Gut microbiome and immune system changes in heat-stressed pigs : A thesis submitted in partial fulfilment of the requirements for the Degree of Doctor of Philosophy at Lincoln University

Abstract

Rising global atmospheric temperatures are expected to negatively affect farmed animals, especially pigs. To study the likely effects, pigs were exposed to chronic heat stress and their growth, changes to the intestinal microbiota diversity and selected immunological changes in the gut lining were investigated. Six test pigs were subjected to a temperature of 35 ± 2°C and six control pigs to a ‘normal’ temperature of 23 ± 2°C (both at a relative humidity of 75-85% for 21 days, in climatecontrolled pens). Bodyweight, rectal and forehead temperatures were measured, and fresh faeces collected daily from each pig. The rectal temperatures, on average, increased in HS pigs, reaching up to 40.5°C, as compared to 37.6°C in control pigs. Similarly, average forehead temperatures had increased to 39.3°C in heat-stressed pigs and were 36.7°C in the control pigs. The average body weight gains in heat-stressed pigs were 2.9, 1.2, and 0.35 kg, respectively at days 7, 14, and 21, significantly lower (P < 0.05) than for the control pigs. While faeces were collected daily, only samples collected on days 1, 7, 14, and 21 were genetically sequenced to observe for changes. In heat-stressed pigs, marked changes in major microbial phyla composition were observed. The number of Gram-positive Firmicutes decreased, whereas Gram-negative Bacteroidetes increased from day 7 onwards. More importantly, major Gram-negative potential pathogens such as Campylobacterales, Proteobacteria, and Spirochaetes had increased in heat-stressed pigs. This increase in Gram-negative bacterial numbers resulted in upregulation of the NF-κB-mediated proinflammatory cytokines IL-6 and IL-8 via the TLR4/NF-κB-signalling pathway, in the gut lining. Increases in IL-6/IL-8 cytokines cause fever, anorexia, and other metabolic changes. The microbiota and immunological changes resulted in diarrhea and potentially exposed pigs to inflammatory bowel disease, which partly explains the lower bodyweights in the heat-stressed pigs. Such losses in production impose financial strains on farmers, while the presence of pathogens may adversely impact food safety for the ever-increasing population, dependent in-part on low-priced meats like pork. To investigate cross-species transfer, gut microbiota from heat-stressed and control pigs were transplanted into ‘specific pathogen-free mice, and the two groups’ gut microbiota sequenced on days 1, 7, 14 and 21. Similar changes to that observed in the pig gut microbiota were evident in the infected mice after 21 days. This study, hence, confirms that heat stress causes compositional and associated immunological changes in the gut microbiota of pigs; and that microbiota changes that occur in one species can be transferred to another species to cause similar clinical signs, gut microbiota, and body weight changes in the recipient species.