"June, 2004" / Includes bibliographical references (leaves 137-156) / xvi, 156 leaves : ill. (some col.), plates (col.) ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)--University of Adelaide, School of Molecular and Biomedical Science, Discipline of Biochemistry, 2004

Reproduction de : Thèse d'exercice : Médecine vétérinaire : Toulouse 3 : 2007. / Titre provenant de l'écran titre. Bibliogr. p. 117-121.

This thesis examines the challenges of decellularizing porcine lungs. The thesis identifies hydrostatic pressure and the role it plays in perfusate distribution within the lung as a potential roadblock for the decellularization of porcine lungs. The thesis examines methods for mitigating the affect of hydrostatic pressure on the decellularization of porcine lungs. Specifically, the thesis details the design and construction of a vessel that rotates lungs during decellularization. This rotating vessel is evaluated in comparison to two other, non-rotation based, decellularization methods. The rotation device is found to more extensively decellularize the lung than either of the other two options. The thesis concludes by examining what other areas of work could be conducted in this field.

This thesis examines the challenges of decellularizing porcine lungs. The thesis identifies hydrostatic pressure and the role it plays in perfusate distribution within the lung as a potential roadblock for the decellularization of porcine lungs. The thesis examines methods for mitigating the affect of hydrostatic pressure on the decellularization of porcine lungs. Specifically, the thesis details the design and construction of a vessel that rotates lungs during decellularization. This rotating vessel is evaluated in comparison to two other, non-rotation based, decellularization methods. The rotation device is found to more extensively decellularize the lung than either of the other two options. The thesis concludes by examining what other areas of work could be conducted in this field.

Chinese Meishan pigs exhibit exceptional prolificacy compared with common domestic breeds and therefore provide a model for investigating the mechanisms involved in a successful reproductive outcome. Previous studies have shown that the Meishan gains its reproductive advantage through reduced conceptus mortality at the time of conceptus attachment to the uterine endometrium and the establishment of pregnancy. The purpose of this thesis was to consider factors that might affect conceptus viability in the Meishan and a control breed (the Large White x Landrace), to determine where differences arise. Meishan and Large White x Landrace function was compared, in the period leading up to the establishment of pregnancy (days eight to 12 after the onset of oestrus, inclusive). Many factors that might impinge on conceptus survival were investigated, including maternal nutrition, metabolism, endocrinology and conceptus and endometrial morphology and biochemistry. Conceptus oestradiol-17β secretion, believed to be the signal for the establishment of pregnancy in the pig, was delayed and more synchronised in the Meishan than in Large White x Landrace. Also, Meishan plasma progesterone concentrations were elevated compared with the Large White x Landrace gilts. Other studies have positively correlated conceptus survival with raised plasma progesterone concentrations, which can be manipulated by altering the nutritional intake of the dam. The role of conceptus oestradiol-17β in the establishment of pregnancy in the Meishan was also investigated. A lack of aromatase enzyme activity, before day 12 after the onset of oestrus, prevents the conversion of testosterone to oestradiol-17β. Thereafter, substrate availability limits the reaction. Exogenous oestradiol benzoate extended the interoestrous interval of treated Meishan gilts, although differences in the physiology of pregnant and pseudopregnant gilts suggested that other conceptus-specific activity influences the establishment of pregnancy in this breed. These studies have highlighted specific aspects of pig reproduction, manipulation of which might improve the prolificacy of domestic breeds. Further investigations are required to establish the whether such techniques would be practical in the commercial setting.

No description available.

Articular cartilage is an avascular, aneural tissue that covers the ends of diarthroidal joints. Once damaged by disease or injury, cartilage lacks the ability to self-repair. Generating tissue engineered cartilage is an exciting field that may provide a possible solution to this problem. The purpose of this study is to determine the efficacy of a through-thickness perfusion bioreactor to generate scaffoldree tissue engineered cartilage. The results of the study show that allowing long-term static culture to cell constructs before perfusion increases the efficacy of the bioreactor. Immediate perfusion of cell constructs in the bioreactor is shown to decrease the efficacy to produce scaffoldree constructs with desirable biomechanical and biochemical properties. The results of the study also show possible options in future works that could increase the efficacy of the bioreactor.

The management of genomic diversity is important within breeding programs and is primarily achieved through controlling the rate of inbreeding. A failure to adequately manage the rate of inbreeding will result in an increased risk of the expression of lethal recessive mutations, inbreeding depression and losses in genetic variance, thereby restricting long-term genetic progress. Each research chapter within this thesis used real data collected from a commercial pig breeding operation to examine a key area of research regarding the management of genomic diversity. The first research chapter examined the selection outcomes from the practical application of Optimal Contributions (OC). These outcomes were examined to determine their alignment with the current theories regarding selection, particularly as to the extent by which selection decisions were influenced by estimated Mendelian sampling terms. This assessment was conducted for the initial selection of individuals as parents, which parents went on to provide a long-term contribution and the magnitude of these contributions. OC was shown to have shifted breeding decisions more closely in alignment with the estimated Mendelian sampling terms. The second research chapter used genomic data to assess the adequacy of the pedigree-based approach for managing diversity during selection. This approach assumes the infinitesimal model with all loci neutral and no impact from selection per se on heterozygosity. Using genomic information, the observed loss of heterozygosity at each marker was compared to the loss of heterozygosity expected from the pedigree-based relationships. Regional disparities between the observed and expected losses in heterozygosity were detected, which were potentially attributable to selection. Runs of homozygosity and the pairwise linkage disequilibrium between markers were also examined within these regions. Regions showing disparity were found to contain well validated quantitative trait loci for important traits. The third research chapter sought to provide a genomic solution to the shortcomings of the pedigree-based approach for quantifying relatedness, identified above. A methodology was devised for tracing identity by descent (IBD) at each allelic position over five ancestral generations, following phasing and imputation of the genomic data. A comparison was made between the inbreeding expected from the pedigree relationships and that observed from the identity by descent of genomic information. In the population studied it was not currently feasible to derive a relationship matrix based exclusively on observed IBD. The fourth research chapter used imputed genomic information to identify haplotypes which had a putative lethal recessive effect. Haplotypes which were never observed in the homozygous form, either in the population or in the offspring produced between carriers, were classified as candidate haplotypes. The top candidates on each chromosome were then examined for a reduction in the total number born when two carriers were mated together. A total of six putative lethal recessive haplotypes were detected relating to at least four putative lethal recessive mutations, where one homozygote was absent and the size of the reduction in litter size matched that expected for a lethal recessive effect. The research chapters contained within this thesis demonstrate the important role that genomics can have in managing inbreeding in addition to generating genetic gain. Genomics is able to provide a more accurate prediction of the Mendelian sampling term, better quantify the relatedness between individuals and detect lethal recessive effects.

The diagnosis of porcine circovirus-associated disease (PCVAD) in pigs requires the detection of characteristic clinical signs and pathological changes, and the detection of virus in tissues of affected pigs. To increase Australia’s capacity to independently diagnose PCVAD in Australia, techniques for the detection of Porcine circovirus type 2 (PCV2) infection in pigs were developed and are reported in this thesis. These techniques were applied to samples obtained from normal pigs and pigs with disease and confirmed the presence of PCV2 and PCVAD in the Australian pig herd. Viral DNA was detected in tissues of infected pigs by both standard PCR and real-time PCR techniques. The real-time PCR was more sensitive. While the conventional PCR was able to detect approximately 100 copies of the viral genome, the real-time PCR was able to detect 20 copies of the genome. An immunohistochemical (IHC) technique which was also developed enabled the visualisation of PCV2 antigen in fixed tissues of pigs with PCVAD. The techniques that were developed were applied to an examination of tissues from pigs affected by illthrift and increased weaner mortality in herds in South Australia, New South Wales and Western Australia. Lesions suggestive of the PCVAD postweaning multisystemic wasting syndrome (PMWS) were detected and virus antigen was detected in association with lesions. The nature of the clinical signs and histopathological lesions detected, coupled with the presence of PCV2 antigen, suggested that PCVAD was present in some Australian pig herds. Phylogenetic analysis of the strains of PCV2-associated with these disease outbreaks demonstrated they were of a type not previously detected in Australia and similar to strains associated with PMWS in North America. To further assist in investigation of PCV2 infections in the Australian pig herd, an enzyme-linked immunosorbent assay (ELISA) was developed that specifically detected antibody to PCV2 and not the related and non-pathogenic Porcine circovirus type 1. The development of this assay required the production of a virus capsid protein antigen using a prokaryotic protein production system. The ELISA was used to test serum samples form the Australian national pig serum bank. A high prevalence of PCV2 infection was detected in most pig herds examined in all Australian states. International trade in pig meat has resulted in many countries placing restrictions on the importation of pig meat, requiring imported pig meats to be cooked to destroy viral agents. This study investigated the in vitro resistance of an Australian strain of PCV2 to heat treatment at temperatures between 56°C and 85°C. The viability of the virus was determined by a combination of reverse transcriptase polymerase chain reaction (RT-PCR), and IHC to visualise viral capsid antigen within infected cells. This study indicated that PCV2 retained its infectivity following heating up to and including 75°C for 15 mins, but was inactivated following heating to 80°C and above. The investigations reported make a significant contribution to PCV2 research in Australia and ensure Australia’s capacity to independently investigate PCVAD in the Australian pig herd.

The prevention and control of salmonellae in commercial swine operations are becoming increasingly important. The current approach focuses on identifying sources and/or origins of salmonellae contamination before swine are processed for human consumption. The objective of the current study was to assess strain variability among salmonellae grouped by serotype and to determine common origins of contamination (farm or slaughter plant). Salmonellae were previously collected from swine at slaughter, serotyped by the National Veterinary Services Laboratory and stored at - 70??C. Pulsed-field gel electrophoresis (PFGE) was performed to genotypically characterize serotypic isolates using restriction endonuclease XbaI. Dendrogram comparisons were also used to assess genotypic similarity when multiple genotypes existed. This study found PFGE to be more discriminatory than serotyping indicating that multiple genotypic strains existed among selected serotypes. On the basis of PFGE results alone, origins of contamination could not be determined in this study. It is suggested by the author, that origins of contamination could be further defined pending future research, in which in-depth longitudinal studies are included. When used as an adjunct to conventional typing methods, PFGE may prove to be a substantial subtyping system in epidemiologic investigations to identify point-of-entry contaminants to the food chain.