Please read the abstract in the section 00front of this document / Thesis (DPhil)--University of Pretoria, 2000. / Veterinary Tropical Diseases / unrestricted

The parasite secretome has been shown to play a key role in both pathogenicity and the regulation of host defence, allowing pathogens, such as helminths, to establish a chronic infection within the host. The recently discovered presence of extracellular vesicles within parasite-derived excretory-secretory products introduces a new mechanism of potential cross-species communication. Extracellular vesicles (EVs), such as exosomes, facilitate cellular communication through the transfer of small RNAs, lipids and proteins between cells and organisms across all three kingdoms of life. In addition to their roles in normal physiology, EVs also transport molecules from pathogens to hosts, presenting parasite antigens and transferring infectious agents. Here, I examine secreted vesicles from the murine gastrointestinal nematode Heligmosomoides polygyrus, and their potential role in the host-helminth interactions. Transmission electron microscopy reveals vesicle-like structures of 50- 100 nM in the ultracentrifuged secretory product, and potential evidence of multi-vesicular bodies in the worm intestine. This, coupled with information from the exoproteome, helped support the hypothesis that exosomes originate from the parasite intestinal tract. I have completed a series of studies looking at the fundamental properties of exosome-cell interactions, providing comparative studies between mammalian and H. polygyrus-derived exosomes. I have determined some of the key factors influencing exosome uptake, including time of incubation, cell type and exosome origin. Through microarray analysis of H. polygyrus exosome-treated small intestinal epithelial cells, we see significant gene expression changes, including those involved in the regulation of signalling and the immune response, such as DUSP1 (dual-specificity phosphatase) and IL1RL1 (the receptor for IL-33). The modest reduction of inflammatory cytokine responses by exosomes in small intestinal cell lines was amplified in immune cells, such as macrophages. Exosomes can significantly reduce expression of classical activation markers, as well as inflammatory cytokine production in the macrophage cell line RAW 264.7, and this is further supported by similar responses in bone marrow-derived macrophages. Owing to their suppressive nature, I demonstrate that immunization of mice with an exosome/alum conjugate generates significant protection from a subsequent H. polygyrus larval challenge, as seen through a reduction in egg counts and worm burden. I have investigated the role of the IL33 receptor (IL-33R); a key molecule associated with parasitic resistance that is suppressed by exosomes in type-2 associated immune responses. Uptake of H. polygyrus-derived exosomes by alternatively activated macrophages caused the suppression of type 2 cytokine/protein release and the reduction of key genes associated with this phenotype. In addition, there was also significant repression of both transcript and surface T1/ST2, a subunit of the IL-33R). Using a model of lung inflammation, in vivo studies demonstrate that, in both prophylactic and co-administration experiments, exosomes modulate the innate cellular response. This is represented by changes in the number of innate lymphoid cells (ILCs), bronchoalveolar lavage eosinophils and type-2 cytokine output. In this system, the expression of T1/ST2 on type 2 ILCs was also significantly reduced. I have extended the investigation on exosome-IL-33R responses by using T1/ST2 knockout mice. Despite generating strong antibody responses, vaccination against exosomes could not protect T1/ST2 knockout mice against a subsequent infection. This work suggests that exosomes secreted by nematodes could mediate the transfer and uptake of parasite products into host cells, establishing cross-species communication to suppress the host ‘danger’ or inflammatory response.

Made available in DSpace on 2016-04-28T20:16:22Z (GMT). No. of bitstreams: 1 2007- Sthefane D avila de Oliveira e Paula-01.pdf: 3469002 bytes, checksum: 6411676587b6fa03e9508964399be8e0 (MD5) Previous issue date: 2007-02-15 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / The aim of the present study was to obtain data on helminth fauna in Columba livia, in municipality of Juiz de Fora, Minas Gerais, Brasil; to describe the structure of the helminth infracommunities present in this host and to clarify questions concerning morphology, sistematics and ecology of some of the component species. The helminthological study of 35 hosts revealed the presence of two digenetic trematodes, Tanaisia (Paratanaisia) bragai (prevalence 51.42%, mean intensity 288.8 ? 403.86 and mean abundance 148 ? 320.9) and T. inopina (prevalence 2.85% and mean abundance 0.68 ? 4.05); five cestodes, Raillietina allomyodes (prevalence 34.28%, mean intensity 6.66 ? 9.14 and mean abundance 2.28 ? 6.11), Raillietina sp. (prevalence 37.14%, mean intensity 9 ? 10.68 and mean abundance 3.34 ? 7.7), Skrjabinia bonini (prevalence 20%, mean intensity 2.14 ? 1.21 and mean abundance 0.42 ? 1), Skrjabinia sp.( prevalence 5.7%, mean intensity 6 ? 7 and mean abundance 0.34 ? 7) and Fuhrmanneta sp. (prevalence 2.85% and mean abundance 0.028 ? 0.16) and four nematodes, Baruscapillaria obsignata (prevalence 51.42%, mean intensity 29.72 ? 44.2 and mean abundance 15.28 ? 34.7); Ascaridia columbae (prevalence 51.42%, mean intensity 60.55 ? 79.88 and mean abundance 31.14 ? 64.2); Tetrameres fissipina (prevalence 14.28%, mean intensity 346.3 ? 504.4 and mean abundance 49.42 ? 212.1) and Synhmanthus (Dyspharynx) nasuta (prevalence 2.85% and mean abundance 0.028 ? 0.16). Among the examined hosts, 97.2 % were found parasitized by at least one helminth species. In accordance with the prevalence of each species T. bragai, A. columbae and B. obsignata were considered secondary species and T. inopina, T. fissipina, S. nasuta, S. bonini, Skrjabinia sp., R. allomyodes, Raillietina sp. and Fuhrmanneta sp. were considered satellite species. All the species exhibited aggregate distributions, wich is the most common distribution pattern in helminth populations. Taxonomic and ecologic aspects of B. obsignata infrapopulations were analised and the results are discussed in terms of possible factors influencing the processes that lead to niche restriction and biased sex ratios in parasite infrapopulations. Additionaly, studies on morphology and sistematics of some component species were performed. Tanaisia (Paratanaisia) bragai and T. inopina specimens were analised by means of confocal laser scanning microscopy. Two new species of Raillietina and Skrjabinia were described and the sequences of the ITS2 ribossomal DNA of these species were determined and combined with other available Raillietina and Skrjabinia sequences in GenBank, in order to perform a phylogenetic study. / O conhecimento da helmintofauna de Columba livia, no Brasil, teve in?cio com as investiga??es a respeito do parasitismo em animais dom?sticos. Esses he lmintos foram relativamente bem estudados, sob o aspecto faun?stico. No entanto, existem lacunas no conhecimento da morfologia, sistem?tica e ecologia das esp?cies. O conhecimento sobre a morfologia e, consequentemente, os caracteres utilizados na sistem?tica, se restringem quase totalmente `as informa??es que a microscopia de luz pode fornecer. N?o existem estudos sobre a ecologia das infracomunidades desses parasitos, no Brasil. Al?m disso, as rela??es entre algumas esp?cies ainda s?o incertas, sendo necess?ria a revis?o da posi??o taxon?mica e filogen?tica desses helmintos. O presente trabalho teve como objetivos gerais: 1- conhecer a helmintofauna de C. livia, no munic?pio de Juiz de Fora, Minas Gerais; 2- determinar a estrutura da comunidade desses helmintos; 3- elucidar quest?es relativas ? morfologia, sistem?tica e ecologia de algumas das esp?cies componentes dessa comunidade. Para este fim, foram necropsiadas 35 aves e os helmintos coletados foram processados segundo as t?cnicas helmintol?gicas conve ncionais e quantificados. Foram calculados o ?ndice de dispers?o, para determinar o padr?o de distribui??o das infrapopula??es; o ?ndice de concentra??o de domin?ncia de Simpson, para avaliar a presen?a de esp?cie dominante na estrutura da comunidade e o ?ndice de Brillouin (H), para determinar a diversidade parasit?ria da comunidade. O estudo helmintol?gico revelou a presen?a de duas esp?cies de digen?ticos, Tanaisia (Paratanaisia) bragai (preval?ncia 51.42%, intensidade m?dia 288.8 ? 403.86 e abund?ncia m?dia 148 ? 320.9) e T. inopina (preval?ncia 2.85% e abund?ncia m?dia 0.68 ? 4.05); cinco esp?cies de cest?ides, Raillietina allomyodes (preval?ncia 34.28%, intensidade m?dia 6.66 ? 9.14 e abund?ncia m?dia 2.28 ? 6.11), Raillietina sp. (preval?ncia 37.14%, intensidade m?dia 9 ? 10.68 e abund?ncia m?dia 3.34 ? 7.7), Skrjabinia bonini (preval?ncia 20%, intensidade m?dia 2.14? 1.21 e abund?ncia m?dia 0.42 ? 1), Skrjabinia sp. (preval?ncia 5.7%, intensidade m?dia 6 ? 7 e abund?ncia m?dia 0.34 ? 7) e Fuhrmanneta sp. (preval?ncia 2.85% e abund?ncia m?dia 0.028 ? 0.16) e quatro esp?cies de nemat?ides, Baruscapillaria obsignata (preval?ncia 51.42%, intensidade m?dia 29.72 ? 44.2 e abund?ncia m?dia 15.28 ? 34.7); Ascaridia columbae (preval?ncia 51.42%, intensidade m?dia 60.55 ? 79.88 e abund?ncia m?dia 31.14 ? 64.2); Tetrameres fissipina (preval?ncia 14.28%, intensidade m?dia 346.3 ? 504.4 e abund?ncia m?dia 49.42 ? 212.1) e Synhimanthus (Dyspharynx) nasuta (preval?ncia 2.85% e abund?ncia m?dia 0.028 ? 0.16). Aspectos da taxon?mia e ecologia das infrapopula??es de B. obsignata foram abordados e os resultados discutidos em termos dos poss?veis fatores com influ?ncia nos processos que levam `a restri??o de nicho e raz?es sexuais enviezadas. Adicionalmente, foram realizados estudos sobre a morfologia e sitem?tica de algumas das esp?cies componentes. Aspectos da morfologia de Tanaisia (Paratanaisia) bragai e T. inopina, foram elucidados utilizando-se como ferramenta a microscopia de varredura a laser confocal. Um estudo filogen?tico de esp?cies dos g?neros Raillietina e Skrjabinia, foi realizado atrav?s da an?lise de seq??ncias da regi?o espa?adora transcrita interna 2 (ITS2) do DNA ribossomal e duas esp?cies novas foram descritas.

Over one third of the human population is currently infected by one or more species of parasitic helminth, but the immune responses elicited by these infections remain poorly defined. Studies in helminth-exposed human populations and laboratory models suggest that helminth infection elicits a range of different effector cell types and that protective immunity and resistance to immune-mediated pathology depends on the balance between these responses. The aim of this thesis was to investigate how cytokines, the molecular mediators of the immune system, can be used to characterise human immune phenotype during natural and experimental helminth infection. Cytokines associated with innate inflammatory (TNFα, IL-6 and IL-9), Thl (IFNγ, IL-2 and IL-12p70), Th2 (IL-4, IL-5 and IL-13), Th17 (IL-17A, IL-21 and IL-23) and regulatory (IL-10 and TGFβ)immune phenotypes were analysed to provide the most comprehensive analysis of cytokine responses in human helminth infection conducted to-date. Using a multivariate statistical approach cytokines were analysed as combined immune profiles to reflect their complex interactions in vivo. In the first part of the study venous blood samples collected from a cross-sectional cohort of 284 Zimbabweans (age range: 3 -86 years) endemically-exposed to Schistosoma haematobium were cultured with antigens from different stages of the parasite's life-cycle(cercariae, adult worms and eggs) and the anti-schistosome vaccine candidate antigen glutathionine-S-transferase (GST). Cytokines responses were quantified in culture supernatants via enzyme-linked immunosorbent assay (ELISA). These assays were repeated 6 weeks after clearance of infection by anti-helminthic treatment. Parasitological and demographic characterisation of the cohort before, 6 weeks, 6 and 18 months after treatment allowed cytokine responses to be related to epidemiological patterns of infection before treatment and the risk of re-infection after treatment. The main findings of this study were:Cytokine responses to the antigens of S. haematobium cercariae are more proinflammatory than those elicited by adult worms and eggs prior to treatment, reflecting the distinct proteomes and exposure patterns of the 3 life-cycle stages Young children (5-10 years old) have a more regulatory and Th17-polarised cytokine response to S. haematobium antigens than older children and adults. These responses are significantly associated with schistosome infection intensity and may contribute to the development of resistance to schistosomiasis with age and exposure to infection Anti-helminthic treatment leads to a shift in S. haematobium cercariae, egg and GST specific cytokine responses towards a more pro-inflammatory phenotype The magnitude of change in S. haematobium-specific cytokine profiles after treatment is dependent on schistosome infection intensity at the time of treatment Individuals who remain un-infected up to 18 months after treatment to clear schistosome infection have a more pro-inflammatory and IL-21-polarised response to S. haematobium antigens 6 weeks after treatment than those who become re-infected, suggesting that post-treatment cytokine profiles promote resistance to re-infection. The second part of the study assayed systemic, parasite and allergen-specific cytokine responses in 45 adults with seasonally exacerbated allergy to grass pollen who were experimentally exposed to Trichuris suis. Cytokine responses in infected individuals were compared to those of 44 un-infected controls. This aspect of the study showed that: Exposure to T. suis promotes systemic and parasite-specific Th2 and regulatory cytokine responses, but does not alter cytokine responses to environmental allergens.

Macrophages are phagocytic cells of the innate immune system, which have a central role in immune surveillance, tissue homeostasis and the immune response to bacterial, viral, protozoan and helminth parasites. It is now appreciated that many tissue resident macrophage (resMΦ) populations, including those in the peritoneal and pleural cavity, are derived prenatally prior to the establishment of definitive haematopoiesis in the bone marrow. Once seeded, these resMΦ populations are long-lived and capable of self-renewal via in situ proliferation driven by CSF-1. An inflammatory insult, such as bacterial infection, results in the recruitment of bone marrow derived macrophages (BMDMΦ) and the disappearance of the resMΦ population. BMDMΦ recruited to the site of infection become classically activated upon engagement of pathogen recognition receptors and subsequent STAT1 induction. Classically activated macrophages (CAMΦ) are highly bactericidal through the production of inflammatory cytokines, which direct the TH1 immune response, and upregulation of iNOS to generate high concentrations of intracellular nitric oxide. During resolution of acute inflammation resMΦ undergo a CSF-1 driven proliferative burst to repopulate the tissue. In contrast to bacterial infection, helminth parasites drive a TH2 immune response characterised by CD4+ T cell production of IL-4, which induces proliferation and alternative activation of the resMΦ population, thereby overcoming the need for an inflammatory influx of BMDMΦ. Alternatively activated macrophages (AAMΦ) are generated through signalling from the IL-4Rα subunit and subsequent expression of the molecules RELMα, YM1 and arginase-1. While both BMDMΦ and resMΦ upregulate RELMα, YM1 and Arg-1 in response to IL-4Rα stimulation, microarray analysis revealed an otherwise diverse transcriptional and cell surface phenotype between these populations. It was hypothesised that the diverse modes of macrophage accumulation enlisted by bacterial and helminth parasites, combined with the distinct alternative activation phenotypes employed by BMDMΦ and resMΦ populations would translate into important functional differences as regards anti-parasitic immunity. Chapter 1 and 2 of this thesis addresses the importance of macrophage origin during infection with the filarial nematode Litomosoides sigmodontis, taking advantage of the naturally occurring resistant C57BL/6 and susceptible BALB/c strains. A large disparity in MΦ accumulation was observed throughout the infection time course, with significantly larger numbers present within the pleural cavity of resistant C57BL/6 mice. This difference in MΦ number was a reflection of enhanced F4/80hi resMΦ accumulation. Through Ki67hi staining and the use of CCR2-/- and partial bone marrow chimeric mice, the expanded F4/80hi population in resistant C57BL/6 mice was shown to be a result of proliferation of the local F4/80hiGATA6+CD102+ resMΦ population. A high degree of BMDMΦ incorporation into the resMΦ pool through assumption of an F4/80hiGATA6+CD102+ phenotype was observed in both naïve and infected bone marrow chimeric animals, supporting a recent publication showing gradual incorporation of these cells into the resMΦ niche with age. Importantly, the degree of BMDMΦ incorporation into the F4/80hi population was equivalent between naïve and infected animals, despite a 27-fold difference in cell number, illustrating that expansion is a result of proliferation of local resMΦ, independent of origin. Susceptibility was marked by reduced resMΦ proliferation and enhanced recruitment of bone marrow derived F4/80lo MΦ and monocytes. These recruited BMDMΦ displaced the resMΦ population, failed to integrate the resMΦ niche and were highly positive for PD-L2, a marker specific to BMD AAMΦ. Prevention of monocyte influx and subsequent resMΦ displacement resulted in increased worm killing and a stronger TH2 immune response in susceptible BALB/c mice, thereby confirming a detrimental role for BMD AAMΦ in worm killing. Conversely, in order to confirm a protective role for the expanded resMΦ in resistant C57BL/6 mice we attempted to deplete the resMΦ population through intrapleural delivery of clodronate-loaded liposomes. Due to technical issues we were unable generate statistically significant results when depleting the resMΦ population, however a trend toward decreased worm killing in the absence of resMΦ is evident. Previously generated microarrays in the lab identified the complement cascade as being highly upregulated by AAMΦ induced in response to Brugia malayi infection compared to BMDMΦ (thioglycollate elicited). To investigate the role of complement in resistance to L. sigmodontis, Chapter 3 briefly phenotypes the macrophage compartment of C3-/- C57BL/6 mice during infection. No differences in worm burden or macrophage phenotype could be detected in C3-/- mice compared to WT controls, however this may be explained through differences in strain or MΦ origin. This chapter provides an important foundation for future studies on complement and its role in worm killing during L. sigmodontis infection. The final chapter of the thesis focuses on examining the bactericidal capabilities of BMD and resMΦ populations. An in vitro system was utilised to assess the interaction of bone marrow derived macrophages (thioglycollate elicited) and ResMΦ (from naïve mice) with Salmonella enterica serovar Typhimurium SL3261. I found that in vitro BMDMΦ are infected by/ingest SL3261 to a much greater degree than resMΦ. The resMΦ population is less efficient at both controlling the spread and killing intracellular SL3261 overtime, compared to the BMDMΦ population. In vivo however, there appears to be no difference in the ability of the monocyte derived F4/80lo MΦ and the F4/80hi resident MΦ to be infected by/ ingest SL3261, nor was a difference in bactericidal ability detected. Ultimately my work highlights that the anti-parasitic functions of MΦ populations are not dictated by origin but rather the activation phenotype upon infection and ability to respond to local stimuli.

Transmissible spongiform encephalopathies (TSEs) or prion diseases, are subacute neurodegenerative diseases that infect humans and animals. Many of these diseases are acquired by peripheral exposure (e.g. orally). After oral exposure prion replication within the Peyer's patches (PP) in the small intestine is necessary for the efficient spread of the disease to the brain. Within the intestine, bacteria and pathogenic microorganisms can affect the status of the gut associated lymphoid tissue (GALT). GALT consists of PP and isolated lymphoid follicles (ILF) that maintain homeostasis and protect from infections. Therefore, factors which modify GALT status, might dramatically affect oral prion disease pathogenesis by influencing the uptake of prions from the gut lumen or expanding their distribution within the host. Chronic intestinal helminth infections are common in animals and in man, and can cause significant pathology within the intestine. Little is known of the effects that intestinal helminth infections may have on oral prion diseases susceptibility. Therefore, in this study the influence that co-infection with Heligmosomoides polygyrus (a natural pathogen of the mouse small intestine) may have on oral prion disease pathogenesis and susceptibility was determined. The studies consisted of groups of 4 (for H. polygyrus characterization and for early prion detection) and 8 (for H. polygyrus-prion co-infection to terminal stage) mice infected with H. polygyrus (orally) alone or subsequently infected with ME7 scrapie prions (orally) at different time-points after parasitic infection. The effects of the H. polygyrus infection alone, and on oral prion disease pathogenesis and susceptibility were then determined. Initially the characterization of H. polygyrus infection on the host intestine revealed that this parasite caused significant pathology in the small intestine and affected the GALT microarchitecture. In the PP follicles, H. polygyrus infection increased the area of follicular dendritic cell expression, altered the positioning of mononuclear phagocytes and increased M cell density. H. polygyrus infection also reduced the number of ILF in both the small and large intestines. Additional studies in mice co-infected with a low dose of prions, revealed that these pathological changes affected the survival time and disease susceptibility. Data also show that the extent of the effects on prion disease pathogenesis and susceptibility were dependent on the stage of the helminth infection at which the mice were orally-exposed to prions. Data demonstrate that co-infection with the gastrointestinal helminth H. polygyrus can influence oral prion disease pathogenesis and susceptibility. Helminth infections can significantly modify the microarchitecture of the gut and the GALT. Data presented suggest the pathological changes that pathogens such as small intestinal helminths cause, may also influence the uptake of prions from the gut lumen after oral exposure.

Approximately two billion people are infected with helminths worldwide. In order to develop a vaccine against these pathogens, more needs to be known about the immune response to helminths. Eosinophils are important for resistance to some helminth species and their recruitment to infected tissues, attachment to parasites and degranulation may all be critical processes for immunity. Complement may contribute to these processes via generation of chemotactic factors (C3a and C5a) or opsonisation of the parasite with C3b/iC3b. The importance of complement during helminth infection is unclear, though complement does promote leukocyte-mediated killing of several helminth species in vitro. The aim of the present study was to investigate the role of complement in immunity of mice to Nippostrongylus brasiliensis, with a focus on whether complement facilitates eosinophildependent resistance to this parasite. A new fluorescence-based method for quantifying in vitro complement deposition and leukocyte adherence on N. brasiliensis was developed. C3 from human serum was deposited on infective-stage L3 via the classical or lectin complement pathways. In contrast, the alternative complement pathway mediated binding of mouse C3 and eosinophil-rich mouse peritoneal leukocytes to L3. Interestingly, the ability of complement and leukocytes to bind to the parasite changed as it matured. Larvae recovered from the skin 30 min post-injection (p.i.) were coated with C3, however those harvested 150 min p.i. exhibited reduced C3 binding capacity. Binding of C3 and eosinophils to larvae recovered from the lungs 24-48 h p.i. (L4) was also diminished compared to that seen on L3. Adult intestinal worms bound C3 and leukocytes only when treated ex vivo with serum and cells. Mice lacking in classical (C1q-deficient), alternative (factor B-deficient) or all complement pathways (C3-deficient) were then employed to determine if complement was important for resistance of mice to N. brasiliensis. IL-5 Tg mice deficient in individual complement genes were generated to assess whether complement contributed to eosinophildependent resistance to the parasite. Factor B deficient mice exhibited impaired C3 deposition on larvae, eosinophil recruitment, eosinophil degranulation and larval aggregation in the skin 30 min p.i. Eosinophil recruitment was similarly abolished by treatment of mice with the C5aR inhibitor PMX53. However at 150 min p.i., larval aggregation, eosinophil and neutrophil recruitment, leukocyte adherence and eosinophil degranulation were largely complement-independent. Ablation of factor B or C3 caused minor but significant increases in lung-larval burden during primary, but not in secondary, infections. Critically, a lack of C3 or factor B in IL-5 Tg mice failed to greatly impair the strong innate anti-parasite resistance typical of these animals, suggesting that eosinophils can provide immunity to N. brasiliensis infection in the absence of complement. This was unexpected, given the evidence from this and previous studies which suggested that in vitro, complement is important for promoting eosinophil-dependent killing of N. brasiliensis and other helminth species. The mechanism(s) by which eosinophils kill N. brasiliensis remain unknown, but may involve the coordination of the complement system with complement-independent factors that act in the early stages of infection. Critically, the influence of complement is limited, because soon after entry into the host, the parasite develops the ability to resist complement activation. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1311182 / Thesis (Ph.D.) -- University of Adelaide, School of Molecular and Biomedical Science, 2008

Host behavior, age, sex, diet, and condition, as well as variation in parasite specificity, drive variation in parasite infection, and ultimately determine the host parasite community. The objectives of this thesis were to 1) examine intraspecific variation in arctic fox parasites, 2) determine relationships between diet and parasites in sympatric arctic and red fox, and 3) compare wolf parasites and diet. Male arctic fox had more cestodes than females and juveniles had more nematodes than adults, likely due to diet and exposure. Red fox carried fewer parasites than arctic fox, likely due to diet, evolved resistance behaviors and higher immune investment, but diet affected cestode abundance in both species. Wolves that ate more white-tailed deer had more cestodes, suggesting increasing deer populations could enhance parasite transmission to moose. However, body condition was unaffected by parasites, suggesting northern canids may have not reached a threshold of infection.

Host behavior, age, sex, diet, and condition, as well as variation in parasite specificity, drive variation in parasite infection, and ultimately determine the host parasite community. The objectives of this thesis were to 1) examine intraspecific variation in arctic fox parasites, 2) determine relationships between diet and parasites in sympatric arctic and red fox, and 3) compare wolf parasites and diet. Male arctic fox had more cestodes than females and juveniles had more nematodes than adults, likely due to diet and exposure. Red fox carried fewer parasites than arctic fox, likely due to diet, evolved resistance behaviors and higher immune investment, but diet affected cestode abundance in both species. Wolves that ate more white-tailed deer had more cestodes, suggesting increasing deer populations could enhance parasite transmission to moose. However, body condition was unaffected by parasites, suggesting northern canids may have not reached a threshold of infection.

The withdrawal of water sources concluded the reuse of treated wastewaters, especially for non-potable purposes. Agricultural use of the reclaimed wastewaters is one of the reuse options. However health considerations of the reuse of reclaimed wastewaters for public related purposes are underestimated, since wastewaters contain a variety of microbial pathogens, which may be transmitted to workers and consumers through the crops irrigated. Of these, parasitic eggs have a special place, as they are capable of surviving in the soil for months or even years, depending on environmental conditions. There is insufficient accumulated information on the health related criteria for the reuse of treated wastewaters in Turkey. The aim of this study was therefore to determine the helminthic eggs in raw sewage and in effluents of ASKi municipal wastewater treatment plant in Ankara. The study involved examining to decide whether these organisms exist in the wastewaters at all, and if so in what concentrations. Modified Bailenger&rsquo / s method, which published in the &ldquo / WHO Laboratory Manual of Parasitological and Bacteriological Techniques&rdquo / and &ldquo / U.S.EPA ICR Microbial Laboratory Manual&rdquo / were used in developing the specific methods used in this study.