Molecular Analysis of Microbial Ecology in Poultry: Campylobacter, Salmonella and Gut Microbiota.

摘要

Conventional strain typing methods have been readily taken over by newer molecular techniques based on nucleic acid typing methods owing to their ease of use, reproducibility, sensitivity and speed. In this study, we have made use of these techniques, which find their implication in assessing ecology and dynamics of microbial community, for which the dissertation has been divided into two sections. In the first section, we have focused on the ecology of food borne pathogens, Campylobacter and Salmonella, which are the leading cause of food borne illnesses around the world. These pathogens are responsible for huge loss not only in the poultry industry but also, causing major illnesses in humans in turn affecting the health care industry. Molecular typing methods were utilized for accomplishing these goals, where we used fla gene typing for understanding the colonization patterns within individual broiler chickens, which reflected on the diversity of Campylobacter population within each broiler. Further, the growth and development of poultry birds are influenced by the gastrointestinal tract microbiota and attempts made to study this complex microbial community in order to understand its interaction with the host has provided an insight on improving poultry health. In this study, we aimed at comprehending such interactions within the chicken gut under the influence of antibiotics, when used as growth promoters. Our results, of metagenomic study based on 16S rRNA gene sequence analysis, indicate that the composition of bacterial phyla in the microbiota is drastically altered which in turn affects the body weight of the host. This alteration in the gut microbial composition can be utilized towards development of alternative growth promoters, and help in evading the development of antibiotic resistance in the chickens as well as its consumers. In the second section, we have developed a novel typing method, multilocus sequence typing (MLST)-seq, to characterize and differentiate food borne pathogens in a simple and robust way by developing two target enrichment strategies 1) Hairpin Selector based ligation and 2) Hairpin-primed Multiplex amplification, for selectively amplifying target genes taking advantage of the Next Generation pyrosequencing technologies. With Salmonella as a model organism each technique has successfully amplified 21 target genes simultaneously from Salmonella genome that were further analyzed for typing Salmonella. MLST-seq has demonstrated a higher discriminatory power than that of existing typing techniques, MLST and pulse-field gel electrophoresis.