Borah, ProbodhPATHAK, BEDANTA2018-11-192018-11-192017-07http://krishikosh.egranth.ac.in/handle/1/5810083621The present study was undertaken with a view to isolate bifidobacteria from various sources, viz. fresh bovine milk, faeces of calves and piglets, ruminal content of slaughtered bovines and intestinal/caecal content of chickenand to characterize the isolates by random amplified polymorphic DNA (RAPD) analysis and sequencing of amplified 16S rRNA gene fragments. A total of 12 (2.52%)out of477 samples from various sources examined were found positive forBifidobacterium spp. yielding the same number of isolates. All of these isolates were recovered from faecal samples of young piglets (6.12%), while samples from other sources examined did not reveal the presence of bifidobacteria. The isolates were characterized and identified by biochemical tests and phosphoketolase (F6PPK) enzyme assay, which showed characteristic features of Bifidobacterium spp. All the 12 isolates were also confirmed by genus-specific PCR based on amplification of 16S rRNA gene. Sequencing of the amplified products and subsequent sequence analysis by NCBI-BLAST revealed that out of the 12 isolates, 3 (25%) belonged to B. breve, 1 (8.33%) to B. pseudolongum, 6 (50%) to B. animalis subsp. lactis and 1 (8.33%) to B. thermacidophilum subsp. porcinum. However in respect of one isolate, the species could not be ascertained by 16S rRNA sequencing due to poor sequence data, although it was confirmed as belonging to Bifidobacterium spp. by genus-specific PCR. RAPD analysis could not clearly differentiate the strains belonging to different species. Phylogenetic analysis based on the bandings patterns also failed to differentiate the isolates belonging to different species of Bifidobacterium. In-vitro assessment of their antibacterial effect of the isolates revealed that two isolates ofB. animalis subsp.lactishadhigher inhibitory activity compared to the other isolates. Based on higher inhibitory effect on selected pathogens, five isolates were selected for further characterization in respect of their probiotic potential in vitro.The inhibitory effect of all these five isolateson the selected pathogens persisted even after heat-treatment at 100°C for 5 min. However, the inhibitory effect didn’t persist after treatment with pronase-E and proteinase-K. All the five isolates tested were able to grow at a minimal pH at 4.0. Two of the tested isolates belonging toB. animalis subsp.lactisspecies exhibited comparatively higher acid tolerance even after 180 min of exposure and showed the lowest percentage of mortality of 16.21 and 21.79 per cent, and 13.40 and 17.32 per cent, respectively on exposure to lysozyme (0.5 mg/ml) and ox bile salts @ 0.3% (w/v). In antibiotic resistant test, all the five isolates showed almost uniform susceptibility to different antimicrobial agents tested, except tetracycline. The present studysuggestedthat the two isolates from faeces of piglets obtained in the present studyof B. animalis subsp.lactispossess promising probiotic potentialand further in vivo study is required to be donefor validation of their probiotic activity.ennullThesis