http://localhost:8080/xmlui/handle/123456789/94 2026-05-01T23:30:21Z http://localhost:8080/xmlui/handle/123456789/505 Title: A STUDY ON ISOLATION, CHARACTERISATION AND IDENTIFICATION OF ENDOPHYTIC BACTERIA FROM SOME MEDICINAL PLANTS OF KOKRAJHAR DISTRICT, ASSAM Authors: DAS, DEBAJANI Abstract: ABSTRACT Endophytic bacteria are those that multiply intracellularly or intercellularly in their host plants at least once during their life cycle without showing any obvious symptom of disease. Plant-associated bacteria represent a vast and untapped source of unique phytochemical compounds, biofertilisers, and plant growth promoters that can be used as sustainable and natural alternatives to agrochemicals. Numerous issues in agriculture and health can be resolved by endophytic bacteria, including reducing environmental pollution brought on by the long-term use of chemical fertilisers and producing medications that will aid in the fight against drug resistance. In addition to their capacity to stimulate plant growth, a number of endophytes (which were previously isolated from a variety of plants) have shown antibacterial and anti-cancer properties. The ability of many endophytic bacteria to produce extracellular enzymes, such as lipases, amylases, pectinases, cellulase, xylanase and proteases, has been demonstrated. These enzymes can be used in various industries, including the food and beverage, textile, and leather sectors. There are several different types of endophytic bacteria that are economically important and can be isolated from medicinal plants. Although many medicinal plants have not yet been fully studied. With these considerations, the present study was carried out to isolate, characterise and identify endophytic bacteria present in some medicinal plants, like Glycosmis pentaphylla, Hygrophila auriculata and Phlogacanthus thyrsiformis, collected from different locations of the Kokrajhar district, Assam. For the isolation of endophytic bacteria, surface sterilisation plays an important role. In this study, surface sterilisation was done by using 70% ethanol and sodium hypochlorite in different concentrations and for different durations. The conformation of proper surface sterilisation was done by performing a sterility test. Here, 16 endophytic bacteria have been successfully isolated from the leaf, root and stem of three medicinal plants using the optimised surface sterilisation method. A total of 5 endophytic bacteria were isolated from the different tissues of G. pentaphylla: GPL-1, GPL-2 and GPL-3 from the leaves; GPS-4 from stems and GPR-5 from roots. Also, from H. auriculata, a total of 5 endophytic bacteria were isolated: HAL-1 and HAL-2 from leaves; HAS-3 and HAS-4 from stems; HAR-5 from roots. Whereas from P. thyrsiformis, 6 endophytic bacteria were isolated: PTS-1 and PTS-2 from stems; PTL-3 and PTL-4 from leaves; PTR-5 and PTR- 6 from roots. Morphological, microscopic and biochemical characterisation of the isolates vi | P a g ewas done using some standard protocol. For microscopic analysis, both Gram staining and scanning electron microscopy analysis were performed. Of the 16 isolates, 10 exhibited Gram-positive nature and 6 exhibited Gram-negative nature. Under microscope, almost all isolates were rod-shaped except PTL-4, which is circular in shape. In biochemical characterisation, almost all isolates showed positive for the catalase test and negative for the indole test. Almost all isolates showed positive for the oxidase test except PTL-3, PTL-4, PTR-6, HAL-2, HAS-3 and HAR-5, whereas GPL-2 is slightly positive as the purple colour is produced after some seconds (delayed production). All the isolates showed positive for the citrate test except PTR-6. In the methyl red test, all isolates showed negative results except HAL-2, which showed a positive result. Out of sixteen, eleven isolates showed positive and five showed negative for the Voges- Proskauer test. Antibiotic sensitivity tests were also performed on the isolates against a number of standard antibiotics. Isolates PTS-1, PTR-5, GPL-2, GPS-4, GPR-5, HAL-1, HAL-2, HAS-3, HAS-4 and HAR-5 showed resistant to ampicillin, whereas PTS-2, PTL- 3, PTL-4, PTR-6, GPL-1, and GPL-3 showed susceptibility to ampicillin. All the isolates showed resistant to penicillin. For gentamicin, almost all isolates were susceptible except GPR-5 and HAS-3, which showed resistant to gentamicin. Except for GPR-5, all isolates showed susceptibility to ciprofloxacin. Against cefotaxime, PTS-1, GPL-2, GPS-4, GPR- 5, HAL-1, HAL-2, HAS-3, HAS-4, and HAR-5 were resistant; PTR-6 showed intermediate resistant, whereas isolates PTS-2, PTL-3, PTL-4, PTR-5, GPL-1, and GPL- 3 showed susceptibility to cefotaxime. For molecular identification, the genomic DNA was extracted from all 16 bacterial isolates using the CTAB method with some modifications. After that, PCR amplification was done for all isolates using the universal primer for the 16S rDNA gene, namely, 27F (forward primer) and 1492R (reverse primer). Amplified products were checked in 1.5% agarose gel, and then sequencing was performed. After getting the sequence, identification was done by performing BLAST followed by constructing a phylogenetic tree using MEGA 11 software. Then, the endophytic bacteria were identified from three medicinal plants, which are as follows: Pseudomonas oryzihabitans strain HAL1DD (HAL-1), Proteus mirabilis strain HAL2DD (HAL-2), Stenotrophomonas geniculata strain HAS3DD (HAS-3), Agrobacterium cavarae strain HAS4DD (HAS-4), Lysinibacillus macrolides strain HARDD (HAR-5) from H. vii | P a g eauriculata; Alkalicoccobacillus gibsonii strain GPDD1 (GPL-1), Bacillus cereus strain GP2DD (GPL-2), Bacillus subtilis strain GP3DD (GPL-3), Bacillus cereus strain GP4DD (GPS-4), Bacillus australimaris strain GPRDD (GPR-5) from G. pentaphylla; Pseudomonas aeruginosa strain DD3 (PTS-1), Agrobacterium larrymoorei strain DDBU3 (PTS-2), Solibacillus silvestris strain DDBU6 (PTL-3), Kocuria assamensis strain DDBU9 (PTL-4), Alkalicoccobacillus gibsonii strain PTR1DD (PTR-5), Prescottella equi strain PTR2DD (PTR-6) from P. thyrsiformis. All the sequences were submitted to NCBI GenBank for accession numbers. Some plant growth promotion activities, such as phosphate solubilisation, production of ammonia, indole-3-acetic acid (IAA), and salt tolerance ability of the isolates, were carried out. Seven of the sixteen isolates exhibited positive results in the phosphate solubilisation activity. The highest solubilisation index was observed in PTS- 1 isolates, followed by PTR-6, GPR-5, PTR-5, GPL-3, PTS-2, and HAS-3. For ammonia production, almost all isolates showed positive results except PTS-1 and PTR-5. Eleven isolates out of 16 had shown positive results for their capacity to produce IAA in the presence of 400 μg/ml of L-tryptophan. The test was performed spectrophotometrically by comparing with standard IAA. Highest production was observed in HAR-5 (258.6 ± 2.05 μg/ml), which is followed by HAL-1 (121 ± 1.63 μg/ml), PTL-3 (81 ± 1.24 μg/ml), GPL-3 (80.16 ± 2.89 μg/ml), GPS-4 (76.66 ± 2.49 μg/ml), GPL-1 (76.33 ± 1.24 μg/ml), GPL-2 (75.83 ± 1.31 μg/ml), PTS-2 (67 ± 1.24 μg/ml), PTL-4 (45 ± 0.47 μg/ml), GPR-5 (31.66 ± 2.49 μg/ml), and PTS-1 (15 ± 0.81 μg/ml). A salt tolerance (1 to 10 % NaCl) ability test was also performed for the isolates. The highest tolerance to NaCl was observed in PTS-2, which is 10%, followed by GPS-4 (8%), HAS-3, GPL-1, GPL-2, and GPL-3 (7%). And all other isolates showed at least tolerance to 4% NaCl. Overall, all the isolates showed high tolerance to NaCl concentrations. The ability of the isolates to produce extracellular enzymes such as lipase, cellulase, amylase, protease, pectinase, and xylanase was also evaluated using a standard protocol. Protease production was demonstrated by the majority of isolates, with the exception of PTL-4 and HAS-3. PTL-4, PTR-5, GPL-3, HAL-1, HAL-2, and HAS-3 showed positive for amylase production. HAL-1, HAL-2, HAR-5, GPL-1, GPL-2, GPS- 4, PTL-4, and PTR-6 all showed positive lipase production. Positive activity for cellulase viii | P a g e 2025-02-01T00:00:00Z http://localhost:8080/xmlui/handle/123456789/502 Title: A study on in vitro propagation and comparative analysis of bioactive compounds in some selected wild and tissue-cultured medicinal plants of Bodoland Territorial Region, Assam” Authors: Baro, Tikendrajit Abstract: Abstract The primary source for food, shelter, and pharmaceutical products are primarily derived from plants, continuous consumption of plants for its medicinal and other uses leads to the extinction of the species from the wild. Plant tissue culture may be an alternate and ultimate solution to address this issue, in this technique any plant cell, tissue, or organ are used to grow in the culture media supplemented with plant growth regulators inside a controlled and aseptic environment. This technique is very important for mass propagation and rapid production of uniform, and disease-free plants; it is also very useful for the preservation of endemic plants and plant genome transformation. It is also very much effective tool for production of secondary metabolites and bioactive compounds from plants. Plant genome modification, plant improvement, transformation for vaccine production, and production of secondary metabolites are widely performed using this technique. In the present experiment five medicinal plants were selected from Tamulpur and Kokrakjar district of Assam, those includes- Torenia crustacea, Lindernia pusilla, Phlogacanthus thyrsiformis, Enydra fluctuans, and Hygrophila auriculata. This experiment aimed to standardize in vitro mass propagation technique, study of genetic stability in micropropagated plants using RAPD assay, comparison of antioxidant activity using different antioxidant assays, comparison of gallic acid and quercetin content in the wild and in vitro plant using HPLC analysis, and identification of bioactive constituents in wild and micropropagated plants using GC-MS analysis. The explant surface sterilization for in vitro propagation was conducted using 0.1% mercuric chloride for 1-5 min followed by incubation of the sterilized explants inside the growth chamber under a controlled and aseptic condition in the growth medium supplemented with growth regulators (BAP, IAA, NAA). During the entire process of tissue culture, the in vitro plant genome may undergo somaclonal variation due to the different in vitro conditions and growth regulators. For the detection of the somaclonal variation RAPD technique was conducted using 14 RAPD primers (OPC 01 to OPC 10, OPA1, OPA 02, OPA 12 and OPA 13).Different nutrient sources and in vitro conditions in the micropropagated plants may vary in the production of the secondary metabolites and active compounds from the wild plant. Therefore, the comparative antioxidant capacity experiments (total phenol content, total flavonoid content, total antioxidant capacity, and DPPH free radical scavenging activity) in wild and in vitro plants were conducted. Again, the quantitative gallic acid and quercetin content in the wild and in vitro plants were studied using HPLC analysis, and finally the bioactive compounds in the wild and in vitro plants were identified using GC- MS analysis. In the in vitro propagation experiment of T. crustacea, most effective explant surface sterilization was observed in 2 min treatment (63.33±4.71%) using 0.1% mercuric chloride, after 21 days of culture. In case of L. pusilla, 2 min and 3 min of treatment showed the maximum explant survival rate (76.67±4.71%) after 21 days of culture. In case of P. thyrsiformis, 2 min and 3 min treatment showed maximum survival rate (76.67±4.71%) of the explants after 21 days of explant culture, in case of E. fluctuans, 3 min and 4 min showed highest explant survival rate (76.67±4.71%) was observed after 28 days of the culture initiation, finally in case of H. auriculata, 4 min showed highest (56.67±4.71%) explant survival rate after 21 days of explant culture. Most effective shoot proliferation and multiplication of T. crustacea and P. thyrsiformis were observed in the MS media supplemented with 1mg/L BAP and 0.2 mg/L NAA and the highest average shoot length was observed and maximum average number of rooting was observed in MS media supplemented with 1mg/L IAA. In case of L. pusilla most effective shoot proliferation and multiplication were observed in the MS media supplemented with 1mg/L BAP and 0.2 mg/L NAA and the highest average shoot length was observed and maximum average number of rooting were observed in the MS media supplemented with 0.5mg/L IAA. Most efficient shoot multiplication of E. fluctuans explants were observed in the MS medium with 2 mg/L BAP (BM6), highest root formation was obtained in the MS medium supplemented with 1mg/L IAA. Finally in case of H. auriculata most effective shoot proliferation and multiplication were observed in the MS media supplemented with 1mg/L BAP and 0.5 mg/L NAA and the highest average shoot length was observed. Maximum average number of rooting were observed in the MS media supplemented with 1mg/L IAA. ixSomaclonal variation was observed in all the genome of in vitro propagated plant genomes from the RAPD experiment. From the comparative antioxidant study of wild and in vitro plants, the in vitro propagated plants showed higher antioxidant potential as compared to the wild plant. Also, from the comparative quantitative detection of gallic acid and quercetin in the wild and in vitro plant, the in vitro plant produced higher gallic acid and quercetin content than the wild plant. The active compounds in the wild and in vitro plants were identified using GC-MS analysis based on the peaks observed on the chromatograms. Diverse group of active compounds were identified with important characteristics including antimicrobial, antioxidant and antiviral activities were exhibited in the in vitro and wild plants. The study concludes that 0.1% mercuric chloride is effective for the explant surface sterilization of all the explants. Again, MS medium supplemented with higher concentration of BAP (1mg/l BAP) and lower concentration of NAA (0.2-0.5mg/L NAA) was most effective for the shoot multiplication and elongation, finally rooting was observed best in the MS medium supplemented with IAA (0.5mg/L to 1mg/L IAA). RAPD assay suggested that the presence of somaclonal variation in the in vitro plants. The antioxidant study suggested that the in vitro plants possessed higher antioxidant activity as compared to the wild plant and the comparative quantitative gallic acid and quercetin content analysis using HPLC suggested that the in vitro plants carried higher concentration of gallic acid and quercetin content as compared to the wild plant. In the screening of bioactive compounds some important bioactive compounds were identified in different medicinal plants, and the active components were varied in the in vitro plants than the wild plant. The somaclonal variation in the in vitro plants and the variation in the antioxidant properties and active compounds in the plant extracts may be caused due to the different in vitro conditions during the tissue culture process and use of different growth regulators in the culture medium. The performed research will help in commercial production of Lindernia crustacea, Lindernia pusilla, Phlogacanthus thyrsiformis, Enydra fluctuans, and Hygrophila auriculata. Production of secondary metabolites, and extraction of bioactive compounds form these plants are possible using the performed experiment. x 2026-02-01T00:00:00Z http://localhost:8080/xmlui/handle/123456789/381 Title: “MICROBIAL AND BIOCHEMICAL STUDY OF FERMENTED FISH NAPHAM TRADITIONALLY PREPARED BY BODO TRIBE OF KOKRAJHAR, ASSAM” Authors: Narzary, Yutika 2022-01-01T00:00:00Z http://localhost:8080/xmlui/handle/123456789/372 Title: “A Study on Growth Parameters, Genetic and Nutritional Characterization of Candidate Cordyceps spp.” Authors: BISWAS, VASHKAR Abstract: /media/bu_library/Vashkar Biswa/04_ abstract.pdf 2024-01-01T00:00:00Z