Development of diagnostic protocol for assessment of microbiological quality of fresh vegetables
| dc.contributor.advisor | Param Pal | |
| dc.contributor.author | Sood, Bhavish | |
| dc.date.accessioned | 2017-11-26T05:02:30Z | |
| dc.date.available | 2017-11-26T05:02:30Z | |
| dc.date.issued | 2017 | |
| dc.description.abstract | The epidemiological surveillance for microbiological quality of seven fresh vegetables (N=725) sourced from Village fields along Buddha Nallah, Supermarkets and Street vendors was conducted and concluded the high incidence of 11 important food borne pathogens with highest positive percentage of K. pneumonia (69%) in village fields, Y. enterocolitica in supermarkets (63.54%) and vendor samples (78.98%). The highest mean total plate count was observed in Spinach (>6 log cfu g-1) collected from village fields and vendors. Most probable number (MPN) analysis reflected the deteriorated quality of irrigation water with index range upto >1100/100ml (above desirable limit 1000/100ml). Pathogenic bacterial load was dominated in internal part of tomato (3.98 log cfu g-1) and on the surface region of cucumber (4.12 log cfu g-1). Phenotying of isolates of 11 pathogens was carried out to study the diversification of pathogens on basis of biochemical, virulence and antibiogram profiling. Clustering analysis of isolates (Unweighted Pair Group Method with Arithmetic Mean) revealed their significant diversity among themselves which provides insight of the indigenous pathogens inhabiting the vegetable commodity. Antibiotic profiling using 25 antibiotics belonging to 10 classes was established for all the isolates and complete tolerance to five antibiotics [Clindamycin (2mcg), Tetracycline (30mcg), Cloxacillin (5mcg), Metronidazole (5mcg) and Imipenem (10mcg)] was observed in Salmonella enterica. The bacterial isolates showed high MAR index range of 0.08-0.84. The species-specific virulence markers (enterotoxin, effector proteins encoding genes) used in PCR assay to detect the gene distribution pattern among the pathogens and their presence prove the potentiality of causing illness. A Bacteriological Food testing Kit (BFTK) (12h detection time) was devised based on the biochemical characteristics of indicator (Escherichia coli, Staphylococcus aureus, Shigella spp., Salmonella spp., Listeria monocytogenes) and emerging (Yersinia enterocolitica, Aeromonas hydrophila, Campylobacter jejuni, Bacillus cereus and Klebsiella pneumoniae) and validated with food samples (N=1000) against BIS Standard Methods at 5% level of significance. Another rapid yet inexpensive detection technique based on the Multiplex PCR was devised in which hblD gene (430bp fragment) of Bacillus cereus; ystA gene (79bp fragment) of Yersinia enterocolitica, invA gene (280bp fragment) of Salmonella enterica and iap gene in Listeria monocytogenes (225bp fragment) were targeted. Furthermore, a combined culturing (BFTK) and molecular tool (multiplex PCR) was found promising to detect pathogens in time slot of 15 h using specific markers with detection limit of 106 cells per target bacteria. A simulation study on concentration method stepped with vacuum filtration, low speed centrifugation and high speed centrifugation has shown high rate of recovery of all the 11 pathogens with sensitivity limit not less than 104 cfu/10g by reducing the sample to final volume of 2ml. A useful intervention was made for decontamination of vegetables by effectively using 1000ppm of citric acid in wash water. | en_US |
| dc.identifier.uri | http://krishikosh.egranth.ac.in/handle/1/5810036472 | |
| dc.keywords | diagnostic protocol, vegetables, tomato, cucumber, antibiotics, vacuum filtration, decontamination | en_US |
| dc.language.iso | en | en_US |
| dc.pages | 309 | en_US |
| dc.publisher | Punjab Agricultural University, Ludhiana | en_US |
| dc.research.problem | Development of diagnostic protocol for assessment of microbiological quality of fresh vegetables | en_US |
| dc.sub | Microbiology | en_US |
| dc.subject | null | en_US |
| dc.theme | Development of diagnostic protocol for assessment of microbiological quality of fresh vegetables | en_US |
| dc.these.type | Ph.D | en_US |
| dc.title | Development of diagnostic protocol for assessment of microbiological quality of fresh vegetables | en_US |
| dc.type | Thesis | en_US |