Mohan Chandra Kalita

Fatty acid profiling and molecular characterization of some freshwater microalgae from India with potential for biodiesel production.

We determined the fatty acid compositions of six species of freshwater microalgae belonging to the Chlorophyta, which were isolated from freshwater bodies in Assam, India. All six microalgae -Desmodesmus sp. DRLMA7, Desmodesmus elegans DRLMA13, Scenedesmus sp. DRLMA5, Scenedesmus sp. DRLMA9 Chlorella sp. DRLMA3 and Chlorococcum macrostigmatum DRLMA12-showed similar fatty acid profiles 16:0, 16:4, 18:1, 18:2, and 18:3 as major components. We also compared fatty acid compositions during the late exponential and stationary growth phases of D. elegans DRLMA13 and Scenedesmus sp. DRLMA9 in BG11 medium. We observed enhanced percentages of total saturated and monounsaturated fatty acids with a concomitant decrease in polyunsaturated fatty acid content upon the prolonged cultivation of both microalgae. Distinct morphological features of microalgal isolates were determined by scanning electron microscopic (SEM) studies. An ornamented cell wall was found in D. elegans DRLMA13, which is characteristic of small spineless species of Desmodesmus. The isolated microalgae were further distinguished through analysis of internal transcribed spacer 2 (ITS2) secondary structures and compensatory base changes (CBCs). Analysis of CBCs showed the relatedness of Chlorella sp. DRLMA3 with other Chlorella-like organisms, but it does not belong to the clade comprising Chlorella sensu stricto, which includes Chlorella vulgaris. The CBC count between Scenedesmus sp. DRLMA9 and other species of Scenedesmus provides evidence that this isolate represents a new species.

In Vitro Clonal Propagation of Curcuma caesia Roxb and Curcuma zedoaria Rosc from Rhizome Bud Explants

Two species of Curcuma (C. caesia and C. zedoaria) have been propagated through tissue culture using rhizome bud explant. The best response for shoot multiplication was obtained on MS basal medium supplemented with 4 mg l−1 BAP and 1.5 mg l−1 NAA for C. caesia (3.5 ± 0.79 shoots per explant) and 1 mg l−1 BAP + 0.5 mg l−1 NAA for C. zedoaria (4.5 ± 0.15 shoots per explant). A maximum of 9.2 ± 0.15 and 8.9 ± 0.09 roots per explant were obtained for C. caesia and C. zedoaria, respectively when MS was supplemented with 0.5 mg l−1 IAA. The rooted plants could be established in soil.

Identification of drought tolerant progenies in tea by gene expression analysis.

Understanding the genes that govern tea plant (Camellia sinensis) architecture and response to drought stress is urgently needed to enhance breeding in tea with improved water use efficiency. Field drought is a slow mechanism and the plants go through an adaptive process in contrast to the drastic changes of rapid dehydration in case of controlled experiments. We identified a set of drought responsive genes under controlled condition using SSH, and validated the identified genes and their pattern of expression under field drought condition. The study was at three stages of water deficit stress viz., before wilting, wilting and recovery, which revealed a set of genes with higher expression at before wilting stage including dehydrin, abscissic acid ripening protein, glutathione peroxidase, cinnamoyl CoA reductase, calmodulin binding protein. The higher expression of these genes was related with increase tolerance character of DT/TS-463 before wilting, these five tolerant progenies could withstand drought stress and thus are candidates for breeding. We observed that physiological parameter like water use efficiency formed a close group with genes such as calmodulin related, DRM3, hexose transporter, hydrogen peroxide induced protein, ACC oxidase, lipase, ethylene responsive transcription factor and diaminopimelate decarboxylase, during wilting point. Our data provides valuable information for the gene components and the dynamics of gene expression in second and third leaf against drought stress in tea, which could be regarded as candidate targets potentially associated with drought tolerance. We propose that the identified five tolerant progenies on the basis of their drought tolerance can thus be utilised for future breeding programmes.

Development of an Efficient Bacterial Consortium for the Potential Remediation of Hydrocarbons from Contaminated Sites.

The intrinsic biodegradability of hydrocarbons and the distribution of proficient degrading microorganisms in the environment are very crucial for the implementation of bioremediation practices. Among others, one of the most favorable methods that can enhance the effectiveness of bioremediation of hydrocarbon-contaminated environment is the application of biosurfactant producing microbes. In the present study, the biodegradation capacities of native bacterial consortia toward total petroleum hydrocarbons (TPH) with special emphasis to poly aromatic hydrocarbons were determined. The purpose of the study was to isolate TPH degrading bacterial strains from various petroleum contaminated soil of Assam, India and develop a robust bacterial consortium for bioremediation of crude oil of this native land. From a total of 23 bacterial isolates obtained from three different hydrocarbons contaminated samples five isolates, namely KS2, PG1, PG5, R1, and R2 were selected as efficient crude oil degraders with respect to their growth on crude oil enriched samples. Isolates KS2, PG1, and R2 are biosurfactant producers and PG5, R1 are non-producers. Fourteen different consortia were designed involving both biosurfactant producing and non-producing isolates. Consortium 10, which comprises two Bacillus strains namely, Bacillus pumilus KS2 and B. cereus R2 (identified by 16s rRNA sequencing) has shown the best result in the desired degradation of crude oil. The consortium showed degradation up to 84.15% of TPH after 5 weeks of incubation, as revealed from gravimetric analysis. FTIR (Fourier transform infrared) and GCMS (Gas chromatography-mass spectrometer) analyses were correlated with gravimetric data which reveals that the consortium has removed a wide range of petroleum hydrocarbons in comparison with abiotic control including different aliphatic and aromatic hydrocarbons.

Characterization of the biofuel potential of a newly isolated strain of the microalga Botryococcus braunii Kützing from Assam, India.

Botryococcus braunii GUBIOTJTBB1 was isolated from a freshwater reservoir in Assam, India and its taxonomic identity was confirmed by 18S rRNA sequence analysis. Biofuel potential of the microalga strain was assessed from batch culture under laboratory conditions, based on its lipid content and energy value of the dried biomass. Total lipid of 57.14% and hexane extractable crude hydrocarbon of 52.6% were recorded maximum at 56 and 28 days respectively, which vary upon culture durations. The energy value (54.69 kJ/g) of the strains sundried biomass was found higher than that of petroleum diesel fuel and nearly twice than other microalgae strains compared. The strain GUBIOTJTBB1 was found superior in terms of total lipid and hydrocarbon contents comparing to the previously reported Indian strains of B. braunii. With further improvements in growth, the strain could become an ideal feedstock for potential biofuel production in the prevailing climatic conditions of the region.

Broad Spectrum Antimicrobial Activity of Forest-Derived Soil Actinomycete, Nocardia sp. PB-52

A mesophilic actinomycete strain designated as PB-52 was isolated from soil samples of Pobitora Wildlife Sanctuary of Assam, India. Based on phenotypic and molecular characteristics, the strain was identified as Nocardia sp. which shares 99.7% sequence similarity with Nocardia niigatensis IFM 0330 (NR_112195). The strain is a Gram-positive filamentous bacterium with rugose spore surface which exhibited a wide range of antimicrobial activity against Gram-positive bacteria including methicillin-resistant Staphylococcus aureus (MRSA), Gram-negative bacteria, and yeasts. Optimization for the growth and antimicrobial activity of the strain PB-52 was carried out in batch culture under shaking condition. The optimum growth and antimicrobial potential of the strain were recorded in GLM medium at 28°C, initial pH 7.4 of the medium and incubation period of 8 days. Based on polyketide synthases (PKS) and non-ribosomal peptide synthetases (NRPS) gene-targeted PCR amplification, the occurrence of both of these biosynthetic pathways was detected which might be involved in the production of antimicrobial compounds in PB-52. Extract of the fermented broth culture of PB-52 was prepared with organic solvent extraction method using ethyl acetate. The ethyl acetate extract of PB-52 (EA-PB-52) showed lowest minimum inhibitory concentration (MIC) against S. aureus MTCC 96 (0.975 μg/mL) whereas highest was recorded against Klebsiella pneumoniae ATCC 13883 (62.5 μg/mL). Scanning electron microscopy (SEM) revealed that treatment of the test microorganisms with EA-PB-52 destroyed the targeted cells with prominent loss of cell shape and integrity. In order to determine the constituents responsible for its antimicrobial activity, EA-PB-52 was subjected to chemical analysis using gas chromatography-mass spectrometry (GC-MS). GC-MS analysis showed the presence of twelve different chemical constituents in the extract, some of which are reported to possess diverse biological activity. These results confirmed that the presence of bioactive constituents in EA-PB-52 could be a promising source for the development of potent antimicrobial agents effective against wide range of microbial pathogens including MRSA.

In vitro Clonal Propagation and Organogenesis in Spilanthes acmella (L) Murray: A Herbal Pesticidal Plant of North-East India

Multiple shoots were regenerated in MS medium using different concentrations of BAP and Kn and different combinations of BAP with IAA, NAA and IBA. Highest multiplication of shoots was obtained with BAP (0.75 mg l−1) with 28.4 shoots per explant after 60 days of culture. Shoots rooted best on IBA (0.5 mg l−1), numbering 48.8 per explant. Organogenesis was maximum in callus cultured on MS medium supplemented with BAP (2.0 mg l−1) and IAA (1.0 mg l−1).

Foliar Spray with Vermiwash Modifies the Arbuscular Mycorrhizal Dependency and Nutrient Stoichiometry of Bhut Jolokia (Capsicum assamicum)

Vermiwash (VW), a liquid extract obtained from vermicomposting beds, is used as an organic fertilizer for crop plants. The current study investigated the effect of a vermiwash foliar spray on the response of bhut jolokia (Capsicum assamicum) exposed to two different arbuscular mycorrhizal fungi (AMF: Rhizophagus irregularis, RI and G. mosseae, GM) in acidic soil under naturally ventilated greenhouse conditions. The VW spray significantly influenced the growth of plants receiving the dual treatment of AMF+VW. Plant growth was more prominent in the GM+VW treatment group than that in the RI+VW treatment group. The plant-AMF interactions in relation to growth and nutrient requirements were also significantly influenced by the application of VW. Interestingly, the VW treatment appeared to contribute more N to plants when compared to that under the AMF treatment, which led to changes in the C:N:P stoichiometry in plant shoots. Furthermore, the increased potassium dependency, as observed in the case of the dual treatments, suggests the significance of such treatments for improving crop conditions under salt stress. Overall, our study shows that the VW foliar spray modifies the response of a crop to inoculations of different AMF with regard to growth and nutrient utilization, which has implications for the selection of an efficient combination of nutrient source for improving crop growth.

Computational Identification, Target Prediction, and Validation of Conserved miRNAs in Insulin Plant (Costus pictus D. Don)

Insulin plant (Costus pictus D. Don) is an economically important medicinal plant for the content of its high value secondary metabolites, bioactive compounds, and remarkable flowering features. MicroRNAs are a class of short (∼21 nucleotides), endogenous, noncoding RNA molecules that play a vital role in regulating gene expression. Here, we used a computer-based homology approach to identify conserved miRNAs in Transcribed Sequence Assemblies (TSA) of C. pictus. It led us to identify 42 miRNAs of 13 different families in C. pictus for the first time. Using quantitative polymerase chain reaction (qPCR) assays, we further confirmed the expression of 8 miRNAs (miR394, miR159b, miR166k, miR172, miR159f, miR166, miR144, and miR858) in young and mature leaf tissues. A total of 109 potential target genes of the identified miRNAs were subsequently predicted in rice (Oryza sativa L.) genome. The target genes encode transcription factors, enzymes, and various functional proteins involved in the regulation of several metabolic pathways. The findings in the present study lay the foundation for further research on miRNAs and miRNA-mediated gene regulation in this important medicinal plant.

HPLC analysis of harringtonine and homoharringtonine in the needles of Cephalotaxus griffithii alkaloid fraction and cytotoxic activity on chronic myelogenous leukaemia K562 cell

Harringtonine (HT) and homoharringtonine (HHT) are Cephalotaxus alkaloids with considerable antileukaemic activity. The objectives of this research were to (1) determine the content of HT and HHT present in Cephalotaxus griffithii needles alkaloid fraction (CGAF) and (2) compare the antiproliferative activity of CGAF, with that of HT and HHT on chronic myelogenous leukaemia K562 cell. The concentration of HT and HHT was found to be 122.14 and 16.79 mg/g of CGAF, respectively. Treatment of K562 cells with CGAF, HT and HHT decreased the viable cells in a dose- and time-dependent manner. Interestingly, the maximum cell death was found in CGAF, with IC50 value which was 3- to 4.6-fold lower than those of HT and HHT. Our results indicate that HT content in the needles of C. griffithii is higher than HHT, and alkaloids other than HT and HHT in CGAF are predominantly responsible for K562 cell death.