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The main objective of the present thesis was to investigate the impact of the dietary supplementation of the probiotic Enterococcus faecium (E. faecium) NCIMB 10415 and different zinc oxide (ZnO) concentrations on the Campylobacter (C.) coli burden in pigs (Chapter 3).
Chapter 2 provides a detailed literature review that emphasises the importance of pigs as a reservoir for Campylobacter spp. and thus, for possible human Campylobacter infections. Moreover, this chapter introduces possible pre-harvest intervention strategies to reduce the C. coli load in pigs for improved veterinary public health: Campylobacter spp. are a major cause of foodborne diarrhoeal disease in humans worldwide. Thermophilic Campylobacter spp. have their natural reservoir in warm-blooded animals with C. coli as the dominant species in pigs. The prevalence of C. coli in pig herds varies between 50 and 100% with excretion rates of up to 107 cfu/g faeces. Several studies showed that up to 10% of pork meat and pork products at retail are contaminated with Campylobacter spp. Moreover, C. coli accounts for 5–10% of the human Campylobacter infections. Thus, during the slaughter process, Campylobacter spp. can be transmitted to the carcass and subsequently to humans through the consumption of contaminated pork. Hence, there is an urgent need for proper intervention measures to reduce human C. coli infections. As Campylobacter spp. do not grow outside the host, a decreased contamination of Campylobacter spp. of the final product is best achieved if the colonisation in live animal can be prevented or reduced. The administration of antimicrobial feed additives as pre-harvest intervention measure are currently being intensively explored.
In the first step of the presented thesis, a Campylobacter spp. colonisation model in pigs was established. It enabled the examination of the C. coli colonisation of different sites within the pig gut which might serve as risk factors for the carcass contamination during slaughter (Chapter 4). Therefore, pigs were experimentally inoculated with the porcine C. coli 5981 strain. Then, the Campylobacter spp. excretion dynamics, the C. coli colonisation rates within different gut sections along the gastrointestinal tract and the translocation to extragastrointestinal sites was studied. Based on this colonisation model, the impact of the dietary supplementation of different ZnO concentrations and the probiotic E. faecium NCIMB 10415 to reduce the faecal C. coli load was tested in weaned piglets (Chapter 5 & 6). The diet of weaned piglets was supplemented with three levels of ZnO (40, 100 and 3,100 mg ZnO/kg feed) and with two levels of E. faecium NCIMB 10415 (0 and 109 cfu/kg feed). In both feeding trials, all piglets were naturally colonised with C. coli with excretion rates of 103–108 cfu/g faeces. The results of the colonisation trial revealed that the entire gastrointestinal tract, but also gut-associated lymphatic tissues constitute big reservoirs for C. coli and serve as potential reservoirs for the carcass cross-contamination during slaughter. Furthermore, we were able to demonstrate that only the dietary supplementation of 3,100 mg ZnO/kg feed, but not E. faecium, led to significant reduced faecal C. coli counts in pigs. Supplementation of the postweaning diet with this pharmacological dose significantly reduced the C. coli excretion level by 10-fold within one week (p = 0.001) compared to piglets that received a low and medium ZnO diet. The reduction of C. coli excretion by high ZnO has been shown to be a transient phenomenon and only lasts as long as the ZnO feeding remains high. Reducing the pharmacological concentration of 3,100 mg ZnO/kg feed to the physiological requirement of 100 mg ZnO/kg feed led to increased C. coli numbers within a few days again. A 10-fold increased zinc concentration in the faeces of pigs that were dietary supplemented with 3.100 mg ZnO/kg feed was measured compared to the control groups. Despite the negative impact on the C. coli growth in pigs fed a high ZnO concentration, several side effects must be considered: The possible elevated zinc concentration in meat and the increased zinc excretion into the environment.
By in vitro growth inhibition assays a high susceptibility of C. coli against ZnO could also be observed. At concentrations ≥ 2.6 mM ZnO a decline in cell numbers occurred. It turned out that the antimicrobial activity is caused, at least partly, by the production of reactive oxygen species. The expression of catalase A, the main peroxide stress defence protein in Campylobacter spp. was up-regulated by 5-fold after ZnO treatment. Besides the activation of the oxidative stress response, the general- and heat stress response was also triggered. The expression of the chaperones clpB, groES and dnaK was up-regulated after ZnO exposure by 2.2-, 2.5- and 2.6-fold, respectively. In contrast, the expression of the zinc influx transporter znuABC and zinc efflux transporter zntA were almost unaffected after ZnO stress.
In conclusion, supplementation of high dietary ZnO, but not E. faecium NCIMB 10415 significantly reduced the C. coli load in weaned piglets. A short-term dietary administration of 3,100 mg ZnO/kg feed to pigs before slaughter seems to be a promising intervention measure to reduce the C. coli load. This reduction might further lead to reduced Campylobacter spp. contamination rates of pork and subsequently to a reduced incidence of C. coli.