Browsing by Author "Notcovich, Shirli"
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- ItemAn experimental challenge model in lactating dairy cows using Streptococcus uberis for antibiotic efficacy testing : a thesis presented in partial fulfilment of the requirements for the degree of Master of Philosophy in Veterinary Science at Massey University, Palmerston North, New Zealand(Massey University, 2013) Notcovich, ShirliThe aim of this project was to develop a challenge model to test the efficacy of novel intramammary antimicrobial treatments for clinical mastitis. The use of the model, can reduces the costs of testing efficacy and accelerate the process of registration of new products. It provides controlled conditions which safeguard animal welfare. The experimental challenge model using Streptococcus uberis developed in this thesis can provide the pharmaceutical industry and animal health research groups with a cost-effective method to test the efficacy of new antimicrobial products for treatment of mastitis in a safe and controlled environment. Two Cloxacillin-based antimicrobials with different formulations and treatment frequency were tested for their efficacy to cure S. uberis infections after infections were induced using the challenge model developed as described in the third chapter of this thesis. The objective of the first study presented in this thesis was to choose one suitable strain from four strains of S. uberis, to be used in future challenge studies. Four strains were tested for their virulence and susceptibility to antibiotic therapy. A further study objective was to determine the dose (number of pathogens infused, expressed as colony forming units (CFU)) required for the tested strains to produce an acceptable proportion of clinical mastitis cases to enable future studies. The strain which accomplished the desired characteristics was then chosen and was utilised for experimental challenge in further studies (Chapters 4 and 5). The overall incidence of clinical mastitis obtained in this study at a quarter level was 54% (26/48). This study showed significant differences in the ability of different strains of S. uberis to cause clinical mastitis when inoculated via the intramammary route. However, only one of the four strains tested demonstrated favourable characteristics as a strain to be used in experimentally induced clinical mastitis studies. Chapters 4 and 5 describe two challenge studies conducted using the experimental challenge model (Chapter 3) to test the efficacy of different antimicrobial drug formulations. In Chapter 4, the cure rate of one cloxacillin based product applied every 24 hr. was compared with the cure rate of a penicillin-based product applied every 12 hr. During the observation period of this investigation all challenged cows developed clinicalThe aim of this project was to develop a challenge model to test the efficacy of novel intramammary antimicrobial treatments for clinical mastitis. The use of the model, can reduces the costs of testing efficacy and accelerate the process of registration of new products. It provides controlled conditions which safeguard animal welfare. The experimental challenge model using Streptococcus uberis developed in this thesis can provide the pharmaceutical industry and animal health research groups with a cost-effective method to test the efficacy of new antimicrobial products for treatment of mastitis in a safe and controlled environment. Two Cloxacillin-based antimicrobials with different formulations and treatment frequency were tested for their efficacy to cure S. uberis infections after infections were induced using the challenge model developed as described in the third chapter of this thesis. The objective of the first study presented in this thesis was to choose one suitable strain from four strains of S. uberis, to be used in future challenge studies. Four strains were tested for their virulence and susceptibility to antibiotic therapy. A further study objective was to determine the dose (number of pathogens infused, expressed as colony forming units (CFU)) required for the tested strains to produce an acceptable proportion of clinical mastitis cases to enable future studies. The strain which accomplished the desired characteristics was then chosen and was utilised for experimental challenge in further studies (Chapters 4 and 5). The overall incidence of clinical mastitis obtained in this study at a quarter level was 54% (26/48). This study showed significant differences in the ability of different strains of S. uberis to cause clinical mastitis when inoculated via the intramammary route. However, only one of the four strains tested demonstrated favourable characteristics as a strain to be used in experimentally induced clinical mastitis studies. Chapters 4 and 5 describe two challenge studies conducted using the experimental challenge model (Chapter 3) to test the efficacy of different antimicrobial drug formulations. In Chapter 4, the cure rate of one cloxacillin based product applied every 24 hr. was compared with the cure rate of a penicillin-based product applied every 12 hr. During the observation period of this investigation all challenged cows developed clinical mastitis in at least one quarter. The incidence of clinical mastitis at the quarter level was high, with 91.25% (73/80) of challenged quarters being affected. After diagnosis of infections, the cows were randomly allocated to two treatment groups and treated accordingly. Clinical cases in which the quarter did not respond to three applications of the allocated antimicrobial product received an extended treatment of the same product. As the allocation to the extended treatment was not random, clinical and bacteriological cures were statistically evaluated for the short treatment only. Clinical cure rates for the short treatment (3 syringes) were 52.63% and 43.75% for the cloxacillin- and penicillin-based products, respectively. There was no significant difference between the treatments (P = 0.8) in their efficacy for the treatment of experimentally induced S. uberis clinical mastitis. In Chapter 5, two long-acting cloxacillin containing products were compared in their efficacy to cure experimentally induced S. uberis infections. One commercially available product was compared with a novel long acting product (applied every 48 hr.). Out of 80 challenged quarters, 41 quarters developed clinical mastitis after inoculation (51.25%). Treatment with the novel product resulted in a total treatment success rate of 93.1% based on clinical examination, and 96.0% based on the bacteriological cure rate. Treatment with the control product resulted in total treatment success rate of 100% based on clinical and bacteriological cure rate. There was no significant difference between the products (P=0.19) in their efficacy for the treatment of experimentally induced S. uberis clinical mastitis. Results in this thesis showed that experimental challenge models can be a useful tool in animal research to test the efficacy of new products in a safe and cost effective manner.
- ItemThe 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, ShirliThe 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.