plant growth promoting traits along with biofertilizer cultures were evaluated for growth promotion of mustard under pot culture conditions during 2019-2020. Actinomycete isolate AK17 was adjudged best in improving growth andseed yield of mustard crop when coinoculated with Azotobacter chroococcum and PSB. The isolates AK20 & AK37 were found promising in improving vegetative growth only. The actinomycete isolate AK17 identified by 16S rDNA sequencing showing 99% similiarity with Streptomyces pseudgriseolus, could further be explored as bioinoculant in agricultural practices.ThesisItem Open Access Impact of chitinase producing bacteria on thrips infestation in citrus plantation(CCSHAU,HiSAR, 2020) Sharma, Nidhi; Pathak, D.V.The studies on chitinase activity of bacterial isolates against thrips on Kinnow were carried out at citrus orchard with the objective Impact of chitinase producing bacteria on thrips infestation in citrus plantation. Plant growth promoting rhizobacteria (PGPR) are root associated bacteria and benefit plants by providing growth promotion through a variety of mechanism. The public concern over the harmful effects of chemical pesticides on the environment and human health has enhanced the search for safer, environmentally friendly control alternatives. Due to the importance of chitinolytic enzymes in insect, nematode, and fungal growth and development, they are receiving attention in regard to their use as biopesticides and microbial biocontrol agents. Therefore, the present study was aimed to isolate and characterize the chitinolytic bacterial isolates and also evaluate their efficacy. A total of eighty morphologically distinct bacterial isolates were retrieved from soil samples of different microbiomes on King‘s B media and these bacterial isolates were screened for chitinase production out of these twenty isolates was showed zone of clearance on the collodial chitin agar plates where zone of clearance and enzyme activity index were used for initial screening of chitinolytic bacteria. Twenty out of eighty bacterial isolates were selected as chitinase producers on the basis of zone size ranging from 12.40 mm to 38.50 mm and were further screened for quantitative chitinase production. Quantitatively it was screened for chitinase enzyme activity on the basis of release of N-acetylglucosamine units (U) by action of chitinase, maximum enzyme activity was recorded for isolate B1 (3.905) followed by B5 (3.52) and D2 (1.35). A total of twenty bacterial isolates, were studied on the basis of their morphological and microscopic characterstics which showed a variation in margins, elevation, shape, pigmentation, Gram‘s reaction and cell shape. These bacterial isolates was biochemically characterized, isolate B5 was found positive for various biochemical tests such as citrate utilization test, Voges Proskauer (VP) test and negative for methyl red test. All the twenty isolates was screened for multifarious plant growth promoting traits viz. IAA production and HCN production. Three bacterial isolates which exhibited maximum chitinase activity were further evaluated for their biocontrol potential against thrips of citrus under field conditions. Kinnow were sprayed with three different bacterial isolates B1, B5, D2 (106 cfu/ml), dimethoate phosphamide spray and nimbicide at different concentrations (1.5ml/L, 2ml/L respectively). One isolate B1 showed maximum biocontrol efficacy in reduction of thrips population -79.66 percent against thrips in kinnow in comparison to control where -37.70 percent of reduction of thrips population was recorded. The percent infestation recorded for B1 isolate was 20 percent as compared to control where infestation was 36 percent. The mean population density recorded for B1 was 0.4 as compared to control where density of thrips was 0.96. The identification of B1 was tentatively confirmed by biochemical characterization and it was identified as strain belonging to Bacillus sp.ThesisItem Open Access Studies on molecular mechanisms underlying rhizobia promiscuity in different Khariflegumes(CCSHAU, Hisar, 2020) Rana, Barkha; Gera, RajeshMungbean, mothbean, cowpea, clusterbean, urdbean and pigeonpea are economically important kharif legumes commonly grown in rain-fed areas particularly South-West Haryana and Rajasthan state. It is possible to improve its symbiotic property for different kharif legumes through identification and manipulation of genetic factors that controls the promiscuity of Rhizobium. So the present investigation was undertaken to study molecular mechanism underlying rhizobia promiscuity in different kharif legumes. A total of 10 rhizobial isolates along with three rhizobial mutants were obtained from Molecular Microbiology laboratory of Microbiology Department that were retrieved from arid and hyper-arid zones of South-West Haryana and Rajasthan state. All selected rhizobial isolates were checked for their authenticity in six different kharif legumes after 45 days of growth under sterilized conditions. Promiscuous rhizobial mutants (PKR177m30, PKR188m85 and PKR194m93) have the ability to nodulate all six kharif legumes with high nodulation efficiency. Evaluation of these isolates for cellulase activity both on CMC agar plates and in liquid broth revealed that promiscuous rhizobial mutants showed highest cellulase activity both on CMC agar plates and in liquid broth as compared to other rhizobia. Molecular mechanism of rhizobial promiscuity was studied by amplifying and detecting relevant nodulation genes such as nodD, nodT, nodXandnolA in different rhizobia. PCR conditions were optimized to amplify a particular gene. All the 13 selected rhizobial isolates showed the presence of nodD and nodX genes, whereas amplification of nodT was observed in the isolates capable of nodulating 6, 5 and 4 different legumes. However, the expression of nolA gene was fairly good with single amplified PCR product in the rhizobia capable of nodulating 6, 5 and 4 different kharif legumes, amplification of nolA with single PCR amplified product of 672 bp was observed in isolates nodulating 6, 5 and 4 different while rest of the isolates showed poor expression of nolA gene with multiple bands 672 & 323bp. The mutant PKR188m85 performed better in terms of nodulation efficiency and plant growth parameters in pigeonpea, mungbean, clusterbean and mothbean whereas PKR177m30 performed better in cowpea and urdbean followed by PKR188m85 than their respective parent and reference strains under sterilized conditions.ThesisItem Open Access Biochemical characterization and toxicity Studies of biosurfactants produced by yeast(CCSHAU, 2019) Dolly Rani; Sangwan, SeemaThe biosurfactant production potential of two yeasts, Meyerozyma guilliermondii YK 32 and M. guilliermondii YK 22 was investigated using total eight types of carbon sources followed by extraction and characterization of final product. Maximum biosurfactant production monitored in terms of oil displacement (9.0cm) was obtained using a mixture of olive oil and whey at 4%(v/v) concentration of each in case of M. guilliermondii YK 32. Its cell free supernatant yielded maximum 12.77g/L crude biosurfactant using acetone precipitation method. Total eight types of crude biosurfactants obtained using different carbon sources (BS1-BS8) were subjected to further characterization. The preliminary characterization using TLC revealed yellow spots in case of BS1, BS4, BS5 and BS8 having Rf 0.33, 0.41, 0.45 and 0.38, respectively indicating the presence of carbohydrate and lipids while a ninhydrin spray detected purple to reddish purple spots in BS2, BS3, BS6 and BS7 with Rf value varied from 0.23 to 0.93 depicting protein and lipid contents. The biosurfactant produced using a mixture of olive oil and whey (BS6) as substrate contained highest protein content (28.61 mg/g) followed by BS2 (20.90 mg/g) and BS1 (15.94 mg/g) while a mixture of molasses and whey supported the production of crude biosurfactant (BS8) containing highest carbohydrate (28.82 mg/g) as well as lipid content (A260nm, 16.85). All the crude biosurfactants exhibited neutral behavior during ionic charge determination. Contrary to chemical surfactant (1%SDS) which inhibited the germination of chickpea, the increasing concentration of biosurfactants supported the germination of chickpea giving a maximum germination index equaled to 423 % at 0.8% concentration of biosurfactants BS4.Further, all the biosurfactants were non toxic to yeast multiplication and did not show any antimicrobial activity against Rhizobium, a bioinoculant for nitrogen fixation. Being supportive to chickpea germination and nontoxic to yeast, the neutral biosurfactant produced by M. guilliermondii YK 32 may be tested for application in agricultural practices, cosmetics and pharmaceutical industries.ThesisItem Open Access Bioremediation of phenolic compounds in sewage water using bacterial consortia(CCSHAU, 2019) Diksha; Rakesh KumarSewage water is a rich source of organic waste and nutrients in the form of nitrates and phosphates, due to which it can be used in agriculture after proper treatment. Sewage water contains various harmful compounds like phenolics which need to be treated before discharge. The direct discharge of sewage water into natural water sources leads to the obstructive impacts on the water sources and aquatic life, sewage water becomes an open source of harmful microbes and pollutants. To demolish these problems the treatment of domestic waste water is must and its reuse for the domestic as well as agriculture purpose can become an eco-friendly solution for the conservation of water. A total of 29 bacterial isolates were retrieved from seven sewage water samples collected from Sewage Treatment Plant, CCS HAU, Hisar. All bacterial isolates were screened for phenol degradation in mineral salt medium supplemented with 250, 500 and 750 ppm concentration of phenol. Out of 29, only 8 bacterial isolates were able to grow in mineral salt medium having 750 ppm concentration of phenol. These eight bacterial isolates were further observed for growth upto 1500 ppm concentration of phenol, out of eight, only four bacterial isolates (NI6, NI5, NR1 and JC1) were able to grow upto 1250 ppm concentration of phenol. Cultural conditions, inoculum size and aeration were optimised in culture medium as well as sewage sludge. Fifteen percent inoculum and 2 LPM aeration rate were found to be optimum for efficient phenol degradation. Least residual phenol in culture medium and sewage sludge at 15% inoculum was left with bacterial isolate NI6 (265.2 ppm and 607.30 ppm respectively). Same was observed in culture medium and sewage sludge at 15% inoculum and 2 LPM aeration in lab scale fermenter for bacterial isolate NI6 (352.3 ppm and 761.27 ppm respectively). A consortium of four bacterial isolates (25% each of NI6, NI5, NR1 and JC1) was found to leave least 276.93 ppm residual phenol in sewage sludge as compared to individual isolates. Consortium was further tested for removal of phenolic compounds, COD, BOD and Heavy metal at STP, CCS HAU, Hisar. At 9th day of application, highest phenolic compounds degradation was observed in sewage sludge (120 ppm) and outlet of clarifier (172.9 ppm), thereafter phenolic compounds concentration started to increase. At 9th day COD reduced from 460 mg/L to 60 mg/L and 100 mg/L to 40 mg/L while BOD decreased from 153 mg/L to 18 mg/L and 28 mg/L to 17 mg/L in sewage sludge and outlet of clarifier respectively. Removal of Zinc and Chromium was observed in outlet of clarifier and sewage sludge.ThesisItem Open Access Characterization of thermotolerant rhizobacteria to control root rot disease in clusterbean (Cyamopsis tetragonoloba L.)(CCSHAU, 2019) Sachin; Pathak, D.V.Cluster bean [Cyamopsis tetragonoloba (L.) Taub.] is an under exploited leguminous crop which is commonly known as guar, chavli kayi, guari, khutti etc. It is grown in India, especially in South Haryana, Rajasthan, Andhra Pradesh and Maharastra in the summer season. India contributes around 80% of cluster bean to the world. Rhizoctonia solani Kuhn, a basidomycetes fungus, is the main causal organism of this disease. The control of fungus is difficult because of wide range of its host, large sclerotia are insensitive to chemical fungicides. A total of fifty eight bacterial isolates were retrieved from rhizosphere of mainly two crops (Wheat and congress grass). All the bacterial isolates were screened for Gram’s staining, spore staining, IAA production, NH3 excretion, growth on chitin as carbon source, antifungal activity against R. solani, Germination index, pre emergence mortality and Relative root elongation index (RREI). On the basis of Gram’s staining 45 isolates were found to be Gram’s +ve, remaining were Gram nagetive. Endospore staining showed 45 isolates as spore formers. A total of 56 isolates were capable of IAA production and maximum IAA producer was NAA4 (25.03 μg ml-1). Out of 58 isolates 53 isolates were capable of ammonia excretion and maximum NH3+ excretion was 4.16 μg ml- 1 by isolate no. KMI1. Only five isolates (KMI1, NAB1, NAE2, NAD1 and NAC2) were showed chitinase activity using chitin as carbon source. Ten bacterial isolates were showed antifungal activity against R. solani under in vitro conditions. Impact of 20 best cultures selected on the basis of biochemical characterization, was studied for germination in cluster bean under in vitro conditions. The maximum root length was observed with isolate NAG6 (7.86 cm) and maximum shoot length was 8.82 cm with NAG6. Maximum RREI % was observed with NAG6 which was 242.59 %. Minimum pre emergence mortality was observed with KME2. It was only 3 %. Germination index varied from 50 to 90 % and maximum GI was 90% in five cultures KMB7, KMH3, KML2, NAB1, NAD1. The best five cultures on the basis of antifungal activity, chitinase activity and other biochemical characterization were selected to evaluate under pot house conditions. Maximum root length was observed with NAE2 along with RDF (18.11cm) and shoot length was observed with NAE2 along with RDF (20.76 cm). Maximum GI was observed in case of NAE2 along with RDF (82.96 %). Minimum pre emergence mortality % observed was 8.00 % in NAE2 along with RDF. Minimum post emergence mortality % observed was 9.04 % in NAE2 along with RDF. The maximum dry shoot weight was observed in case of NAE2 along with RDF (5.075 mg). The maximum dry root weight was 0.403 mg in NAE2 along with RDF followed by 0.392 mg in NAD1 along with RDF. The maximum N uptake was observed with NAE2 along with RDF (64.75 mg/pl.). NAE2 culture performed better for root rot control and most of plant growth parameter under pot house conditions. This culture was identified as Bacillus subtilis subtilis using 16sRNA RFLP.ThesisItem Open Access Green synthesis and characterization of plant derived nanoparticles for enhancement of biogas production(CCSHAU, 2019) Dilbag; Malik, KamlaEnergy crisis and environmental pollution are serious problems of sustainable development of the world. The problem is far more acute in the perspective of developing economies in countries like India where there is significant pressure on the available natural sources of energy. One of effective approaches to solve these energy problems is developing biomass energy such as biogas and biofuel. Biogas production is a well-established conversion technology to obtain energy from biomass that is alternative to fossil fuel. Nanotechnology can pave the way for betterment of biogas production through the use of nanocatalysts which ensure more efficient bioconversion strategies, better breakdown of substrates and more optimized output delivery. Generally, nanoparticles are prepared by a variety of chemical and physical methods which are quite expensive and potentially hazardous to the environment which involve use of toxic and perilous chemicals that are responsible for various biological risks. This may be the reason which leads to the biosynthesis of nanoparticles via green route that does not employ toxic chemicals and hence proving to become a growing interest to develop environment friendly processes. Therefore, in the present investigation was carried out green synthesized plant derived iron nanoparticles used for enhancement of biogas production from cattle dung. The nanoparticles were synthesized from neem (Azadirachta indica) and curry (Murraya koenigii) leaves with various conc. (30-70 ml) supplemented with cattle dung for biogas production. Characterization of the synthesized iron nanoparticles was done by UV-visible spectrophotometer, PSA, zeta potential analyzer (ZPA) and transmission electron microscopy (TEM). The highest absorption peak was observed at 432 nm.TEM image reveals that the FeNPs were in smaller size and spherical in shape with a diameter of 72.5 nm (NE) and 25 nm (CE), respectively. The maximum degradation (43.1%) of total solids was observed in T-5. Similar trend was observed in VS degradation and maximum degradation (15.7%) was observed in T-5. Biogas production from FeNPs (NE & CE) T-5 after 8 weeks of digestion was 104.3 litres and in T-9, it is found to be 98.6 litres as compared to control in which biogas production was found to be 85.2 litres. There was 22.4% increase in biogas production as compared to control after 8 weeks of batch anaerobic digestion. Cumulative biogas production with the addition of FeNPs attained the highest biogas yield (2537 l) in J2 as compared to J1 (2201 l) and methane contents were 77.8% as compared to control 72.3% after 8 week of digestion.ThesisItem Open Access Biocontrol of root-knot nematode (Meloidogyne javanica) in brinjal (Solanum melongena L.) using plant growth promoting rhizospheric bacteria(CCSHAU, 2018) Antil, Sonam; Rakesh KumarEgg plant (Solanum melongena L.) or brinjal is a nightshade species belonging to family Solanaceae and is the second most important vegetable crop after tomato and shows high susceptibility to infection by the root knot nematode Meloidogyne javanica. Management of the root-knot nematodes via biological methods such as PGPR is a novel technology emerging for sustainable agriculture. In this context, a total of 63 isolates were isolated from nematode affected rhizospheric soil of brinjal, tomato crops and earthworm culture media. All the 63 isolates were characterized for various plant growth promoting traits. Out of 63, fifteen isolates produced siderophores and maximum production was observed by isolate LBB-1and KMT-2. Chitinase activity was observed in 14 isolates and isolates KMT-2 and KMT-8 showed maximum chitinase activity. Sixteen isolates were found to be positive for HCN production and maximum production was seen in isolates NAE-1, KMB-3, KMT-2 and KMS-6. Fifty seven isolates were capable of excreting ammonia and maximum ammonia excretion was observed in KMT-2 (4.8μg/ml) and LBB-1 (4.7 μg/ml). All the bacterial isolates except one were producing IAA and high level of IAA production was observed in KMT-1 (45.9 μg/ml), KMB-2 (38.8 μg/ml), KMB-4 (27.7μg/ml) and KMT-5 (26.4μg/ml). A total of 39 isolates solubilized phosphate and highest P-SI was observed in isolate KMT-1(2.0) and LBB-4 (1.7). Twelve isolates- NAB-4, NAS-1, NAE-1, LBB-1, LBS-1, KMB-3, KMT-2, KMT-4, KMT-5, KMT-8, KMS-3 and KMS-6 resulted in minimum hatching in egg masses and higher mortality rate in J2 of M. javanica (upto 92%). Based on all the screening tests, a total of 12 were selected for pot house experiment on brinjal crop. Rhizospheric microbial count increased in all treatments from 0 DAP to 45 DAP. Under pot house conditions, T14 (RDF +KMT-4), T15 (RDF +KMT-5), T16 (RDF + KMT-8) and T17 (RDF +KMS-6) showed maximum plant growth, reduced galls and eggs in roots, and final nematode population in soil.
