Avnish Kumar, Mohsin Shawl, Rashmi Jain and Abhishek Mathur
Endophytes are the microbes that live within the host plant tissues without causing any visible disease symptoms. Depending on their nutritional requirements they can live as biotrophic parasites or saprotrophs. They also represent a huge reservoir of microbes that are explored very poorly. The present study was done first ever in India to investigate the biodiversity of endophytic bacteria and fungi in parts of the plant, Murraya koenigii. The results have shown that the both bacterial and fungal isolates were diverse both in morphology and characteristics. In the present investigation, the results were found to be very surprising and interesting. Total of 08 bacterial and 11 fungal endophytes were isolated from the plant parts. It was found that the inner core of leaves, stems and roots of Murraya koenigii L. were found to have the presence of bacterial and fungal endophytes as observed on LB and PDA plates. Different colonial growth was observed in the petriplates while some were obtained as mixed cultures. The pure cultures were maintained separately for further use. In case of bacterial endophytes as observed on LB agar plates. 3 isolates were obtained from stem, 3 were obtained from leaves and 2 were obtained from roots. In case of fungal endophytes as observed on PDA plates, 6 isolates were obtained from stem, 4 were obtained from leaves and single isolate was obtained from roots. It was found that bacterial endophytic fractions were most effective against the pathogens and drug resistant strains in comparison to fungal endophytic fractions. The results showed that bacterial endophytic fractions of stems viz. S1, S2 and S3 possessed significant antimicrobial activity against E. coli NCIM 2065, Lactobacillus plantarum NCIM 2083, Micrococcus luteus ATCC 9341, Salmonella abony NCIM 2257 while no activity was observed against Methicillin resistant strains of S. aureus and any of the fungal cultures, Candida albicans NCIM 3471; Aspergillus niger NCIM 1196. Amongst bacterial endophytic fractions of leaves, L1 and L3 showed no antibacterial activity against any of the bacterial pathogens while showed potency against fungal cultures studied. Only L2 bacterial endophytic fraction showed potency against all the bacterial and fungal pathogens studied except Lactobacillus plantarum NCIM 2083. The L2 bacterial endophytic fraction also showed antibacterial activity against one of the Methicillin resistant strain of S. aureus. Bacterial endophytic fractions of roots viz. R1 and R2 showed potency against all the bacterial and fungal cultures except E. coli NCIM 2065. No antimicrobial potential of any of the fungal endophytic fraction of stem viz. S1-S6 against any of the test organism was revealed. Although S3 fraction showed slight antibacterial activity against one of the Methicillin resistant strain of S. aureus. Simultaneously fungal endophytic fraction of leaves viz. L1, L3 and L4 also showed no activity against any of the test organism. L4 fraction showed significant antifungal activity against Candida albicans NCIM 3471. The endophytic fraction of leaves, L2 showed significant activity against one of the Methicillin resistant strain of S. aureus, E. coli NCIM 2065 and Lactobacillus plantarum NCIM 2083. The isolated bacterial endophytes were screened for gram staining and different biochemical tests. The results confirmed that out of 08 isolated cultures of bacterial endophytes, 05 were gram positive, non-motile, non-spore forming bacilli and 03 were gram negative, motile, non spore forming bacilli. Gram positive colonies showed positive sugar (Glucose, sucrose, fructose, rhamnose, mannose) fermentation test, catalase test, amylase test, urease test, esterase test and methyl red test while Gelatin liquefication test, Indole test, Vogues Prosker test, Citrate utilization test, H2S production tests were found to be negative. Gram negative colonies showed positive sugar (Glucose, sucrose, lactose, cellobiose) fermentation test, Mac Conkey growth, Vogues Prosker test, Citrate utilization, Indole and acetate utilization test. The data of the tests were evaluated by the previous studies which confirm gram positive bacilli as Bacillus megaterium while gram negative bacilli were nominated as Enterobacter cloacae. The fungal endophytes isolates were classified by colony and hyphal characters as stained by lactophenol. Cultures were deposited at National Centre of Fungal Taxonomy (NCFT), New Delhi. The results confirmed that amongst, 11 fungal endophytes isolates, 05 were found to be Aspergillus niger, 02 were Aspergillus flavus, 02 were Candida albicans, 01 was Phoma hedericola and 01 was Penicillium sublateritium.
