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    Characterising the responses of farm mammals to a thoracic squeeze and the relationship to tonic immobility : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Veterinary Science at Massey University, Manawatū, New Zealand
    (Massey University, 2023) Holdsworth, Sophia Ellen
    Applying compression to the thorax of newborn farm mammals causes immobility accompanied by reduced responsiveness. Traditionally, this response was proposed to arise in neonatal foals due to the ‘thoracic squeeze’ mimicking the compression occurring during birth. Recent findings regarding the responses to the squeeze suggest a link to Tonic Immobility (TI). TI is a temporary and reversible state of reduced responsiveness and immobility with characteristic physiological changes. It is elicited by the collective actions of handling and sustained physical contact with additional pressure (restraint) and may be facilitated by inversion. TI is reported in young and adult animals of numerous species. The aim of this thesis was to examine whether responses to the thoracic squeeze are consistent with TI. First, behavioural responses to a squeeze were explored in lambs, with a focus on determining whether responses persisted beyond the neonatal period. Cortisol responses of healthy neonatal piglets to the squeeze were then investigated to explore similarities in Hypothalamic-Pituitary-Adrenal responses between the squeeze and TI. The final study examined electroencephalographic (EEG) responses of healthy neonatal piglets to a thoracic squeeze under light anaesthesia, to determine whether the squeeze causes changes in brain activity or exerts anti-nociceptive effects such as those reported during TI. The results demonstrated that responses to a thoracic squeeze persist beyond the neonatal period in lambs, and responses are generalised across multiple mammalian species. Furthermore, cortisol responses of piglets to a thoracic squeeze followed a similar pattern to that previously observed during TI in other species. Also consistent with some TI studies, the initial handling and restraint required to apply the squeeze appeared to induce the cortisol response in piglets. No inferences could be made about the effects of a thoracic squeeze on state of awareness in neonatal piglets, or the squeeze’s effect on nociception due to methodological limitations. Nevertheless, the results of this research support the hypothesis that the thoracic squeeze may be classified as a stimulus for inducing TI. Further work is required to characterise the effects of the squeeze on awareness and nociception and to explore the affective experiences of animals subjected to the squeeze.
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    The physiology of the keratin plug formation in the teat canal of dairy cattle and its interaction with current and novel methods for prevention of intramammary infections : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Veterinary Science, Massey University, Turitea, Palmerston North, New Zealand
    (Massey University, 2021) Notcovich, Shirli
    The incidence of intramammary infections (IMI) in dairy cows in the early dry period is the highest of the lactation cycle when methods to prevent IMI are not applied. This high incidence is comparable only with that observed near calving. At the end of lactation, the teat is sealed by a plug formed mainly by keratinised cells, detritus and proteinaceous material. Research suggests that the keratin plug acts as a physicochemical barrier throughout the dry period that impedes the entrance of bacteria. However, the physiological mechanism of keratin plug formation is still uncertain. The main objectives of this thesis were to characterise the physiological functions of the teat canal (TC) during the early dry period and assess how they associate with the presence of IMI. A further objective was to evaluate the modes of action of a current mastitis preventative containing bismuth subnitrate and a novel formulation of micronized keratin that is under investigation as a teat seal for preventing IMI during the early dry period. To address these objectives a novel biopsy method was developed to allow investigation of the physiological characteristics of the epithelial tissues of the TC. A transcriptomic analysis of the TC epithelium after drying off showed that epithelial cells decreased expression of mitotic and immune-response related genes. A Streptococcus uberis strain was used in a challenge study aiming to examine mechanisms of colonization in the TC and the response of the epithelial tissue to progressing infection. This Streptococcus uberis challenge did not result in colonization of the TC nor in IMI with S. uberis. Nevertheless, a reduction in the thickness of the stratum granulosum and the keratin layer of the TC epithelium was observed. This coincided with an increase in TC colonization by non-pathogenic bacteria and a decline in the concentration of certain cytokines after drying off. These changes observed in the TC epithelium support previous reports showing increased incidence of IMI by non-pathogenic bacteria during the early dry period. Antimicrobial effects and neutrophil cell responses were evaluated in vitro in two studies to test previously hypothesised action mechanisms for bismuth subnitrate and a novel keratin-based internal teat sealant (ITS) formulation. Bismuth subnitrate showed an inhibitory effect on bacterial growth, contrary to the current description of ITS as non-pharmacological, inert physical barriers. No activation of a cellular response was observed for keratin or bismuth formulations in vitro. Bismuth subnitrate and keratin were also tested in vivo for their effect on the formation of the keratin plug. The hypothesis of this study was that these treatments induce expression of mitogenic genes that induce a faster sealing of the teat canal. There was no modification of gene expression after treating cows with bismuth subnitrate or the novel keratin-based ITS formulation during the formation of the natural keratin plug, and no modification of the closure status of the teat canal lumen, suggesting that neither of the two treatments induced an improved sealing of the teat canal after drying off through increased keratin production. These findings contribute to the knowledge of keratin plug formation and physiological characteristics of the TC during involution. They align with and partially explain some of the literature-reported events observed during the early dry period. The knowledge gained provides support for future product development aimed to increase protection of the mammary gland during the dry period.
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    Behavioural effects of a thoracic squeeze on healthy neonatal mammals of precocial species : a thesis presented in partial fulfilment of the requirements for the degree of Master of Science in Physiology at Massey University, Manawatū, New Zealand
    (Massey University, 2020) Holdsworth, Sophia Ellen
    Some neonatal farm mammals that are apparently healthy do not behave normally after birth and without intervention, often die. Recently, a thoracic squeeze has been applied to these neonates, which observed their entrance into a ‘non-responsive state’, from which they rapidly recover when the squeeze is released to express normal healthy behaviours. In addition, healthy newborn foals also become non-responsive during application of the thoracic squeeze. To date, there have been no systematic studies into the effects of the thoracic squeeze on healthy neonates of other precocial mammalian species. This preliminary research aimed to describe the responses of healthy newborn piglets and calves to a standardised application of the thoracic squeeze, and evaluate the effectiveness of the method of squeeze application in eliciting a non-responsive state. Neonatal piglets and calves were squeezed around the chest with either a soft fabric rope or a modified inflation cuff. Physiological parameters were monitored for abnormal changes, while induction behaviour and neural reflex activity were measured. Behavioural data indicated that the thoracic squeeze was effective at eliciting a non-responsive state in all piglets in the study and maintaining this state in half of all the piglets induced, with reduced or absent reflexes in nearly all piglets. In contrast, 81% of the calves were induced into a lower-responsive state, with half of all calves squeezed maintaining this state for the full observation period and present reflexes observed for nearly all calves. Furthermore, the cuff appeared to be more effective at inducing both piglets and calves into a non-responsive or lower-responsive state than the rope, with a faster application and removal. These findings suggest that the non-responsive state reported in foals can be generalised to neonates of other precocial species. The thoracic squeeze has been demonstrated to be inherently safe and effective at inducing and maintaining a non-responsive state in neonates. This pilot study provides a foundation for further research using the inflation cuff to explore the mechanisms underlying the thoracic squeeze and ways in which the squeeze can be effectively applied in an industry context.
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    The effect of early life nutrition on rumen microbial community development and impact on lifetime performance in ruminants : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Veterinary Sciences at Massey University, Palmerston North, New Zealand
    (Massey University, 2019) Cristobal Carballo, Omar
    Manipulation of the rumen microbiota in adult ruminants has been intended to improve animal performance and decrease greenhouse gas emissions, but results have only shown a short- or non-lasting effect after intervention. Changes in the ruminal microbiota during rumen development have recently shown promising results in the short-term. Therefore, the purpose of the present body of work was to determine how dietary management and chemical interventions, during rumen development, modify the ruminal microbial community composition, and whether these changes affect rumen fermentation and development, and consequently, performance in the young ruminants. The objectives of this thesis were to: (i) evaluate the impact of early weaning on rumen development and function in artificially-reared lambs; (ii) characterize the impact of early weaning in lambs on the rumen microbiota in the first 16 weeks of life and examine the relationships between rumen microbiota composition and rumen fermentation profiles, rumen development and blood metabolites; (iii) assess whether contrasting feeding regimes in the first 7 months of life lead to an imprint in the rumen microbial community structure, fermentation profiles and methane emissions in the rumen of calves; (iv) and evaluate the effect of methane inhibitors on the rumen microbial community composition, fermentation pathways, and gas emissions in calves. A series of three experiments were carried out in young ruminants separated from their mothers after colostrum intake, to address the objectives of this thesis. In experiment one, 3-5-day-old lambs were euthanized at weeks 0, 4 and 16 of rearing to investigate objectives (i) and (ii). Early weaning of lambs increased plasma hydroxybutyrate at week 4 of rearing, while dry matter intake, fermentation profiles and rumen morphology were similar between groups. Papillae morphology and muscular thickness differed between ruminal sites at 4 and 16 weeks of rearing, but not between treatments. Diversity and relative abundance of ruminal bacteria was affected by feeding management, whilst the archaea community showed few changes. Changes in the proportions of abundant bacteria genera from Bacteroidetes and Firmicutes were associated with fermentation profiles, rumen morphology and blood metabolites; however, further investigations are required to explain these associations. In experiment two, ~1-week-old calves were reared with two divergent feeding systems and different post-weaning forage quality with a common pasture diet after 7 months of age to investigate objective (iii). Consumption of pre-weaning concentrate compared to forage produced lower methane yields and greater total short chain fatty acids (SCFA) concentrations and propionate proportions; whist ruminal microbes showed greater proportions of saccharolitic bacteria and Methanobrevibacter boviskoreani, but lower hemicellulolytic and cellulolytic bacteria, and Mbb. gottschalkii. Post-weaning, high-quality forage produced greater total SCFA concentration and propionate proportions than low-quality forages, while methane yield was similar. Hemicellulolytic bacteria and Methanosphaera spp. were greater in high-quality forages, while cellulolytic bacteria and Methanomassiliicoccales spp. were greater in low-quality forages. No pre-weaning effect was observed. Finally, the consumption of a common diet after 7 months of age resulted in similar methane emissions, fermentation profiles and microbial communities. In experiment three, ~1-week-old calves fed either concentrate starter diets or starter diets plus methane inhibitors were tested to evaluate objective (iv). Inhibitor intake decreased methane yield, but increased hydrogen yield and the proportion of propionate and had no effect on dry matter intake, total SCFA concentrations or animal growth. Within the abundant bacteria, the proportions of hydrogen utilizing and producing bacteria increased and decreased, respectively. Archaea diversity and proportions were affected during the period of methane inhibitor intake. However, similar gas emissions, fermentation profiles, and microbial communities were observed between groups at 24 and 49 weeks of age. Collectively, these results showed that reducing the age at weaning and introducing the solid feed to lambs at ~1 week of life accelerated some aspects of rumen morphology and function. Dietary management and methanogen inhibitor interventions affected the composition of the ruminal microbiota and fermentation profiles during treatment, however, no permanent changes in the microbial community and resulting ruminal fermentation were observed post-treatment in young ruminants.