Veterinary Science - Theses
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ItemThe impact of pharmacokinetics on the emergence of in vitro bacterial resistance to cefovecin( 2017)Introduction Antibiotic resistance is a global issue in both human and veterinary medicine with substantial social and economic impact. Cefovecin, a third-generation cephalosporin antibiotic has a half-life of approximately 166 hours in the cat. This contrasts with the significantly shorter two-hour half-life of cephalexin, an oral first-generation cephalosporin. Sub-inhibitory antibiotic cultures were used to investigate the impact of cephalosporin antimicrobial concentration and time on the development of in vitro bacterial resistance. It was hypothesized that the long half-life of cefovecin would cause an increased emergence of bacterial resistance. Materials & Method Five clinical isolates of Escherichia coli from the U-Vet Werribee Animal Hospital laboratory collection were subjected to sub-inhibitory conditions. Escherichia coli National Collection of Type Cultures England (NCTC) 10418 and NCTC 12441 (American Type Culture Collection (ATCC) 25922) reference strains were used as controls. Minimum inhibitory concentrations (MIC) of cefovecin and cephalexin were determined by broth microdilution techniques as described by the Clinical and Laboratory Standards Institute (CLSI) guidelines. Antibiotic concentrations of 0 - 0.95 x MIC in Mueller-Hinton cation adjusted broth (CAMHB) were used as sub-inhibitory culture conditions. The batch-cultures were incubated on an orbital shaking platform at 37 ◦C for a total period of 144 hours. The bacteria were diluted 1:1000 into fresh antibiotic media every 24 hours. Following this, the E. coli isolates from the 0.95 x MIC cefovecin sub-inhibitory experiments were cultured in antibiotic-free media for a further 144 hours to test the persistence of any increase in resistance. Results E.coli isolates cultured in antibiotic-free CAMHB for 24 hours showed no significant change in MIC to either drug, however E. coli isolates cultured in sub-inhibitory antibiotic concentrations for periods of up to 144 hours showed up to 16 times increase in MIC for cefovecin and up to 8 times increase in MIC for cephalexin. Cultures with antibiotic concentrations closer to the MIC of the E. coli isolate increased their MICs both more rapidly and to greater multiples. The 0.95 x MIC cefovecin E. coli isolates that were cultured in antibiotic-free media for a further 144 hours showed either a 2-fold or 4-fold decrease in MIC, although none returned to the initial MIC. This research provides preliminary evidence that cefovecin should be used with prudence to limit in vivo resistance emergence.
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ItemBacterial enteritis in young Merino sheep.( 2017)Diarrhoea during winter is a common problem of sheep in the high rainfall regions of south-eastern Australia. Gastrointestinal nematodes are the primary cause of this problem, especially in young Merino sheep (those <16 months old). However, a syndrome of persistent diarrhoea (‘winter scouring’) is also relatively common in young sheep with low or moderate nematode burdens, with outbreaks usually occurring between June and October. Affected animals have profuse, watery diarrhoea, and often show signs of abdominal discomfort. Before this study, the cause of this syndrome and the epidemiology and risk factors for outbreaks and faecal shedding of the causative organisms, were not well understood. To determine the primary cause of the syndrome, 45 outbreaks of ‘winter scouring’ were investigated on 24 farms across south-eastern Australia. Necropsies were conducted and faecal samples were collected from animals in affected flocks. Faecal and tissue samples were cultured for Yersinia, Salmonella and Campylobacter species, and faecal WECs conducted to assess nematode parasitism. The sensitivity of Yersinia isolates to four common antimicrobial compounds was also assessed. Yersinia pseudotuberculosis serotype III and virulent Y. enterocolitica were the most frequently isolated bacteria, with outbreaks typically occurring when more than 10% of the mob were shedding either of these organisms. Campylobacter and Salmonella did not play a role in any of the outbreaks investigated. Over 64% of all Yersinia isolates were resistant to sulphonamides, but 98% were susceptible to oxytetracycline. These results highlighted the need to investigate the epidemiology and risk factors of yersiniosis in more detail to support the development of more effective control strategies. As a first stage of this process, a longitudinal study was conducted on four farms to evaluate the patterns of faecal shedding of Yersinia and assess some potential risk factors for outbreaks of yersiniosis, including live-weight, average daily weight gain and WEC. Faecal shedding of both species occurred predominantly during the winter, but shedding of Y. enterocolitica also occurred at other times. Therefore, in addition to scouring in winter, Y. enterocolitica can also cause diarrhoea at other times. In contrast to the outbreak investigations, the prevalence of faecal shedding of Yersinia species was often very high (>30% of animals within a mob) without any clinical disease. This suggests that complex interactions between risk factors contribute to yersiniosis, and that simply monitoring the prevalence of faecal shedding of Yersinia species will not be a reliable predictor of an outbreak of yersiniosis. High WECs (>300 eggs per gram) were associated with an increased risk of shedding each Yersinia species, and so effectively controlling gastrointestinal nematodes should also decrease the risk of yersiniosis. Animals that gained weight were less likely to shed Y. enterocolitica, but more likely to shed Y. pseudotuberculosis. Despite this, the current strategy of ensuring weaner sheep (those 3-12 month of age) continue to gain weight during autumn and winter is still recommended. The effect of an autogenous bacterin against Y. pseudotuberculosis serotype III was assessed on two farms that consistently had severe outbreaks of yersiniosis. There was no outbreak in the year of the study (2014), and vaccinated weaners had similar live weight, growth, faecal shedding of both Yersinia spp. and antibody titres to Yersinia outer membrane proteins to unvaccinated controls. Consequently, further studies to evaluate the efficacy of vaccines during an outbreak of yersiniosis are required.