Profiles, Diversity and Antibiotic Response Patterns of Bacterial Isolates from Fish and Processed Fish Products Retailed in Kirinyaga County, Kenya
Kyule, Domitila Muendo
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Fish contain omega 3 polyunsaturated fatty acids (PUFA’s), vitamins and other micronutrients. However, fish and seafood rank among foods highly associated with food borne disease outbreaks especially in poor hygienic conditions while handling or processing foods. The objectives of this study were to determine the risk factors associated with fish contamination, levels of microbial contamination and molecular profiles of bacterial isolates. In addition, antimicrobial response patterns were also determined against commonly used antibiotics. A semi-structured questionnaire was administered randomly to individual fish handlers while fish products (raw fish, samosa, sausage, cake, skewer, hot dog, fried fish, fish balls, fingers and burger) were randomly sampled from fish vendors. Microbial load was determined using serial dilutions and the load determined by pour plates. The bacteria were enumerated and sub-cultured to obtain pure cultures on Nutrient media. Bacterial morphologies were determined and confirmed by biochemical characterization using citrate utilization, methyl red, SIM, urease, indole and carbohydrate fermentation tests. Total genomic bacterial DNA was extracted using boiling extraction method and 16S rRNA gene was amplified using PCR while the amplifications were confirmed using gel electrophoresis. The amplified products were then directly sequenced using Sanger method in both directions using the same primers as in the PCR amplification. Kirby-Bauer agar diffusion method was used to test antibiotic susceptibility test. Data was analyzed using ANOVA and means were separated using Tukey’s HSD test at 5 % significance level (SAS version 9.0). The generated sequences of 16S rRNA gene were analyzed and edited using Finch-Tv software Ver. 1.4.0. Data analysis was carried out using SAS software Ver. 9.4. There was a significant difference (p = 0.001) and interaction (p = 0.037) of average bacteria count obtained from different fish products and markets. Fish treatment resulted to a significant difference (p = 0.001) total CFUs. Based on morphological characteristics, a total of 64 morpho-groups were obtained. Based on molecular characterization, most of the identified bacterial isolates belonged to Bacillus sp. Other identified bacterial isolates comprised of B2SmCTM1, which was identified as Klebsiella michiganensis, isolate C1FCTM1 identified as Salmonella enterica, isolates F3WTRM5, M10SmCTM1 and M10SaCRM5 identified as Escherichia coli. In addition, isolates J6BCTM5, L10WTTM5 and J6WTTM5 were concluded to be Staphylococcus xylosus. The study revealed that respondents were aware that high temperatures promoted bacterial growth thus majority (68 %) refrigerated their fish products. The storage of fish products in display units and shelves without ice may justify high levels of contamination experienced in the region. Fish and fish products sold in various markets in Kirinyaga county-exhibited varied bacterial contamination with fish samosas having the highest bacterial count. There was a significant difference in the action of tetracycline, streptomycin, ampicillin, ciprofloxacin, gentamycin and penicillin against the bacterial isolates (P = 0.001). Ciprofloxacin was the most effective antibiotic based on antimicrobial response patterns of the selected bacterial isolates while penicillin was the least effective antibiotic. Extensive empowerment of the fish handlers and hoteliers on the proper handling of the raw fish and fish products to minimize the instances of microbial contamination should be promoted.