Browsing by Author "Mahatma, Lalit"
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ThesisItem Open Access Biochemical and Molecular Characterization of Cellulase Producing Microorganisms from Termite(Plant Molecular Biology and Biotechnology Dept., ACHF, NAU, Navasari, 2016-07) Gadhiya, Komal J.; Mahatma, LalitThree bacterial isolates viz., Pseudomonas viridivida NAULK-1, Pseudomonas fluorescens NAULK-2 & Serratia plymuthica NAULK-3 were isolated from the termite (Heterotermes tenuis) gut. These were Gram negative, short rod shaped occurring singly and/or cluster. P. fluorescens NAULK-2 was short rod forming capsule. No capsule was seen in other two isolates. All three isolates were motile. All three bacteria can grow at 5-8 pH, however, showed distinct cultural characteristics on Luria Burtani (LB) agar plate. Three cellulose degrading enzyme viz., endoglucanase, exoglucanase and β-glucosidase were produced by all the three microorganism. Thereby they were capable to hydrolyze long chains of the cellulose in a progressive process because of exoglucanases, cleaves intramolecular β-1,4-glucosidic linkages and decreases the specific viscosity because of endoglucanases enzyme. Wood principally contains Cellulose, Hemicellulose and Lignin. Lignins are particularly important in the formation of cell walls, especially in wood and bark. Lignin fills the spaces in the cell wall between cellulose, hemicellulose, and pectin components, especially in xylem tracheids, vessel elements and sclereid cells. It is covalently linked to hemicellulose and therefore crosslinks different plant polysaccharides, conferring mechanical strength to the cell wall and by extension the plant ABSTRACT Abstract…. as a whole. Therefore, the bacteria having only the three cellulose degrading enzyme viz., endoglucanase, exoglucanase and β-glucosidase cannot degrade wood independently. S. plymuthica NAULK-3 had only these three enzymes. Isolate P fluorescens NAULK-2 produce all the three cellulose degrading and xylan degrading enzymes viz., endoglucanase, exoglucanase, β-glucosidase and xylanase. It was more efficient then the S. plymuthica NAULK-3 in degradation of wood, however, were not capable to degrade fully. P. viridivida NAULK-1 produced four different enzymes produced by the P. fluorescens NAULK-2 and additionally it produced laccase to digest lignin. Accordingly, it would be most capable to digest the wood as it have almost all the enzymes to degrade all the principal components of the plant wood. All the three isolates required neutral pH, however, P. viridivida NAULK-1 required 45oC temperature, P. fluorescens NAULK2 required 350C temperature and S. plymuthica NAULK-3 required 30oC temperature for their optimum expression. Expression of all these were maximum at 28 days post inoculation on the CMC and cellulose. Microorganism began the utilization of the natural agricultural/ agroindustrial wastes viz., wheat bran/straw, saw dust and rice straw within 7 days and the optimum time to degrade different substrate varied according to the substrate utilized and microorganism used. Among the different substrates, what straw/bran and rice straw was easiest to degrade by all the isolates, almost equal time was taken by all the isolated to degrade both the natural products, however, among the different isolates P. viridivida NAULK-1 degraded it earliest (21 days for the 50 per cent digestion and 28 days for the complete digestion), followed by P. fluorescens NAULK-2 (28 days for the 50 per cent digestion and 35 days for the complete digestion). Isolate S. plymuthica NAULK-3 took 35 days for the 50 per cent digestion and 42 days for the complete digestion. In the case of saw dust as substrate P. viridivida NAULK-1 took 28 days for Abstract…. the 50 per cent digestion and 35 days for the complete digestion. Isolate P. fluorescens NAULK-2 took 35 days for the 50 per cent digestion and 49 days for the complete digestion and isolate S. plymuthica NAULK-3 took 42 days for the 50 per cent digestion and 56 days for the complete digestion. The data were in accordance to the synthetic media, however, the long time to degrade the same over the synthetic media might be because of the complex structure of these natural molecules. For bacterial genomic DNA analysis 27F & 1525R primer were used to amplify ~1.5 kB DNA band size at 500C annealing temperature. In PCR all the isolates gave ~1.5 kB DNA band.ThesisItem Open Access Biological and Molecular Characterization of Chilli Leaf Curl Virus (ChiLCV) in South Gujarat(DEPARTMENT OF PLANT PATHOLOGY N. M. COLLEGE OF AGRICULTURE NAVSARI AGRICULTURAL UNIVERSITY NAVSARI, 2017-07) DORE R; Mahatma, LalitStudy was undertaken to study the Chilli leaf curl virus (ChiLCV) in South Gujarat. High incidence of the disease in the different fields of NAU as well as the adjoining area could be seen during the survey in 2016. ChiLCD manifesting typical leaf curl symptoms were seen in all the areas surveyed ranging from 10-35 per cent. The variation in disease incidence was mainly due to the different stages of the crop at different place and the different management practices followed. In every infected field surveyed, whiteflies were found invariably. However, jassids and thrips were the other insect pests also noticed. The typical symptoms of the disease observed during the investigation were typical leaf curl symptoms with the upward/downward curling of leaves, puckering and reduced size of leaves along with the thickening and swelling of veins. Affected plants grew slowly and became stunted or dwarfed. Fruit, if produced at all, were small, dry and unmarketable. Quality viral DNA was isolated and in PCR a part of DNA-A molecule of~1200 bp was amplified by the Begomovirus specific primers confirming it to be Begomovirus. A 932 bp nucleotide sequence was obtained on sequencing. Analysis and comparison of the sequence with the other standard DNA-A molecule of the Begomovirus indicated that the amplified fragment have two genes viz., virus coat protein (V1) gene and pre coat protein (V2) gene. The sequenced virus showed highest identity 99% with Chilli leaf curl India virus isolate India:Sonipat:TC290:2010 segment DNA-A, [KJ649706.1]. According to the recent criteria for the nomenclature of the virus was considered and named as Chilli leaf curl India virus – India: Sonipat: Navsari: 2016 abbreviated as ChiLCIV-[IN:Son:Nvs:16]. Alignment of the recently sequenced virus during present investigation [ChiLCIV-[IN:Son:Nvs:16] with other 45 different viruses showed the evolution pattern and similarity of the virus with the other Begomoviruses in the world. Different isolates of Chilli Leaf Curl Virus (ChiLCV) align distinctly in separate clad in the phylogenetic tree prepared.ThesisItem Open Access Biological and molecular characterization of Okra yellow vein mosaic virus (OYVMV) infecting okra in South Gujarat(Plant Pathology Dept., NMCA, Nau, Navsari, 2016) Ghevariya, Tushar V.; Mahatma, LalitIncidence of Yellow vein mosaic disease (YMD) of the Okra ranging from 1050 per cent in the different field of NAU as well as the adjoining area was observed during the survey in 2015 in late winter to summer. Total seven different fields of Okra in the villages around the Navsari Agricultural University, viz., Allu, Vankaner, Sejvad, Hathuka, Bhatlav, shiker and Delvada were surveyed. The crop grown in the Vegetable Research Farm of NAU, Navsari in open fields was also surveyed. The disease appeared with the appearance of true leaves and was seen at all the stages of the growth. YMD in okra appeared with the appearance of true leaves and was seen at all the stages of the growth. The initial symptoms on young leaves were diffuse and mottled in appearance. Clearing of the small veins starts near the leaf margins, at various points however, were more pronounced towards the petiole end as compared towards the expanded margins of the leaf. Gradually the yellowing increases covering the interveinal area and entire leaf appeared yellow. In few cases the leaf necrosis has also been reported. Fruit, stem and above ground parts of the infected plants also turned yellow in color. Early infected plant remains dwarf. Market acceptability of the yellow colored fruits reduced severely causing enormous economic loss. In some of the plants no green colour symptoms were seen, however, had another type of symptoms principally characterized by the vein thickening. Plant if seen from the top or side, appeared rough, rugose, dark green and dwarfed. In case of severe infection the entire leaf crumpled, curled inside, petiole bend and even cannot hold leaf up. Swelling of the vein and enation can also be seen on the backside of the leaf. In graft transmission the similar symptoms were also observed, however, the vein thickenings were not observed. In PCR A ~1200 bp band of DNA-A molecule was obtained by the Begomovirus specific primers. On sequencing the virus had maximum 98 per cent sequence homology with the Okra yellow vein mosaic virus isolate Aurangabad (GU181356.1), therefore, it was considered as a strain of this virus only. Accordingly the sequenced isolate has been considered as tentative strain of Okra yellow vein mosaic virus [Aurangabad] and named as Okra yellow vein mosaic virus [Aurangabad] [India:Nvs:Okra:2016] and was abbreviated as OYVMV [Aurangabad] [India:Nvs:Okra:2016]. Study of the Phylogenetic tree of the viruses from the different crops and geographical area revealed that the virus isolated and sequenced during the present investigation is closely related to the viruses of okra and cotton from the different parts of the world.ThesisItem Open Access Biological and Molecular Characterization of Tomato Leaf Curl Virus (ToLCV) in South Gujarat(PLANT PATHOLOGY DEPT.,N.M.COLLEGE OF AGRICULTURE, NAVSARI AGRICULTURAL UNIVERSITY, NAVSARI, 2016-04) Vanthana, M.; Mahatma, LalitIncidence of Tomato Leaf Curl Disease (ToLCD) in the protected as well open field condition was observed during the present investigation to the tune of 10-50 per cent. The typical symptom includes upward/downward curling along with the vein clearing of leaves. Fruit, if produced at all, were small, dry and unmarketable. Initially the newly emerging plant leaflet started showing mild vein clearing and leaf curling symptoms. Symptoms gradually become severe showing characteristic moderate to severe leaf curling (upward or downward) and shrinking of the leaves. Leaves were often stiff, thicker and of leathery texture rather than flaccid. Affected plants grew slowly and became stunted or dwarfed. The flowers appear normal. No chlorotic or yellowing of the leaf lamina could be seen. The virus was transmitted by approach as well as side veneer grafting. In PCR a part DNA-A molecule of ~1200 bp was amplified by the Begomovirus specific primers confirming it to be Begomovirus. A 1125 bp nucleotide sequence (Accession number KU921251) was obtained on sequencing. Amplified fragment had two genes viz., virus coat protein (V1) gene and pre coat protein (V2) gene. The sequenced virus showed highest identity (94%) with Tomato leaf curl Gujarat virus [Nepal] segment DNA A, [AY234383.1] followed by 93% sequence identity with thirteen different tomato Begomoviruses which were the strains/isolates of Tomato leaf curl Gujarat virus or Tomato leaf curl New Delhi virus [DQ629101.2]. The virus was named as Tomato leaf curl Gujarat virus [Nepal] [India:Nvs: Tom:2016] and was abbreviated as ToLCGV [Nepal] [India:Nvs:Tom:2016]. Alignment of the sequenced virus with the other Begomoviruses of the different crops from the different location indicated that the virus was typically a leaf curl virus. Virus which produced yellow leaf curl symptoms and yellow mottle symptoms were entirely different from the virus characterized during the present investigation. Two distinct categories of virus producing different types of symptoms viz., Leaf Curl (LC) and Yellow Mottle (YM) are included in the ToLCD. It is very difficult to recognize the symptoms of the viral disease by the virus name as found in other viruses. Both the categories of the virus have been reported from the area in the past, however, the study concluded that the ToLCD is predominantly caused by the Tomato Leaf Curl Virus belonging to Leaf Curl category in the area surveyed. All the morphological, biological and molecular results were found supporting, that the virus studied was leaf curl type only.ThesisItem Open Access CHARACTERIZATION OF COAT PROTEIN GENE (AV -1) OF Mungbean yellow mosaic virus (MYMV) INFECTING MUNGBEAN IN SOUTH GUJARAT(PLANT PATHOLOGY DEPT., N.M.COLLEGE OF AGRICULTURE, NAVSARI AGRICULTURAL UNIVERSITY, NAVSARI, 2014-09) Solanki, Hiren P.; Mahatma, LalitAim of the present investigation was to characterize and use of AV-1 gene of the yellow mosaic disease (YMD) causing Mungbean yellow mosaic virus for the cloning in the bacterial system and over expression of the coat protein. The protein over expressed in bacterial system can be used as antigen to elicit immune response in animal and production of antibodies for serological investigation. YMD in Navsari has been observed to caused by MYMV-Vig:IN:NVS:Mg:2012. The strategies included isolation of DNA, in silico characterization of the AV-1 gene and its putative protein for the study of antigenicity and further its over expression. Disease produce typical yellow mosaic symptoms which became brighter and enlarge to some extent with the age; however, depletion of the chlorophyll could not be observed once synthesized. Severity of the symptoms could be seen in the newly eme rging leaves. Good quality viral DNA was isolated from younger symptomatic leaves of mungbean plants by CTAB method and amplified ~700 bp of the DNA-A molecule of the begomovirus infecting mungbean. Sequence of MYMV-Vig:IN:NVS:Mg:2012 (JQ004982.1) previously characterized by this laboratory was retrieved from the NCBI database. The gene was 420 bp long synthesizing 139 amino acids long putative protein. The same sequence was used for the study. Sequence was highly conserved. Epitopes of the amino acids was well spread in the entire amino acid. Epitopes were found shifted towards the N -terminal site of the amino acids. Maximum 13 amino acids long peptide sequence was found in the amino acid sequence starting at 42 positions till 54 t h positions (YDNEPSTATVKND) of the putative amino acid sequence. Study of the amino acid sequence indicated that the hydrophilisity in the sequence was spread all over the sequence; however, as epitopes, it was also little more shifted towards the N - terminal region of the protein. The protein was water soluble. Study of the different properties of protein revealed that the protein being targeted was of good quality. From the study it was evident that the putative protein would be of high antigenicity and would produce good quality antibodies. Attempt was made to transform the AV -1 gene in the pETite vector. It could be transformed in the bacteria system, however, could not over expressed the protein. This might be either due to the toxic effect of the gene product on the bacterial gr owth of alteration of essential part of the genome of the bacteria during the cloning which lead to the lethal effect.ThesisItem Open Access Epiphytology of mungbean Yellow Mosaic Virus (MYMV) in Mungbean (Vigna radiata (L.) Wilczek) Under South Gujarat Condition(Plant Pathology Department, N. M. College of Agriculture Gujarat Agricultural University, 2011-06) Dahyabhai, Patel Vinay Kuamr; Mahatma, LalitMungbean which is also known as green gram (Vigna radiafa (L.) Wilczek), is the third most important pulse crop grown in India. Mungbean suffers from several diseases with substantial losses in yield. Of all the diseases, Mungbean Yellow Mosaic Virus (MYMV) disease is the most destructive, which takes heavy toll in Indian subcontinent and adjacent areas of South-East Asia. Nevertheless, not much work has been carried out on epiphytology and estimation of yield losses due to the disease. Therefore, the present investigation was carried out on the topic entitled "Epiphytology of Mllngbean Yellow Mosaic Virus (MYMV) in mungbean (Vigna radiala (L.) Wilczek) under south Gujarat condition". During the survey, occurrence of Munghean Yellow Mosaic Virus (MYMV) in munghean was noticed in serious proportion causing heavy losses in Navsari and Surat district. The disease incidence in Surat and Navsari districts ranged from OR.66 to 5H.96 per cent. Fixed plot surveys were carried out at different field of NAlJ at different stages of crop in different varieties. None of the variety showed resistance against the disease. The first symptoms of the disease was ohserved as early as the first tri foliate emerged and up-to 79 to 100 per cent incidence was recorded at the time of crop maturity. Whiteflies might have brought primary inoculums of MYMV at-least two kilometers away from the site of experiment anytime in between 14 days before the sowing of crop to the 4 days after the sowing of crop to inoculate the pathogen at the site of experimentation immediately after the emergence of the crop. Then, the virus might have spread from one plant to other plants through its vector, whitefly. Maximum numbers of whiteflies (148) were trapped at North East direction. Whereas 128, 123 and 123 whiteflies were observed in North West, South East, South West direction respectively during summer 20 I O. Average number of whiteflies on 10 plants ranged from 0 to 2.5. Whitefly population had signi ficant • • positIve correlation with maximum temperature, average temperature and relative humidity. In the present investigation 3SoC • ma:(\mum temperature, 31 .1 °C mean temperature, 90 % morning RH. 53 OJ, , • a fter noon RH and 71.5 % mean R H found to be optimum for the build-up of whitefly population. Disease incidence was found significantly positively correlated with whitefly population. Minimum temperature, average temperature, afternoon relative humidity, average relative humidity and wind velocity found to be positively correlated with MYMV incidence . The multiple linear regression equation showed that different weather parameters were together responsible for the whitefly population to the extent of 90.38 per cent. Weather factor (Minimum temperature) was responsible for the disease incidence to the extent of 94.32 per cent . The potential yield losses due to the disease depend upon the stage of infection. An early infected plant suffers 74.50 per cent loss. The plot which was completely protected by the spraying of systemic insecticide yielded 32.59 per cent more yield than the field left unsprayed.ThesisItem Open Access Interaction of Mungbean Yellow Vein Mosaic virus and Rhizobium Sp. in Mungbean [Vigna radiata (L.)(Plant Pathology Department, N. M. College of Agriculture Gujarat Agricultural University, 2014-07) Chudasama, Mitesh Khimjibhai; Mahatma, LalitYellow Mosaic Disease of the mungbean caused by a variant MYMV-Vig: IN: NVS: Mg: 2012 in the South Gujarat. Rhizobium sp. is a symbiotic nitrogen fixing bacteria remain associated with the legumes. The present research work was aimed to find the interaction of Rhizobium sp. and MYMV for the exploiting the mechanism to reduce the disease and improve the yield and productivity. Ten different samples were collected to isolate the Rhizobium strain of mungbean. Through the isolation and initial biochemical characterization. three isolates (NAURh-l. NAURh·2 and NAURh-7) out of the ten isolates were suspected to be the Rhizobium sp. They • I showed white, round, convex, glistening, mucoid and opaque colonies on YEMA. These isolates showed negative reaction in IMViC test, H 2 S test, starch hydrolysis test, caesin hydrolysis test, nitrate reduction test, Gelatin hydrolysis test, whereas positiveAbstract reaction in oxidase and urea hydrolysis test, All the three isolates , look alike, therefore, only one isolate NAURh-2 was promoted for the further research. Among the different antibiotic tested, Sparfloxacin and Tetracycline were placed in the categories of ch loram phen ico I. intermediate; co-tri moxazo Ie, levofloxacin, ofloxacin, cefotaxime, gentamycin, tobramycin and streptocycline were susceptible oxacillin, • and • Im'penem, ciprofloxacin, meropenem, cafalaxin, cephalothion , moxifloxacin, amoxyclav, penicillin, ampicillin, erythromycin, vanomycin, clindamycin and sulphatriad were of resistant categories. Through the Biolog study and subsequently manual analysis of the data and biological studies it was concluded that the isolate (NAURh-2) was Rhizobium iegllmil1osarllm . Cell length of the bacteria varied from 1.2 to 3.0 I'm, whereas cell width varied from 0.5 to 1.0 I'm . The cells from the freshly prepared culture were Gram negative and found regular rod shaped. Cells were either singly or arranged in aggregates. No spore formation was observed even on the old and stressed cultures. Bacteria produces capsule were motile with peritrichous flagella . Nodulation in the crop could be seen at the 12 days onwards after the inoculation. Maximum 28 nodules per plant were observed at 32 post inoculation thereafter , it started decreasing in the present situation. Detail steps of the nodulation were studied. Significantly higher nodules were observed in the plants inoculated from the Rhizobium ieguminosarllm treatment, seed in the control treatment with condition. the Among Rhizobilllll the different iegllmil10sarumAbslraCI IOmllkg was found significantly superior. No effect ~f the early nodulation on the Rhizobium sp . nodulation and vice versa was observed. If this would have been observed as per the few published literature, this would have been exploited for the minimization of losses caused by the MYMV and enhancement of productivity of the high demanded protein source of vegetarian.ThesisItem Open Access INTERACTION OF MUNGBEAN YELLOW VEIN MOSAIC VIRUS AND RHIZOBIUM SP. IN MUNGBEAN [Vigna radiata (L.) Wilczek](PLANT PATHOLOGY DEPT., N.M.COLLEGE OF AGRICULTURE, NAVSARI AGRICULTURAL UNIVERSITY, NAVSARI, 2014-07) Chudasama, Mitesh K.; Mahatma, LalitYellow Mosaic Disease of the mungbean caused by a variant MYMV-Vig: IN: NVS: Mg: 2012 in the South Gujarat. Rhizobiu m sp. is a symbiotic nitrogen fixing bacteria remain associated wit h the legumes. The present research work was aimed to find the interaction of Rhizobium sp. and MYMV for the exploiting the mechanism to reduce the disease and improve the yield and productivity. Ten different sa mples were collected to isolate the Rhizobium strain of mungbean. Through the isolation and initia l bioche mical characterization, three isolates (NAURh-1, NAURh-2 and NAURh-7) out of the ten isolates were suspected to be the Rhizobium sp. They showed white, round, convex, glistening, mucoid and opaque colonies on YEMA. These isolates showe d negative reaction in IMViC test, H 2 S test, starch hydrolysis test, caesin hydrolysis test, nitrate reduction test, Gelatin hydrolysis test, whereas positive reaction in oxidase and urea hydrolysis test, All the three isolates look alike, therefore, only one isolate NAURh-2 was pro moted for the further research. Among the different antibiotic tested, Sparfloxacin and Tetracycline were placed in the categories of intermediate; chloramphenicol, co- trimoxazole, levofloxacin, ofloxacin, cefotaxime, gentamycin, tobramycin and streptocycline were susceptible and ciprofloxacin, cafalaxin, cephalothion, oxacillin, imipene m, meropenem, moxifloxacin, a moxyclav, penicillin, a mpicillin, erythro mycin, vano mycin, clindamycin and sulphatriad were of resistant categories. Through the Biolog study and subsequently manua l analysis of the data and biological studies it was concluded that the isolate (NAURh-2) was Rhizobium leguminosarum. Cell lengt h of the bacteria varied from 1.2 to 3.0 μm, whereas cell widt h varied from 0.5 to 1.0 μm. The cells from the freshly prepared culture were Gram negative and found regular rod shaped. Cells were either singly or arranged in aggregates. No spore formatio n was observed even on the old and stressed cultures. Bacteria produces capsule were motile with peritrichous flagella. Nodulation in the crop could be seen at the 12 days onwards after the inoculation. Maximum 28 nodules per plant were observed a t 32 post inoculation thereafter, it started decreasing in the present situation. Detail Significantly steps higher of nodules the were nodulation observed were in the studied. plants inoculated from the Rhizobium leguminosarum in the contro l condition. Among the different treatment, seed treatment with the Rhizobium leguminosarum 10ml/kg was found significantly superior. No effect of the early nodulation on the Rhizobium sp. nodulation and vice versa was observed. If this would have bee n observed as per the few published literature, this would have bee n exploited for the minimization of losses caused by the MYMV and enhance ment of productivity of the high de manded protein source of vegetarian.ThesisItem Open Access Investigations of Mungbean yellow Mosaic Virus (Mymv) of Mungbean (Vigna Radiata (L.) Wilezek)(Department of Plant pathology College of Agriculture Gujarat Agricultural Universit, 2010-07) Pawar, Dayaneshwar Madhukarrao; Mahatma, LalitMungbean (Vigna radiata (L.) Wilczek) is one of the important pulse crops primarily grown for food in India. During the survey, occurrence of Mungbean Yellow Mosaic Virus (MYMv) in mungbean was noticed in serious proportion causing heavy losses in Navsari, Surat and valsad districts;. Cultivars GM-4 and K-851 were found more severely affected at the flowering stage during summer season . Considering the seriousness of the problem, the present investigation was carried out on transmission and detection of virus. Roving field surveys were under taken in and around Navsari , Surat and Val sad districts, the total 16 locations to find out occurrence of the Mungbean Yellow Mosaic Virus (MYMv) during summer, 20 I O. The MYMv incidence increases with on increase in crop stages. Crop at the first trifoliate leaf stage was 08-12 per cent, second trifoliate leaf stage was 16-20 per cent and before flowering stage 4 I to 55 % incidences were reloaded and at the time of maturity stage showed 65-76 per cent incidence of MYMv. Fields of Sugarcane Research Station, NAU, Navsari farm were observed periodically. Three popular varieties of mungbean viz., K-851, GM-3 and GM-4 were sown in the field. None of the variety showed resistance against the disease and 75 to 100 per cent incidence was reloaded at the time of crop maturity. Among these, K-8S1 showed cent per cent MYMV incidence. The disease appeared in the field as small scattered yellow to golden yellow colour flecks on the infected leaves. These were scattered on the entire leaves and were more concentrated near the leaf venation. The severity of the symptoms could be seen in the newly emerging leaves, where in case of high susceptibility, cent per cent area of the leaf turned yellow. The symptoms could be observed on all the green coloured aerial parts of the plants including cotyledon leaf, trifoliate leaves, stem, petiole, flower part, pod and seeds. Infected plant remained stunted with few pods of small size and shrivelled seeds. The peR based diagnostic protocol was standardized to amplify viral DNA, six different primer pairs were used. Presence of DNA-A and DNA-8 molecules of the virus could be detected by the different sets of the primers. Self designed primer pair, LM-20F + LM-20R was best and replicative. Therefore, the same primer pair was used for the different diagnostic and localization purpose throughout the study. The virus was not found to be transmitted by seeds, mechanical sap and aphid in the susceptible plants. Whitefly (Bemisia tabaci, Gennadius) could acquire virus i~ 30 minutes of acquisition feeding and similarly could inoculate in 30 minutes of inoculation feeding period. There was effect of fasting on the transmission of virus. for cent percent virus transmission, whiteflies needs 6 hours of acquisition and inoculation feeding period. MYMV could be detected from the sepal, standard petal, wing petal, keel petal and androecium . However, MYMV could not be detected from gynoecium. Among the parts of seeds, MYMV could be detected from the pod , seed coat and cotyledon. However, presence of MYMV could not be detected from embryo of the seed . From the results , it is concluded that the seeds act as a passive source of primary inoculation and may be termed as passive transmission of the disease or virus through seeds . Presence of MYMV in the callus induced from the infected cotyledon could be detected. This is the first report of the detection of any Begomovirus from the seeds or any part of the seeds. Further, the virus could be multiplied (cultured) artificially in the laboratory condition.ThesisItem Open Access Isolation And Characterisation Of Antiviral Substances From Plants(Anand Agricultural University; Anand, 2004) Mahatma, Lalit; Mishra, AshokThesisItem Open Access Molecular and Biochemical Characterization of Oil Degrading Bacteria(DEPARTMENT OF PLANT MOLECULAR BIOLOGY AND BIOTECHNOLOGY N. M. COLLEGE OF AGRICULTURE NAVSARI AGRICULTURAL UNIVERSITY Navsari, 2016-07) Topivala, M.A.; Mahatma, LalitPresent investigation was carried out on “Molecular and biochemical characterization of oil degrading bacteria”. Total six isolates from the different area were isolated. All these six isolates showed lipid solubilization index in the range of 5.4 to 2.33. NAULM-1 showed maximum lipid solubilization index (5.4). NAULM-4 showed lowest 2.33 lipid solubilization index. NAULM-2, NAULM-3, NAULM-5 and NAULM-6 gave 3.3, 2.85, 3.2 and 2.43 solubilization index respectively. All the isolates were Gram negative rod-shaped, motile and non spore forming bacteria. On solid media all these bacterial isolates showed raised elevated colonies with undulated margins. Best pH for the lipase production was neutral and the production of the same decreases with the further increase or decrease of the pH. Temperature 35ºC, was found optimum for lipase production by three isolates (NAULM-1, NAULM-2 and NAULM-4 with 8.01 U/ml, 5.72 U/ml and 4.91 U/ml lipase production at 35ºC whereas 30ºC temperature was optimum for NAULM-3 (4.91 U/ml lipase). One per cent concentration of the substrate was most suitable for the lipase production. ABSTRACT Abstract… To supply the organic nitrogen source, casein was best for the isolate NAULM-1, NAULM-2 and NAULM-3. Whereas tryptone was most preferred by NAULM-4. Among the different inorganic nitrogen sources NaNO3 was preferred by isolates NAULM-1, NAULM-2 and NAULM-4 whereas maximum 5.35 U/ml lipase expressions was observed in NAULM-3 in KNO3 as inorganic nitrogen source. Overall NAULM-1 and olive oil was the best combination of isolate and substrate source for the efficient lipase production in vitro. Heavy metal Cu, Pb, Cd. drastically reduced expression of lipase in vitro. The crude enzyme worked best with high stability at pH 7.0 and temperature 35ºC. Activities of the enzyme was not much affected at lower temperature up to 25ºC tested by all the isolates. All the isolates were identified using Biolog Microlog GN2 microplates and by partial sequencing of the 16S rDNA sequencing. Universal primer pairs, 27F and 1525R amplified ~1.5 kb fragment of partial 16S rDNA in all the isolates. On sequencing obtained 1153 bp sequence of NAULM-1, 999 bp sequences of sequence of NAULM-2, 1166 bp sequence of NAULM-3 and 960 bp sequence of NAULM-4. The results of the Biolog and Sequencing matched. On the basis of their identification the isolate NAULM-1 was tentatively named as Pseudomonas pseudoalcaligenes (LCB-14) NAULM-1, isolate NAULM-2 was tentatively named as Pseudomonas aeruginosa (ABG5) NAULM-2, isolate NAULM-3 was tentatively named as Pseudomonas aeruginosa (M2) NAULM-3 and isolate NAULM-4 was tentatively named as Pseudomonas aeruginosa (PS_4) NAULM-4. All these isolates belong to pseudomonas cluster of Gamma-Proteobacteriaceae. Pseudomonas is one of Abstract… the widely recognized species for the production of lipase. P. pseudoalcaligenes had been used in the past for bioremediation and commercial lipase production by many of the organizations. All the isolates can be used for the commercial lipase production; however, Pseudomonas pseudoalcaligenes (LCB-14) NAULM-1 found most preferred and can be exploited for the purpose.ThesisItem Open Access Molecular characterization and metabolic profiling of banana (Musa × paradisiaca L.) pseudostem waste degrading microbes”(NAU, 2015) Patel, Hiren Kanubhai; Mahatma, LalitSix different bacterial isolates were isolated from the different decomposing samples of the banana collected. These were distinct and named as Bacillus licheniformis NAULH-1, Bacillus wakoensis NAULH- 2, Bacillus licheniformis NAULH-3, Bacillus wakoensis NAULH-4, Serratia marcescens NAULH-5 & Brevibacillus agri NAULH-6, respectively. Cellulase (cell), β-glucosidase (Bgl) and exoglucanase (Exgl) gene were present in all six isolates. Laccase (Cot A) and xylanase (xyn) gene was present only in four isolates viz., Bacillus licheniformis NAULH-1, Bacillus wakoensis NAULH-2, Bacillus licheniformis NAULH-3, Bacillus wakoensis NAULH-4. Lignin perooxidase (Lip) gene was found in three isolates (Bacillus licheniformis NAULH-1, Bacillus wakoensis NAULH-2, Bacillus wakoensis NAULH-4) only. Partial sequencing of the laccase gene of all four isolates indicated the presence of Cot-A conserved domain of cupredoxin superfamilies. B. wakoensis NAULH-2 showed CuRo_1_BOD_Cot-A like domain at 1- 140 amino acid sequence having trinuclear copper binding site at 70-122 amino acid sequence. B. licheniformis NAULH-1 had 169 amino acid sequences having two conserve domains of CuRo_2 _Cot-A and CuRo_3 _Cot-A having 1-27 and 55-166 amino acid sequence, respectively. B. licheniformis NAULH-3 had only one domain CuRo_1_BOD_Cot-A domain having 37-111 amino acid sequence, while B. wakoensis NAULH-4 contained two conserve domains CuRo_1_BOD_Cot-A and CuRo_2 _Cot-A having 1-22 and 33-51 amino acid sequence respectively. The data revealed by 16s rDNA sequencing indicated that B. wakoensis NAULH-2 and B. wakoensis NAULH-4 had 99% similarity with the Bacillus sp., while B. licheniformis NAULH-1 and B. licheniformis NAULH-3 showed 99% similarity with Bacillus licheniformis. All six isolates showed production of endoglucanse, exoglucanase and β-glucosidase enzymes. Four isolates viz., B. licheniformis NAULH-1, B. wakoensis NAULH-2, B. licheniformis NAULH-1 and B. wakoensis NAULH-4 produced laccase and xylanase enzymes. The phenotypic microarray of metabolic profiling indicated that all bacterial isolates have specific ability to utilize complex carbohydrates except Brevibacillus agri NAULH-6. In banana pseudostem degradation facilities B. wakoensis NAULH-2 showed fastest and highest degradation of banana pseudostem at neutral pH, while B. wakoensis NAULH-4 showed higher degradation at alkaline pH 8. B. licheniformis NAULH-1 degraded higher amount of banana pseudostem at 30-35OC compare to all isolates, while at higher temperature 40-45OC, B. wakoensis NAULH-2, B. licheniformis NAULH-3 and B. wakoensis NAULH-4 degraded higher amount of banana pseudostem. All the isolates showed optimum efficacy of banana pseudostem degradation at 28 days. Laccase produced by Cot A gene is the major enzyme responsible for lignin degradation. Till date fungal laccase were used for lignin degradation, which is the major obstacle in degradation of banana pseudostem. The identification of new bacterial laccases gene would open up new perspectives for rapid biotransformation of the cumbersome waste in to carbon rich substances for the eco-friendly agricultureThesisItem Open Access Molecular Characterization of Mungbean yellow Mosaic virus Infecting Mungbean (Vigna radiata (L.) Wilczek) in South Gujarat Agro Climatic Conditions of India(Plant Pathology Department, N. M. College of Agriculture Gujarat Agricultural University, 2012-06) Charles, Mugisa; Mahatma, LalitMungbean or Greengram [Vigna radiala (L.) Wilczek] is an important Pulse grown in India is a rich source of protein and Iron. Besides nourishing the people, the cultivation of Mungbean can sustain soils of India by adding Nitrogen through rhizobial symbiosis. However, Yellow mosaic disease (YMD) caused by Mungbean Yellow Mosaic Virus (MYMV) remains the most destructive disease affecting yield potential of mungbean both quantitatively and qualitatively. To assess the incidence of the disease in Navsari roving field surveys were carried out during summer, 20 I I in major mungbean growing areas of Navsari, Gandevi, Jalalpur, Vansda, Chikhli and taluka. The disease incidence ranged from 06.54 to 33.21 per cent. Crop at the first trifoliate leaf stage at NAU farm, Ugat, and Salem villages of Navsari taluka showed 07.84-13.02 per cent incidence. The incidence increased with the crop growth stages. Around 14.78-17.02 per cent incidence was observed in second trifoliate leaf stage at Pepalkhed, Lakhawadi and Vadichota villages of Vansda taluka. Disease incidence of 30.12-33.21 per cent at Sadakpor and Pepal Gabhar villages of Ckikhli Taluka when observed just before the flowering. The symptoms initially appeared as small scattered yellow to golden yellow colour flecks on the infected trifoliate. The scattered symptoms gradually enlarged covering entire leaves. Severity of the symptoms was mOre on newly emerging leaves of the infected plant. peR amplified and sequenced part of DNA-A molecule gave a 1285 bp nucleotide sequence. The sequence has been submitted to GeneBank, NCBI, and was given accession number JQQ04982. Sequence is having one gene on the virus strand and three genes- on the complementary strand. Analysis and comparison of the sequence with the other standard universal ORF of the DNA-A molecule indicated that the amplified fragment have four genes viz., virus coat protein (AV I) gene (partial cds); replication associated protein (AC3) gene (complete sequence): transactivator protein (AC2) gene (complete cds) and replicase (AC!)ThesisItem Open Access Morphological and Molecular Churacterizotion of Arbuscular Mycorrhizal Fungi From the South(DEPARTMENT OF PLANT MOLECULAR BIOLOGY AND BIOTECHNOLOGY N. M. COLLEGE OF AGRICULTURE NAVSARI AGRICULTURAL UNIVERSITY Navsari, 2016-06) Tandel, M.H; Mahatma, LalitThesisItem Open Access TRANSCRIPTOME ANALYSIS OF PIGEON PEA (Cajanus cajan L.) DURING VARIOUS TEMPERATURE, DROUGHT AND HEAVY METAL STRESSES(PLANT MOLECULAR BIOLOGY & BIOTECHNOLOGY DEPT., N. M. COLLEGE OF AGRICULTURE, NAVSARI AGRICULTURAL UNIVERSITY, NAVSARI, 2014-03) Radadiya, Nidhi G.; Mahatma, LalitDuring the present investigation, different biochemical and molecular parameters in pigeon pea were analyzed during various temperature, drought and heavy metal stress conditions. Over all maximum activities of Glutathione-S-transferase was observed in the leaves and roots exposed to the 7 % PEG stress followed by 500 μmol CdCl2 in 3 days after the exposure to the stress. The data suggested that the Glutathione-S-transferase is a principle enzyme expressed in response to the drought and heavy metal stress. Expression of the enzyme was less in the higher concentration of PEG (15%) and CdCl2 (500 μmol). This may be due to the toxicity of these at the higher concentration. Maximum expression of glutathione reductase was observed in 650 μmol CdCl2 in 3 days after the exposure to the stress followed by the 15 % PEG exposure. Data suggested that the glutathione reductase is also one of the key enzyme expressed in response to the drought and heavy metal stress, however, is capable to tolerate higher concentration of PEG and CdCl2 than GST. Maximum expression of ascorbate peroxidase was observed in 7 per cent PEG followed by 15 per cent PEG and chilling stress at 3 days after the exposure to the stress. Data suggested that the ascorbate peroxidase is also a key enzyme for the drought, however, it is also found in the response to the chilling stress. The Glycine betaine (GB) content was higher in drought stress compared to other stresses and control seedlings after 3 days of stress whereas, in chilling stress the GB content was reached up to 208.09 μmol. In heavy metal stress GB level was increased up to 144.06 μmol and 159.86 μmol at 500 μmol and 650 μmol CdCl2 concentration respectively. After 6 days of all stresses seedlings were almost maintain the GB level which noted at 3 days after stress. Among the different polyamines, putrescine was detected in all stressed seedlings of pigeon pea at all the stages of analysis whereas cadavarine was detected in only heavy metal and 15% PEG treated seedlings leaves. Spramine and spermidine were not detected in heavy metal stress but detected in drought and cold stress. In drought stress treatment the per cent area of putrescine was higher at the 3 days after treatment which further decreased slightly at 6 days after treatment. Heavy metal and chilling stress also showed the higher putrescine level than the control seedlings leaves. Spermidine was also detected in drought and chilling stress, however, was not detected in heavy metal stress. Cadavarine was detected in heavy metal stress. Maximum expression of SAMs gene in the leaves were observed at 3 days after drought stress with 15% PEG that is increased by 6.543 fold as compared to control. Similarly, maximum SAMs gene expression in the roots were observed at the same time period and same water potential that is increased by 2.428 fold as compared to control. Conversely, expression of SAMs gene were not detected in leaves and roots at cadmium stress but transcripts were down regulated as compared to the control. Expression of SAMs gene was also studied in leaves and roots at chilling stress (10oC). SAMs gene expression were increased by 6.32 fold in leaves and 2.028 fold in roots at 3 days after stress. After 6 days of stress expression of SAMs gene observed that is increased by 1.537 fold in leaves and 1.064 fold in roots as compared to the control but it was lower than expression of SAMs gene which noted at the 3 days after stress.