The occurrence of shiga-toxin producing Escherichia coli in South African game species

[摘要] ENGLISH ABSTRACT: The consumption of game meat on a global scale is increasing every year and South Africa is noexception. Consumers have started incorporating low kilojoule and low cholesterol products intotheir lifestyles, of which game meat is one such product. These increases give rise to a range ofchallenges for the game meat industry in terms of the microbial safety of the meat.The aims of this study were to optimise a DNA extraction protocol for the isolation of theShiga-toxin producing Escherichia coli (STEC) positive control and PCR conditions for theamplification of the stx1, stx2 and eaeA virulence genes; to determine the prevalence of STECcontamination in South African game species; to determine the microbial population present onSouth African game carcasses after dressing; and to develop and implement an organic acid sprayin order to reduce microbial growth on game carcasses.Optimisation of a DNA extraction protocol for STEC positive control included an ethanolwash and re-suspension in sterile distilled water to purify the DNA from any PCR inhibitors. Primerconcentrations were also optimised to prevent non-specific binding, which causes primer-dimerformation. PCR amplification conditions were compared to determine the optimum annealingtemperature, as a too low or too high temperature led to unsatisfactory amplification results.The optimised protocol and conditions were used to determine the prevalence of STEC inSouth African game species. Animals from two different farms were used in this study. Farm 1consisted of four species, of which two species (Zebra and Black Wildebeest) had STEC present inthe faecal matter. The meat of all four species (Zebra, Black Wildebeest, Impala and Eland)contained STEC suggesting possible cross contamination from the hide to the carcass. Faecalsamples from Farm 2 (Blesbok and Springbok) tested negative for STEC.During the microbial population study it was found that aerobic bacteria prevalence on themeat ranged from 1.60 to 4.97 log cfu·cm-2 whereas total coliforms varied from 5.04 to 5.59 logcfu·cm-2. E. coli prevalence ranged from 0.00 to 1.71 log cfu·cm-2 while Staphylococcus aureusvaried from 0.00 to 2.97 log cfu·cm-2. The results from the faecal matter displayed aerobic bacteriaprevalence in the range of 5.78 to 6.44 log cfu·g-1 while total coliforms ranged from 6.53 to 7.04 logcfu·g-1. E. coli prevalence varied from 3.00 – 4.54 log cfu·g-1 while Staphylococcus aureus rangedfrom 3.63 to 4.40 log cfu·g-1. None of the faecal samples tested positive for Salmonella or Listeria.Throughout the organic acid spray development, different organic acids (lactic acid, citricacid, acetic acid and octanoic acid) and sodium benzoate were tested singly and in combination.Development started with enrichment mediums and progressed to meat samples. Based on theresults of the meat samples a final combination of 5% lactic acid, 0.5% octanoic acid and 0.1%sodium benzoate was chosen to use on game carcasses. Although the combination provided promising results in laboratory trials, its efficacy on the carcasses was unsatisfactory with lowoverall log reductions achieved.To conclude, the high prevalence of STEC (26.9%) detected in the game species wasalarming and requires further investigation.

[发布机构] Stellenbosch University