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ItemStudies on gastrointestinal nematodes of alpacas in AustraliaRashid, Mohammed Haronur ( 2019)The Australian alpaca industry has grown remarkably since the re-introduced of alpaca into the country in the 1980s. This relatively new and small industry faces various challenges that affect the overall production (fibre, meat etc.) of animals. For example, production losses and deaths associated with gastrointestinal nematodes (GINs) have become one of the major concerns for alpaca producers in Australia and elsewhere as there is a scarcity of reliable information about the epidemiology of GINs in alpacas, worm control practices used by alpaca farmers and the efficacy status of widely used unregistered anthelmintics. The present thesis aimed to (i) assess worm control practices used by Australian alpaca farmers, (ii) determine the epidemiology GINs and (iii) the efficacy of commonly used anthelmintics against GINs of alpacas in Australia. A comprehensive literature review along with seven results chapters and a general discussion chapter are included in this thesis. In Chapter 2, an online questionnaire survey was conducted to assess worm control practices used by Australian alpaca farmers. Results showed that more than half of the respondents perceived that GINs was an important health problem. Macrocyclic lactones (MLs) were the most commonly used anthelmintics in alpacas. Almost half of the respondents (47%) used anthelmintics at the dose rate recommended for sheep. The majority of alpaca farmers were unaware of pasture spelling and one-third of respondents had shared paddocks with ruminants. This study led to subsequent studies to understand the epidemiology of GINs of Australian alpacas using sensitive diagnostic methods and assess the efficacy of commonly used anthelmintics against GINs of Australian alpacas. In chapters 3 and 4, sensitive diagnostic methods were developed to determine the faecal eggs counts and identify nematode genus/species in the faeces of alpacas. In chapter 3, a newly developed diagnostic method, the FECPAKG2, was compared to a routinely used method, the McMaster technique, for the counting of GIN eggs in the faeces of alpacas. Data revealed moderate to good agreement between the two methods. This was the first study to assess the agreement of measurements between two methods for estimating nematode eggs in the faeces of alpacas. In Chapter 4, a molecular diagnostic tool based on multiplexed-tandem PCR (MT-PCR) was developed. This tool is faster and is capable of identifying common nematode genera/species of alpacas, including Camelostrongylus mentulatus which was not possible using traditional larval culture method. The epidemiology of GINs of alpacas in Australia was assessed using cross-sectional (Chapter 5) and longitudinal studies (Chapter 6). A range of GINs are prevalent in Australian alpacas with variable worm burdens in different climatic zones and seasons. The results of both studies were comparable. Both studies showed an overall prevalence of GINs in SACs from 61 – 66%. Weaners had the highest prevalence in both studies ranging from 73 - 80%. However, the pattern of prevalence was not same across the climatic zones. In cross-sectional study, the highest prevalence of GINs (77%) were observed in the summer rainfall zone, whereas in longitudinal study the winter rainfall zone had the highest prevalence (68%). In addition, a mixed-effects zero-inflated negative binomial (ZINB) regression model has been used to design parasite control interventions. To assess worm burden and the spectrum of GINs infecting Australian alpacas, 100 gastrointestinal tracts of alpacas were examined (Chapter 7). Results revealed a mean burden of 1,300 worms, with the highest burden of 29,000 worms. Nineteen different species of GINs were identified from Australian alpacas, including three camelid specific nematodes: Graphinema auchenia, Camelostrongylus mentulatus and Trichuris tenuis. Haemonchus contortus was the most prevalent nematode followed by C. mentulatus and Trichostrongylus spp. In Chapter 8, the efficacy of commonly used anthelmintics against GINs of alpacas was assessed. The faecal egg count reduction tests were conducted on 20 alpaca farms across the country. The results showed that a commercially available combination of levamisole, closantel, albendazole and abamectin was the most effective dewormer followed by single anthelmintic such as, monepantel, moxidectin, closantel, fenbendazole and ivermectin. Haemonchus spp. were the most commonly resistant nematodes. This was the first comprehensive study to investigate the efficacy of commonly used anthelmintics against GINs of alpacas. This study provides significant information on GINs of Australian alpacas. Results of this study advance our knowledge on the epidemiology and control of GINs and efficacy status of most commonly used anthelmintics which could be used to develop control strategies against GINs of alpacas in Australia.
ItemMolecular epidemiological investigations of Enterocytozoon bieneusiZhang, Yan ( 2019)Enterocytozoon bieneusi is a microsporidian microorganism that causes intestinal disease in animals including humans. Although E. bieneusi has been discovered and investigated in numerous countries around the world, at the commencement of this PhD project, there was no record of E. bieneusi in animals in Australia and no information on the prevalence and genetic make-up of E. bieneusi populations in this country. Thus, the present project explored E. bieneusi of a number of species of wild and domestic animals as well as humans in parts of Australia using a molecular approach, in order to improve our knowledge of the epidemiology of this microsporidian. Nested-PCR-based sequencing of the internal transcribed spacer (ITS) and/or small subunit (SSU) of nuclear ribosomal DNA was used for the detection of E. bieneusi DNA in faecal samples and the genotypic characterisation of this microbe. In the first instance, three species of wild deer (Cervus elaphus, Dama dama and Rusa unicolor; n = 610) and three species of wild marsupials (Macropus giganteus, Vombatus ursinus and Wallabia bicolor; n = 1365) in Melbourne's water catchment areas were investigated (Chapters 2 and 3). Here, E. bieneusi was detected in 4.1% of sambar deer and 1.4% of marsupials. Phylogenetic analyses of sequence data showed that genotypes D, J, MWC_d1, MWC_d2 and Type IV (in sambar deer) and MWC_m1 and NCF2 (in marsupials) clustered with E. bieneusi genotypes recorded previously in humans, indicating the zoonotic potential of these genotypes. Studies of cattle on farms located near Tarago reservoir (n = 471; Chapter 4) in Victoria, and alpacas in the states of New South Wales, Queensland, South Australia, Victoria and Western Australia (n = 81; Chapter 5) revealed relatively high prevalences of E. bieneusi in both cattle (10.4%) and alpacas (9.9%). The phylogenetic analysis of ITS sequence data revealed six genotypes (BEB4, I and J, TAR_fc1, TAR_fc2 and TAR_fc3) in cattle and three (ALP1, APL3 and P) in alpacas, all recognised to have zoonotic potential, indicating that these herbivores might act as reservoirs for human infection. Subsequently, a study of companion animals (cats and dogs; n = 514) identified genotype D, which is commonly detected in humans (Chapter 6). However, surprisingly, an investigation of humans (n = 605) in the states of Queensland and Western Australia (Chapter 7) did not identify the genotypes previously found in cats, dogs and wildlife in the state of Victoria, although, surprisingly, a known genotype (i.e., ALP1) recorded in farmed alpacas (Chapter 5) was identified in humans for the first time (Chapter 7). This latter finding raises questions about the transmissibility of this genotype from alpaca to humans. Overall, this thesis has recorded E. bieneusi, for the first time, in a variety of animals in Australia, and provides a first glimpse of the epidemiology of this microsporidian in this country (Chapter 8). The findings of this thesis indicate that the animals studied here, including eastern grey kangaroos, swamp wallabies and wombats; sambar deer, cattle and alpacas; and cats and dogs, can all carry E. bieneusi genotypes that might infect humans via water, food and/or the environment. To provide further evidence to support this proposal and much deeper insights, well-designed large-scale (temporal and spatial) epidemiological studies are required. Population genomics of E. bieneusi using advanced next-generation sequencing and informatics tools could significantly support this endeavour. Other future investigations might also, for example, attempt to establish in vitro culture and in vivo systems for E. bieneusi for controlled studies of the biology of distinct genotypes of this microbe as well as approaches for assessing the viability and infectivity of E. bieneusi and anti-infectives against this microbe. Clearly, we are only beginning to understand some aspects of this enigmatic pathogen - E. bieneusi.