Dissertations / Theses on the topic 'Ross River Virus'

Ross River virus (RRV) is a mosquito-borne alphavirus and the aetiological agent of epidemic polyarthritis (EPA). Arthropod borne-Alphaviruses that are related to RRV, such as Chikungunya virus, Sindbis virus and Barmah Forest virus, are usually associated with epidemics of infectious arthritides in different parts of the world. In humans, RRV-induced disease symptoms include fever, rash, myalgia and pain and stiffness of the joints. Muscle and joint pain are the most debilitating symptoms in RRV patients and the best treatment available is non-steroidal anti-inflammatory drugs (NSAID). Previous studies in mice have demonstrated that RRV infection results in inflammation of skeletal muscle and joints and that macrophages play a primary role in disease. The present study was carried out to further elucidate the underlying mechanisms mediating RRV-induced muscle and joint pathology. Previous studies have reported that encephalitic alphaviruses trigger apoptosis of brain cells in mice and that blocking apoptosis reduces mortality rates. In the present study, the ability of RRV to induce muscle apoptosis was investigated in vitro, using a murine myoblast cell line (C1C12), and in vivo, using a mouse model of RRV disease. RRV-infected C1C12 myofibres displayed an array of morphological and biochemical makers of apoptosis. Apoptosis was also observed in the skeletal muscle of RRV-infected C57BL/6J mice. Blocking apoptosis by general caspase inhibition resulted in milder disease symptoms, reduced myofibre damage and decreased inflammation of muscle and joint tissues. The total number of cell infiltrates as well as the number of macrophages infiltrating muscle was significantly reduced by the treatment with a caspase inhibitor. The effects of RRV infection on the skeletal system were also investigated. Primary human osteoblast cells were infected with RRV and monitored for viral-induced cytopathic effect. Osteoblasts supported rapid virus growth and, by 48 hours after infection, succumbed to viral-induced necrosis. In addition, histological examination of bone tissue from RRV-infected C57BL/6J mice showed clear evidence of bone resorption. Tibias from infected mice showed an increased number of activated osteoclasts, a reduction in bone density and thinning of cortical bone. The expression of host factors involved in inflammatory responses and bone remodelling was studied in RRV-infected myofibres and osteoblast cell cultures and in the muscle and joint tissues from infected mice. RRV-infected muscle cells and tissue showed elevated mRNA levels for the chemokines CCL-2, CCL3, CCL5 and CXCL1, all of which are known to mediate the migration of monocytic cells. With the exception of CXCL1, these chemokines were also found to be up-regulated in RRV-infected osteoblast cultures and in joint tissues from infected mice. Muscle and joint tissue from infected mice also showed elevated mRNA levels for type I and type II interferons, TNF- and NOS2. In addition, joint tissues from infected animals contained high levels of IL-6 and IL-1, two cytokines known to mediate bone remodelling. Finally, the therapeutic potential of the drug bindarit was investigated using the mouse model of RRV disease. Bindarit is a known inhibitor of CCL-2 and TNF- and has been found to prevent protein denaturation. Treatment with bindarit resulted in mice developing milder disease symptoms, reduced muscle damage and decreased inflammation of muscle and joint tissues. In particular, bindarit significantly reduced macrophage infiltration into skeletal muscle tissue. This thesis has contributed to the understanding of RRV pathogenesis. It has identified novel mechanisms of RRV-induced muscle and bone pathology and provided further evidence that associate pro-inflammatory host factors to RRV disease. This work has also demonstrated that bindarit should be considered as a candidate for treating RRV disease in humans.

Ross River virus is one of the most severe communicable diseases in Australia. During the 1995/96 outbreak of Ross River virus in south-western Australia, over 1 ,300 human cases were reported. Since the symptoms of the disease are sometimes too weak to be diagnosed, it is important to determine the number of humans who actually contracted the virus during outbreaks. To do this, several mathematical models with different hypotheses are constructed and analysed mathematically. The threshold mathematical conditions of these models suggest that as well as the size of the vector mosquito population, the population size and length of viraemia periods; of host populations and the infection rates between the hosts and vectors play the main roles in the transmission. Several parameters in the transmission are currently unknown, so only simple models of RRV transmission are computer-simulated. Some of the unknown parameters are extrapolated from published studies of other arboviruses. The sensitivities of the models to some of the unknown parameters are also examined. Simulation results indicate the sero-conversion rates and ratios of clinical to subclinical human infections during the outbreaks which occurred in the Peel and Leschenault districts in Southwestern Australia.

[Truncated abstract] Ross River virus (RRV) causes the most common arboviral disease in Australia, with approximately 5000 new cases reported each year, making this virus a major public health concern. The aim of this thesis was to link results from virological, pathogenesis and epidemiological studies to further define RRV disease in the south-west (SW) of Western Australia (WA), a region of endemic and epizootic RRV activity. A crosssectional seroprevalence study was used to show that 7.8 percent of SW communities were seropositive to RRV, comparable to other regions of Australia with similar temperate climates to the SW . . . RRV-specific IgM antibodies were found to persist for at least two years following RRV infection. A murine model was used to conclusively show differences in pathogenesis between RRV genotypes, the SW and northern-eastern (NE) genotypes, which are known to circulate throughout Australia. The SW genotype, unique to the SW of WA induced only poor neutralising antibody production and nonneutralising antibodies after the acute phase of infection. In comparison, the NE genotype which currently predominates in mosquito populations in the SW of WA, induced the most efficient neutralising antibody response and consequently produced the mildest disease in the mouse. These data in the mouse suggest that the infecting genotype will mostly likely influence disease outcome in humans and could at least partially explain why more severe and persistent disease has been reported from the SW of WA. Collectively, results from this thesis provide an important benchmark against which future investigations into BFV and RRV diseases can be measured.

Ross River virus (RRV) is an Alphavirus distributed throughout Australia. It is transmitted by mosquitoes and is known to cause moderate to severe disease symptoms in humans. Along with other alphaviruses such as Sindbis virus and Chikungunya virus, RRV is known to cause arthritic symptoms, characterised by muscle and joint inflammation. Several investigations have established the role of macrophage cells and pro-inflammatory host factors in the development of RRV-induced disease. In this study, we attempted to determine differences between RRV passaged in mammalian and mosquito cells. There is strong evidence that arthropod-borne viruses are able to display enhanced infectivity when passaged into arthropod cell line. We showed that mosquito cell-derived RRV (mos-RRV) was able to replicate to higher titres than mammalian cell-derived RRV. We also showed that mos-RRV failed to induce Type I IFN-associated antiviral responses. The second aim of this study was to investigate the role of TNF-ᬠa pro-inflammatory cytokine implicated in arthritic diseases, in the development of RRV disease. We treated RRV-infected C57BL/6J mice with a commercially available TNF-ᠩnhibitor drug and monitored disease signs. We found that the TNF-ᠩnhibitor does not ameliorate RRV disease (RRVD) symptoms, and that it does not prevent muscle and joint inflammation. We analysed histological sections of muscle and joint tissue of Enbrel-treated and untreated, RRV-infected cells. We also determined and compared host cytokine expression profiles. Finally, we sought to determine the requirement for natural killer (NK) cells in RRV disease. NK cells have been detected in the synovium of RRV-infected patients since early studies, but their role in disease pathogenesis remains unclear. Using a NK-dysfunctional mouse (C57BL/6J-Lystbg), we showed that mice lacking a functional NK system are more susceptible to RRV disease than wildtype, C57BL/6J mice. We monitored disease symptoms following RRV infection and assessed muscle and joint inflammation in Lystbg and C57BL/6J mice. This thesis examines mechanisms of viral infection and immune evasion employed by RRV, as well as into the role of host cells and cytokines in RRVD pathogenesis disease mechanisms. We showed that a functional NK cell system is required for the regulation of RRV-induced muscle and joint inflammation. Our characterisation of the use of a commercial TNF-ᠩnhibitor in RRV-induced disease in mice may provide information on the role of TNF-ᠩn viral arthritis, and may help towards developing safe and effective treatment.

Ross River virus (RRV) belongs to the genus Alphavirus and is a medically important arbovirus that causes musculoskeletal disease in humans with symptoms such as arthralgia, arthritis and myalgia. Disease symptoms consistent with RRV infection were first recorded in 1928 in Australia. Currently, with approximately 5,000 cases of RRV infection reported each year in Australia, RRV is the most widely spread arbovirus throughout the South Pacific region. At present there are no specific therapeutics or vaccines available. RRV disease is treated with analgesics and non-steroidal anti-inflammatory drugs to provide symptomatic relief. Therefore, it is important to investigate RRV disease mechanisms so as to better understand disease pathogenesis, which could lead to identifying potential targets for therapeutic intervention. The host Type I interferon (IFN) system is the primary innate antiviral defence mechanism. The antiviral effects of type I IFN act to both suppress viral replication and modulate innate and adaptive immune responses during viral infection. However, the interplay between the host type I IFN responses and alphavirus infection is currently poorly understood. This thesis focuses on the role of type I IFN system in RRV infection and disease pathogenesis.<br>Thesis (PhD Doctorate)<br>Doctor of Philosophy (PhD)<br>Institute for Glycomics<br>Science, Environment, Engineering and Technology<br>Full Text

The physical and immunological consequences to BALB/c and CBA murine neonates of in utero infection with Ross River virus [RRV] were compared to previously reported teratogenic and tolerogenic properties of other togaviruses. X-ray imaging as a means of determining the incidence, type and extent of teratogenesis induction in neonatal mice lacked sufficient resolution for the determination of structural abnormalities. A previously described histological technique for identifying cartilage and bone defects [Mabruk et al., 1988] was unsatisfactory as disintegration of neonatal material occurred upon storage. This study modified the method to include an inital fixation step in normal buffered formalin prior to Alcian Blue staining, a ten-fold increase in the concentration of the clearing agent and an extension of reagent exposure times. No skin defects nor skeletal deformities, reported in similar studies with related viruses, were noted in the present investigation. Skeletal changes observed following embryonic exposure to RRV at 8 to 18 days gestation were similar to those seen in uninfected control mice. Such changes, which included bones missing or not visible in the extremities of mice, were attributed to incomplete skeletal development resulting from variations both in the murine gestational period [19 to 21 days] and in the collection of neonates post-partum [within 12 hours of birth]. Changes to the structure [malformation] and orientation [misalignment] of the sternebrae, noted in the present investigation, were considered variable but normal representations of ossification centres within the sternum. No difference in the incidence or severity of these anatomical disturbances was observed between uninfected and RRV infected mice in this study. Although the development of six bones within the sternum were expected, 6% [5/83] of neonatal BALB/c mice demonstrated a supernumerary sternebra. This observation was not significant for uninfected and RRV-infected populations. Further, one uninfected CBA mouse presented with only five sternebrae. Consequently, it is concluded that in utero exposure to RRV, strain T48, does not induce teratogenic changes to BALB/c or CBA foetuses. The induction of neonatal immunological tolerance in uninfected mice and mice exposed in utero to RRV was assessed using an indirect enzyme linked immunosorbent assay [ELISA] to quantify the murine serum titres of anti-RRV lgM and lgG subclass antibodies. Further, a virus titration method was utilised to assay murine sera for virus content. Four experimental groups of mice were investigated : U and I defined as uninfected and infected respectively; 0 and M defined as offspring and mother respectively. Serum titres of anti-RRV lgM antibody remained similar or increased significantly during the testing period. In contrast, the serum titres of anti-RRV lgG antibodies were obseNed to significantly increase only in those offspring suckled on uninfected mothers [U0UM and l0UM groups] and to significantly decrease only in those offspring suckled on infected mothers [U0 1M and 10 1M groups]. Further, a significant reduction in the incidence of viraemia and median serum titre of virus was obseNed only in BALB/c mice born to, and subsequently suckled on, uninfected mothers. For both BALB/c and CBA mouse strains, comparisons between groups of mice [U0 UM and 10 UM; U0 1M and 10 1J indicated that the serum titres of anti-RRV lgG subclass and lgM antibodies remained similar or were significantly elevated in the uninfected offspring during the 2 week period following intraperitoneal [i.p.] injection with RRV. Other comparisons [U0UM and U0 1M; 10 UM and 10 1J indicated that although the serum titres of anti-RRV lgG subclass antibodies were similar or significantly elevated in those mice suckled on infected mothers at 1 and 3 days following i.p. injection, this trend was reversed at 14 days post-injection. For these same comparisons, serum titres of RRV-specific lgM antibody remained similar or were significantly higher in those mice suckled on uninfected mothers for the entire testing period. In addition, more offspring born to, and subsequently suckled on, uninfected mothers were viraemic, with significantly higher serum titres of virus, than any other experimental group. Generally, inter-strain comparisons for each experimental group indicated that serum titres of anti-RRV antibodies were either similar in both mouse strains or significantly elevated in CBA mice. Significant variation in the incidence of viraemia and median serum titre of virus was observed only in those mice born to, and subsequently suckled on, uninfected mothers. At 3 days post-injection, more CBA mice were viraemic, with higher serum titres of virus, than BALB/c mice. Post-partum transfer of maternal immunoglobulin moderated the lgG and lgM immune responses in uninfected mice suckled on infected mothers, affirming the importance of maternal antibody in neonatal anti-viral protection. This investigation presents important information on the effects of in utero exposure to RRV in the murine model. A distinct deficiency in lgG production was demonstrated in mice infected in utero with virus then challenged postpartum. Further characterisation of this immunodeficiency indicated that all the lgG pathways in BALB/c mice were affected, while in CBA mice only the lgG1 and lgG3 subclasses were reduced. No correlation between prenatal exposure to RRV and lgM production was evident. These results provide the first evidence for the induction of neonatal immunological tolerance in BALB/c and CBA mice following in utero exposure to RRV. This thesis proposes clonal anergy of B cells, capable of responding to RRV, as the primary mechanism responsible for the immunologically non-responsive state observed in this study. However, the ancillary role of clonal abortion of RRV-specific Band T cell clones in the induction of murine neonatal tolerance cannot be dismissed. The future investigation of specific aspects identified in this study are of considerable importance in vaccine development. The possibility of adverse effects of a live vaccine on an unborn child would greatly affect the decision as to whether a RRV vaccine should be live attenuated or killed.

[Truncated abstract] In an era of emerging and resurging infectious diseases, understanding the ecological processes that influence pathogen activity and the influences of anthropogenic change to those are critical. Ross River virus (RRV, Togoviridae: Alphavirus) is a mosquito-borne zoonosis occurring in Australia with a significant human disease burden. In the southwest of Western Australia (WA) RRV is principally vectored by Aedes camptorhynchus Thomson (Diptera: Culicidae), which is halophilic. The inland southwest, the Wheatbelt region, of WA is substantially affected by an anthropogenic salinisation of agricultural land called dryland salinity, which threatens to influence transmission of this arbovirus. This study assessed the ecological impacts of dryland salinity on mosquitoes, mammalian hosts and their interactions to influence the potential for RRV transmission. Many aquatic insect taxa colonise ephemeral water bodies directly as adults or by oviposition. Using a manipulative experiment and sampling from ephemeral water bodies in the Wheatbelt, I demonstrated that salinity of water bodies can modify colonisation behaviour and the distribution of some organisms across the landscape. Halosensitive fauna selected less saline mesocosms for oviposition and colonisation. In particular, Culex australicus Dobrotworksy and Drummond and Anopheles annulipes Giles (Diptera: Culicidae), potential competitors with Ae. camptorhynchus, avoided ovipostion in saline mesocosms and water bodies in the field. This finding suggests salinity influences behaviour and may reduce interspecific interactions between these taxa and Ae. camptorhynchus at higher salinities. Using extensive field surveys of ephemeral water bodies in the Wheatbelt I found mosquitoes frequently colonised ephemeral water bodies, responded positively to rainfall, and populated smaller water bodies more densely than larger water bodies. The habitat characteristics of ephemeral water bodies changed in association with salinity. Consequently there were both direct and indirect associations between salinity and colonising mosquitoes. Ultimately the structure of mosquito assemblages changed with increasing salinity, favouring an increased regional distribution and abundance of Ae. camptorhynchus. The direct implication of this result is secondary salinisation has enhanced the vectorial potential for RRV transmission in the WA Wheatbelt. ... This thesis contributes to an emerging body of research aimed at delineating important ecological processes which determine transmission of infections disease. Collectively the findings in this study suggest dryland salinity enhances the potential for RRV activity in the Wheatbelt. Currently, human RRV notifications in the Wheatbelt do not reflect the salinity-RRV transmission potential in that area, but appear to be associated with dispersal of RRV from the enzootic coastal zone of southwest WA. I speculate dryland salinity is a determinant of potential for RRV transmission, but not activity. Dryland salinity is predicted to undergo a two to four fold expansion by 2050, which will increase the regional potential for RRV activity. Preservation and restoration of freshwater ecosystems may ameliorate the potential for transmission of RRV and, possibly, human disease incidence.

Insect pests (such as mosquitoes) and their associated impacts have become important social, economic and environmental health issues. Mosquitoes transmit diseases, are widely perceived as a nuisance and are becoming a serious health concern for the public. The incidence of contracting mosquito-transmitted diseases has markedly increased in recent decades in Australia (Russell 1994). Currently, Ross River virus is the most prevalent mosquito-transmitted viral disease in Australia with up to 8,000 cases reported annually (Curran et al. 1996). The absence of documented evidence about the full social costs of mosquito-transmitted diseases is a critical issue as there is currently no economic rationale underlying existing resource allocation for intervention programs. This study of the full social costs of Ross River virus was conducted between April and July 2002. Demographic, health-state and disease-related data were collected using survey questionnaires for 201 notified Ross River virus victims from across Queensland. Two self-administered surveys were conducted at the time approximately onset and six months from onset while a phone survey was conducted at 12 months after onset. Direct impacts such as the costs of health care (medical consultations, pathology services and medicines), non health care resources (treatment-related transport) and indirect impacts such as the opportunity cost of lost productivity (due to disability and treatment-related waiting times) of the viral illness were recorded in the questionnaires and were valued using market prices. A non-market valuation method (willingness-to-pay), in conjunction with a self-assessed standard health measure (Short Form 36) were used to quantify more intangible health-related quality of life effects such as change in physical, mental and social functioning. Estimated full social costs of the disease were analysed across age and gender groups. Based on the mean cost estimates for the study sample, the total disease costs have been extrapolated by local government areas as the appropriate administrative areas. A wide range of social and economic costs of the virus has been addressed in this thesis. However, the derived costs cannot be summed into a total estimate as several of these values overlap in terms of coverage. Therefore, only the major cost components, with a minimum of overlap, have been used to estimate the aggregate social cost of the disease. Given the methodological and empirical limitations of the study, the most accurate estimate of the average per capita full costs of Ross River virus in Queensland is estimated at $AUD (2002) 1,070 per case. The estimate of the full social cost of Ross River virus disease can be a vital input for many relevant policy applications. For example, disease costs together with resource costs of current interventions, are essential inputs for ongoing economic evaluations of mosquito control programs at local level. In this thesis, the cost minimisation approach has been presented to evaluate mosquito control programs at the local level where the policy analyst’s task is to minimise the overall social costs (that is, disease costs plus control costs). These economic evaluations have substantial potential benefits to society in terms of the efficient allocation of scarce resources. In addition, estimated disease cost is a significant input for economic impact assessment of regional disease outbreaks. It also can be used to highlight disease impact upon the economy and community and hence draw attention to the scale and scope of such problems to policy makers at all levels so that they can respond appropriately to the mosquito problem, and mosquito-transmitted diseases, as priority issues in the political agenda.<br>Thesis (PhD Doctorate)<br>Doctor of Philosophy (PhD)<br>Australian School of Environmental Studies<br>Science, Environment, Engineering and Technology<br>Full Text

The incidence of many arboviral diseases is largely associated with social and environmental conditions. Ross River virus (RRV) is the most prevalent arboviral disease in Australia. It has long been recognised that the transmission pattern of RRV is sensitive to socio-ecological factors including climate variation, population movement, mosquito-density and vegetation types. This study aimed to assess the relationships between socio-environmental variability and the transmission of RRV using spatio-temporal analytic methods. Computerised data files of daily RRV disease cases and daily climatic variables in Brisbane, Queensland during 1985-2001 were obtained from the Queensland Department of Health and the Australian Bureau of Meteorology, respectively. Available information on other socio-ecological factors was also collected from relevant government agencies as follows: 1) socio-demographic data from the Australia Bureau of Statistics; 2) information on vegetation (littoral wetlands, ephemeral wetlands, open freshwater, riparian vegetation, melaleuca open forests, wet eucalypt, open forests and other bushland) from Brisbane City Council; 3) tidal activities from the Queensland Department of Transport; and 4) mosquito-density from Brisbane City Council. Principal components analysis (PCA) was used as an exploratory technique for discovering spatial and temporal pattern of RRV distribution. The PCA results show that the first principal component accounted for approximately 57% of the information, which contained the four seasonal rates and loaded highest and positively for autumn. K-means cluster analysis indicates that the seasonality of RRV is characterised by three groups with high, medium and low incidence of disease, and it suggests that there are at least three different disease ecologies. The variation in spatio-temporal patterns of RRV indicates a complex ecology that is unlikely to be explained by a single dominant transmission route across these three groupings. Therefore, there is need to explore socio-economic and environmental determinants of RRV disease at the statistical local area (SLA) level. Spatial distribution analysis and multiple negative binomial regression models were employed to identify the socio-economic and environmental determinants of RRV disease at both the city and local (ie, SLA) levels. The results show that RRV activity was primarily concentrated in the northeast, northwest and southeast areas in Brisbane. The negative binomial regression models reveal that RRV incidence for the whole of the Brisbane area was significantly associated with Southern Oscillation Index (SOI) at a lag of 3 months (Relative Risk (RR): 1.12; 95% confidence interval (CI): 1.06 - 1.17), the proportion of people with lower levels of education (RR: 1.02; 95% CI: 1.01 - 1.03), the proportion of labour workers (RR: 0.97; 95% CI: 0.95 - 1.00) and vegetation density (RR: 1.02; 95% CI: 1.00 - 1.04). However, RRV incidence for high risk areas (ie, SLAs with higher incidence of RRV) was significantly associated with mosquito density (RR: 1.01; 95% CI: 1.00 - 1.01), SOI at a lag of 3 months (RR: 1.48; 95% CI: 1.23 - 1.78), human population density (RR: 3.77; 95% CI: 1.35 - 10.51), the proportion of indigenous population (RR: 0.56; 95% CI: 0.37 - 0.87) and the proportion of overseas visitors (RR: 0.57; 95% CI: 0.35 - 0.92). It is acknowledged that some of these risk factors, while statistically significant, are small in magnitude. However, given the high incidence of RRV, they may still be important in practice. The results of this study suggest that the spatial pattern of RRV disease in Brisbane is determined by a combination of ecological, socio-economic and environmental factors. The possibility of developing an epidemic forecasting system for RRV disease was explored using the multivariate Seasonal Auto-regressive Integrated Moving Average (SARIMA) technique. The results of this study suggest that climatic variability, particularly precipitation, may have played a significant role in the transmission of RRV disease in Brisbane. This finding cannot entirely be explained by confounding factors such as other socio-ecological conditions because they have been unlikely to change dramatically on a monthly time scale in this city over the past two decades. SARIMA models show that monthly precipitation at a lag 2 months (=0.004,p=0.031) was statistically significantly associated with RRV disease. It suggests that there may be 50 more cases a year for an increase of 100 mm precipitation on average in Brisbane. The predictive values in the model were generally consistent with actual values (root-mean-square error (RMSE): 1.96). Therefore, this model may have applications as a decision support tool in disease control and risk-management planning programs in Brisbane. The Polynomial distributed lag (PDL) time series regression models were performed to examine the associations between rainfall, mosquito density and the occurrence of RRV after adjusting for season and auto-correlation. The PDL model was used because rainfall and mosquito density can affect not merely RRV occurring in the same month, but in several subsequent months. The rationale for the use of the PDL technique is that it increases the precision of the estimates. We developed an epidemic forecasting model to predict incidence of RRV disease. The results show that 95% and 85% of the variation in the RRV disease was accounted for by the mosquito density and rainfall, respectively. The predictive values in the model were generally consistent with actual values (RMSE: 1.25). The model diagnosis reveals that the residuals were randomly distributed with no significant auto-correlation. The results of this study suggest that PDL models may be better than SARIMA models (R-square increased and RMSE decreased). The findings of this study may facilitate the development of early warning systems for the control and prevention of this widespread disease. Further analyses were conducted using classification trees to identify major mosquito species of Ross River virus (RRV) transmission and explore the threshold of mosquito density for RRV disease in Brisbane, Australia. The results show that Ochlerotatus vigilax (RR: 1.028; 95% CI: 1.001 - 1.057) and Culex annulirostris (RR: 1.013, 95% CI: 1.003 - 1.023) were significantly associated with RRV disease cycles at a lag of 1 month. The presence of RRV was associated with average monthly mosquito density of 72 Ochlerotatus vigilax and 52 Culex annulirostris per light trap. These results may also have applications as a decision support tool in disease control and risk management planning programs. As RRV has significant impact on population health, industry, and tourism, it is important to develop an epidemic forecast system for this disease. The results of this study show the disease surveillance data can be integrated with social, biological and environmental databases. These data can provide additional input into the development of epidemic forecasting models. These attempts may have significant implications in environmental health decision-making and practices, and may help health authorities determine public health priorities more wisely and use resources more effectively and efficiently.

The incidence of many arboviral diseases is largely associated with social and environmental conditions. Ross River virus (RRV) is the most prevalent arboviral disease in Australia. It has long been recognised that the transmission pattern of RRV is sensitive to socio-ecological factors including climate variation, population movement, mosquito-density and vegetation types. This study aimed to assess the relationships between socio-environmental variability and the transmission of RRV using spatio-temporal analytic methods. Computerised data files of daily RRV disease cases and daily climatic variables in Brisbane, Queensland during 1985-2001 were obtained from the Queensland Department of Health and the Australian Bureau of Meteorology, respectively. Available information on other socio-ecological factors was also collected from relevant government agencies as follows: 1) socio-demographic data from the Australia Bureau of Statistics; 2) information on vegetation (littoral wetlands, ephemeral wetlands, open freshwater, riparian vegetation, melaleuca open forests, wet eucalypt, open forests and other bushland) from Brisbane City Council; 3) tidal activities from the Queensland Department of Transport; and 4) mosquito-density from Brisbane City Council. Principal components analysis (PCA) was used as an exploratory technique for discovering spatial and temporal pattern of RRV distribution. The PCA results show that the first principal component accounted for approximately 57% of the information, which contained the four seasonal rates and loaded highest and positively for autumn. K-means cluster analysis indicates that the seasonality of RRV is characterised by three groups with high, medium and low incidence of disease, and it suggests that there are at least three different disease ecologies. The variation in spatio-temporal patterns of RRV indicates a complex ecology that is unlikely to be explained by a single dominant transmission route across these three groupings. Therefore, there is need to explore socio-economic and environmental determinants of RRV disease at the statistical local area (SLA) level. Spatial distribution analysis and multiple negative binomial regression models were employed to identify the socio-economic and environmental determinants of RRV disease at both the city and local (ie, SLA) levels. The results show that RRV activity was primarily concentrated in the northeast, northwest and southeast areas in Brisbane. The negative binomial regression models reveal that RRV incidence for the whole of the Brisbane area was significantly associated with Southern Oscillation Index (SOI) at a lag of 3 months (Relative Risk (RR): 1.12; 95% confidence interval (CI): 1.06 - 1.17), the proportion of people with lower levels of education (RR: 1.02; 95% CI: 1.01 - 1.03), the proportion of labour workers (RR: 0.97; 95% CI: 0.95 - 1.00) and vegetation density (RR: 1.02; 95% CI: 1.00 - 1.04). However, RRV incidence for high risk areas (ie, SLAs with higher incidence of RRV) was significantly associated with mosquito density (RR: 1.01; 95% CI: 1.00 - 1.01), SOI at a lag of 3 months (RR: 1.48; 95% CI: 1.23 - 1.78), human population density (RR: 3.77; 95% CI: 1.35 - 10.51), the proportion of indigenous population (RR: 0.56; 95% CI: 0.37 - 0.87) and the proportion of overseas visitors (RR: 0.57; 95% CI: 0.35 - 0.92). It is acknowledged that some of these risk factors, while statistically significant, are small in magnitude. However, given the high incidence of RRV, they may still be important in practice. The results of this study suggest that the spatial pattern of RRV disease in Brisbane is determined by a combination of ecological, socio-economic and environmental factors. The possibility of developing an epidemic forecasting system for RRV disease was explored using the multivariate Seasonal Auto-regressive Integrated Moving Average (SARIMA) technique. The results of this study suggest that climatic variability, particularly precipitation, may have played a significant role in the transmission of RRV disease in Brisbane. This finding cannot entirely be explained by confounding factors such as other socio-ecological conditions because they have been unlikely to change dramatically on a monthly time scale in this city over the past two decades. SARIMA models show that monthly precipitation at a lag 2 months (=0.004,p=0.031) was statistically significantly associated with RRV disease. It suggests that there may be 50 more cases a year for an increase of 100 mm precipitation on average in Brisbane. The predictive values in the model were generally consistent with actual values (root-mean-square error (RMSE): 1.96). Therefore, this model may have applications as a decision support tool in disease control and risk-management planning programs in Brisbane. The Polynomial distributed lag (PDL) time series regression models were performed to examine the associations between rainfall, mosquito density and the occurrence of RRV after adjusting for season and auto-correlation. The PDL model was used because rainfall and mosquito density can affect not merely RRV occurring in the same month, but in several subsequent months. The rationale for the use of the PDL technique is that it increases the precision of the estimates. We developed an epidemic forecasting model to predict incidence of RRV disease. The results show that 95% and 85% of the variation in the RRV disease was accounted for by the mosquito density and rainfall, respectively. The predictive values in the model were generally consistent with actual values (RMSE: 1.25). The model diagnosis reveals that the residuals were randomly distributed with no significant auto-correlation. The results of this study suggest that PDL models may be better than SARIMA models (R-square increased and RMSE decreased). The findings of this study may facilitate the development of early warning systems for the control and prevention of this widespread disease. Further analyses were conducted using classification trees to identify major mosquito species of Ross River virus (RRV) transmission and explore the threshold of mosquito density for RRV disease in Brisbane, Australia. The results show that Ochlerotatus vigilax (RR: 1.028; 95% CI: 1.001 - 1.057) and Culex annulirostris (RR: 1.013, 95% CI: 1.003 - 1.023) were significantly associated with RRV disease cycles at a lag of 1 month. The presence of RRV was associated with average monthly mosquito density of 72 Ochlerotatus vigilax and 52 Culex annulirostris per light trap. These results may also have applications as a decision support tool in disease control and risk management planning programs. As RRV has significant impact on population health, industry, and tourism, it is important to develop an epidemic forecast system for this disease. The results of this study show the disease surveillance data can be integrated with social, biological and environmental databases. These data can provide additional input into the development of epidemic forecasting models. These attempts may have significant implications in environmental health decision-making and practices, and may help health authorities determine public health priorities more wisely and use resources more effectively and efficiently.

Arthritogenic alphaviruses, such as Ross River virus (RRV) are associated with worldwide outbreaks of human polyarthritis/arthralgia. The pathogenesis of RRV and other alphaviruses is poorly understood. Studies have shown potential links between the different strains of RRV and variation in their pathogenesis and virulence. Currently there is believed to be two circulating strains of RRV, the south western (SW) from the south west region of Western Australia and the north eastern (NE) from the east coast of Australia. Studies have suggested that the persistence of RRV may be the result of an impaired immune response. This study was designed to determine if the SW and NE isolates of RRV have the ability to induce apoptosis in DCs and fibroblasts and discover any possible variation in their apoptosis-inducing capacity. Both Vero cells and murine bone marrow DCs (BMDCs) were infected with the SW74249 (SW) and SW82627 (NE) strains of RRV. A time course analysis of two apoptotic markers and a cell viability marker for both cell types was conducted by flow cytometry. The results indicate RRV- induced apoptosis in both Vero cells and BMDCs, with RRV inducing a stronger pro-apoptotic response in BMDCs than Vero cells, 24 h after infection. Between the two strains there was little variation in the Vero cells over time. In the BMDCs there was some variation with the RRV-SW strain inducing a higher percentage of cell death than the RRV-NE strain, 24 h after infection. Collectively, the data indicates that RRV has the capacity to induce a pro-apoptotic response in DCs, with the SW presenting as more aggressive compared to the NE, potentially leading to greater virulence. This data could help to explain the mechanism of RRV persistence in vertebrate hosts, as well as the reported differences in severity and duration of human clinical symptoms. Immunotherapy aimed at correcting the patient’s dysfunctional immune system, may represent a new strategy for the successful medical treatment of RRV infection.

Arthritogenic alphaviruses such as Ross River virus (RRV) and Chikungunya virus (CHIKV) are mosquito-borne viruses that have been responsible for outbreaks of arthritic disease in Australia and worldwide, respectively. Characteristic disease of arthritogenic alphaviruses include the development of joint inflammation and arthritic disease which can last months to years after infections. Arthritic disease caused by RRV and CHIKV shares similar characteristics with rheumatoid arthritis (RA), such as the development of incapacitating, joint inflammation, as well as the upregulation of proinflammatory mediators. While most studies, using experimental animal models of viral infection, have shown a role played by innate immune cells in alphaviral inflammation, the involvement of adaptive immune cells remains poorly understood. Studies investigating RA mouse models and RA patients have identified distinct subsets of T lymphocytes as important contributors in joint inflammation and disease. In particular, CD4+ T cells that produce IL-17 act by upregulating proinflammatory cytokines promoting inflammatory cell recruitment to the joints. The role of IL-17-producing cells in alphavirus-induced arthritis has not been investigated, we therefore aimed to investigate the role of IL-17+ cell subsets in a mouse model of RRV-induced joint inflammation. Mice were infected with RRV and ankle joints were harvested at the peak of inflammation (10 days post-infection). Flow cytometry analysis revealed the presence of IL-17A+ and IL-17F+ cell subsets produced by CD4+ and CD8+ T cells in the joints of infected mice. Further, gene expression analysis and protein analysis of joint tissue homogenates revealed elevated mRNA expression and protein levels of IL-17A at 10 days post-infection (dpi) in RRV-infected mice. In addition, RRV-infected joints showed a significant increase in pro-inflammatory cytokines such as IFN-γ, TNF and IL-6. We next sought to determine whether in vivo neutralisation of IL-17 could reduce joint inflammation in RRV-infected mice. Using a monoclonal antibody targeting IL-17A, we found that antibody-treated mice showed a reduction in disease severity from 8-10 dpi, compared to untreated RRV-infected mice. Blocking IL-17A had no significant effect on cell subsets in joint infiltrates, and did not reduce RRV viral load in joint tissue. However, protein analysis showed a significant reduction in IFN-γ in antibody-treated mice compared to untreated mice. This indicates that although blockade of IL-17A did not have an effect on infiltrating cells or viral load, disease severity was substantially reduced. Finally, we also showed that targeting the IL-17A/F heterodimer using a specific anti-IL-17A/F antibody reduced disease severity, indicating that both isoforms of IL-17 may play a role in RRVD pathogenesis. In this study, we characterise for the first time two distinct subsets of IL-17-producing cells in an animal model of alphavirus-induced joint inflammation. In addition, we show that inhibiting IL-17A, as well as IL-17A/F heterodimers in vivo prior to the onset of RRV disease can help alleviate joint inflammation in RRV-infected mice. Thus, we provide evidence that IL-17+ T cell subsets can contribute to joint inflammation in RRV infection, and that targeting IL-17 is likely to be a promising approach in the development of future therapies to treat alphaviral infections.<br>Thesis (Masters)<br>Master of Medical Research (MMedRes)<br>School of Medical Science<br>Griffith Health<br>Full Text

Arboviruses contribute a significant burden to human and animal health. Circulation of arboviruses comprises three components; blood-sucking arthropods, vertebrate hosts, and viruses that can infect vertebrates and invertebrates. Interaction of these components is dependent on ecological factors (such as species distributions and climate), epidemiological factors (including vector and host immunity) and behavioural determinants (such as vector feeding host preference or host defensive behaviours). Identifying these drivers of disease emergence can be complicated but informs efforts to mitigate on-going circulation. This dissertation generates new perspectives on the transmission dynamics of Australia’s most common arbovirus, Ross River virus (RRV), with a particular focus on non-human reservoirs. Specifically, I (a) critically analysed current and historic knowledge for non-human reservoirs and vector feeding patterns, (b) examined the natural exposure of RRV in human, free-living and domestic vertebrate populations, and (c) assessed the vertebrate and vector community ecology across areas of varying human notification rates. I began by undertaking a systematic literature review (Chapter 2) assessing evidence for non-human reservoirs of RRV. This chapter synthesised published serological, virus isolation and experimental infection studies in light of the long-held dogma that marsupials are the primary reservoir of RRV. A key finding of this chapter was emerging evidence that placental mammals and birds were also capable of transmitting RRV to mosquito vectors, suggesting a broader reservoir potential than marsupials alone. To further assess the current and historic knowledge, a meta-analysis was performed on mosquito blood meal analysis studies (Chapter 3). It was evident from this chapter that Australian mosquitoes have highly varied feeding patterns which did not reflect their taxonomic classification or larval ecology. To understand the natural exposure of RRV in vertebrate populations I used both existing literature and performed serological surveys. Although humans are largely thought to be RRV dead-end hosts (species which are incapable of pathogen amplification), circulation of the virus in human populations is well documented and provides insights into spatial-temporal patterns of transmission relevant to the understanding of non-human reservoirs. In Chapter 4, I assessed spatial-temporal patterns of seroprevalence in human populations reported from across the natural distribution of RRV in Australia and the Pacific Island Countries from 1958 to present day. A key finding was that RRV circulated in human populations at least since 1975 when human seropositivity between 20 and 34% was reported in Indonesia and Papua New Guinea. This is important because these countries have different vertebrate fauna to Australia, suggesting different transmission cycles. I then assessed RRV exposure in non-human vertebrate species, focussing on South East Queensland, a RRV endemic area (Chapter 5). Samples were collected through a network of veterinary clinics over a 12-month period, and a total 595 samples from 31 species were obtained. The results showed that taxonomic relatedness is not an important determinant for seropositivity, but rather the ecology and physiology of a species including diet, body size and longevity is most important for exposure to RRV. This study not only tests the greatest diversity of vertebrate species in a single RRV seroprevalence study, but is also novel because it provides methodological advances for analysing seroprevalence data for other zoonotic pathogens. In my last data chapter (Chapter 6), I assessed the vertebrate-vector communities across sites with varied RRV human notification rates. Field surveys assessing ‘abundance’ and ‘diversity’ in light of human notifications were undertaken for six months. Human notifications were positively correlated with vertebrate biomass and total mosquito abundance. Although informative in highlighting variables for ongoing investigations, the data from this chapter was insufficient to determine whether this pattern was unique to the specific habitats in which the data was collected, or a true relationship between disease in humans and non-human reservoirs. Nevertheless, this result lends support to the complex transmission dynamics of RRV and the need to apply multi-disciplinary approaches to understand transmission. Throughout the dissertation, I present compelling new evidence that species other than marsupials are involved in the transmission ecology of RRV. My analyses support the notion that transmission is locally dependent on the availability, diversity and ecology of vertebrate hosts, in combination with the diversity and abundance of mosquito vectors. Moreover, by uncovering the feeding patterns of vectors, identifying exposure rates of vertebrate species, in combination with abundance and availability of both vectors and vertebrates, the research in this dissertation provides much of the information required to build a next-generation matrix model for RRV. Throughout the dissertation I have addressed my research aims and present important new methodological techniques (such as combining serological and ecological analyses) for assessing vertebrate reservoirs of arboviruses, whilst carrying out investigations of disease ecology.<br>Thesis (PhD Doctorate)<br>Doctor of Philosophy (PhD)<br>School of Environment and Sc<br>Science, Environment, Engineering and Technology<br>Full Text

Arthritogenic alphaviruses such as chikungunya virus (CHIKV) and Ross River virus (RRV) are transmitted by mosquito vectors, and can cause musculoskeletal disease manifestations such as, excruciating pain and inflammation in joints and the surrounding muscle tissues, in infected individuals. Despite the recent findings of cartilage damage, erosion and focal necrosis in murine models of both chikungunya viral disease (CHIKVD) and Ross River viral disease (RRVD), the role of human chondrocyte/cartilage in alphaviral infection has never been previously explored. In this thesis, it was shown that, both the lower and the higher density cultures (high-density cell cultures and micromass pellet cultures) of primary chondrocytes, supported the RRV-T48 infection and produced the upregulation of soluble inflammatory host factors during the infection. Interestingly, the heterogeneity of chondrocyte infectibility was also noted between two human chondrocyte donors. Chondrocytes that were derived from one patient could not sustain an efficient RRV-T48 replication when compared to chondrocytes that were isolated from another patient. Although this infection profile was surprising, the sample pool for the donor chondrocytes was extremely small thus, other confounding factors may have also contributed to the difference in infection. Additionally, the presence and co-localisation of RRV antigens within the chondrocytes of murine joints, proved that RRV could get to the cartilage and either cause damage directly or trigger the body systems to potentiate the damage. The second half of the thesis focused on exploring a new class of heparan sulfate (HS)-modulating compounds, as targeted treatments for arthritogenic alphaviral infection/diseases. Current drugs for alphaviral diseases are non-specific and include the use of non-steroidal anti-inflammatories which only provide temporary or partial relief in patients. Recently pentosan polysulfate (PPS), a sulfated polysaccharide and a HSmimetic, was found to have reduced the recruitment of inflammatory infiltrates and to have protected the cartilage matrix from degradation in RRV-infected mice. Following which, in this thesis, the roles of other HS-modulating compounds such as polynuclear platinum complexes (PPCs) and HS-mimetic PG545, were investigated for the treatment and/or inhibition of arthritogenic infection. The HS-modulating compounds were characterised for 50% inhibitory doses, in addition to strength of association studies that were performed using the surface plasmon resonance (SPR) and the isothermal titration calorimetry (ITC) binding methods. Both the high signal responses generated in the SPR and the ITC, demonstrated that PG545 had a direct potent inhibitory effect on the RRV virus particles. Therefore, PG545 was further evaluated in the well-established murine model of acute RRVD, and in in-vitro systems. Increasing concentrations of the compounds were able to inhibit plaque formation in RRV, Barmah Forest virus (BFV), and in both the East/Central/South African (ECSA) and the Asian CHIKV strains, as determined by the 50% plaque inhibitory assays. In mice, disease state and compound efficacy were primarily characterised by assessing hind limb weakness, expression levels of soluble host factors, the components of the cartilage matrix, viral titre and histopathology. Prophylactic, standard treatments of PG545 to RRV-infected mice had significantly reduced viral loads in muscles and joints corresponding to a reduction in their clinical scores of limb weakness and immune infiltrate recruitment, at 10 day-post infection (dpi). Also, at peak disease (10dpi), RRV-infected PG545-treated mice had lower expression levels of host factors, IL-6 and CCL-2. Furthermore, PG545 treatment also demonstrated a reduction in the matrix degrading enzyme heparanase (HPSE) in both muscle and joint samples. Serum biochemistries also showed that PG545 treatment in RRV-infected mice, reduced RRV-induced elevations of aspartate aminotransferase (AST) and alanine aminotransferase (ALT). It was also noted that, the standard dosing of PG545 only caused minimal lymphoid tissue anomaly in mice, while the prolonged PG545 dosage (to shorten clinical disease) resulted in both transient hepatosplenomegaly and anaemia. Moreover, both the histopathology and gene expression data, demonstrated that the splenomegaly was due to the expansion of the red pulp and the atrophy of the white pulp. It should also be noted that the long half-life of PG545 (as previously published), could have also contributed to the drug-induced transient pathogenesis, seen in mice that were treated with PG545 for a prolonged duration. Taken together, these findings suggest that although PG545 has a direct inhibitory effect on both RRV-infection as well as the RRV-induced inflammatory disease in host organisms, careful dosing needs to be observed due to the stability and long half- life of compound.<br>Thesis (PhD Doctorate)<br>Doctor of Philosophy (PhD)<br>Institute for Glycomics<br>Griffith Health<br>Full Text

This research investigated the epidemiology and ecology of two mosquito-borne viruses of public health importance in the Asia-Pacific: dengue and Ross River viruses. Patterns of disease caused by these two viruses were analysed in space and time across rural and urban settings, and some of the influencing demographic, geographic and environmental factors were explored. The outcomes of this research included increased understanding of the complex influences on mosquito-borne virus transmission in different human habitats, and the development of unique field and lab-based approaches for studying how the interactions of mosquitoes, wildlife, humans and their environments can cause disease.

In this study, I have investigated the relationship between mosquito control and mosquito-borne disease rates within Queensland, Australia. The thesis considers the most prevalent arbovirus causing human disease in Australia (Ross River virus) and estimates, how much Ross River virus disease is avoided through local government mosquito control in Queensland, and then compares the monetary value of avoided Ross River virus disease with the financial costs of local government mosquito control. A survey to collect information about mosquito control costs and practices was designed and implemented in each of the 125 local governments in Queensland. This survey collated previously dispersed information; because, although local governments in Queensland are legally obliged to perform mosquito control for disease prevention and nuisance reduction, there is no formal or regular reporting of mosquito control costs and practices to the State. A substantive conclusion from this research is that mosquito control has resulted in lower Ross River virus disease notifications in some local government areas. Ross River virus disease notifications are consistently lower in local government areas that implement mosquito control programs that pre-empt mosquito outbreaks using routine surveillance and then reduce mosquito abundance using mosquito control. Furthermore, there is evidence that local governments using extensive freshwater mosquito control, in addition to saltwater mosquito control, have relatively lower annual Ross River virus disease rates and lower standard deviations of the annual Ross River virus disease rates (indicating the freshwater mosquito control is important in suppressing outbreaks of Ross River virus disease). In contrast, mosquito control practices in the inland local government areas tend to be reactive to community complaints of mosquito abundance causing nuisance, and generally include ad-hoc mosquito control treatments. There is no evidence that reactive, adhoc mosquito control programs result in reduced Ross River virus disease notifications. The numbers of avoided Ross River virus notifications were estimated for the local governments that are located in the south eastern coastal region of Queensland. It has been estimated that an annual average of 2206 Ross River virus disease notifications have been avoided through effective mosquito control; and, for each actual notification of Ross River virus disease in the southern coastal local governments, two notifications have been avoided. The survey revealed that in excess of $10 million was spend by local governments implementing mosquito control in Queensland in 2004. The majority of this expenditure occurs in the more densely populated local governments located in the southern coastal strip of the state. A comparison of the financial costs of mosquito control and the financial value of avoided disease produced a cost-benefit ratio of 0.37, meaning that on average, 37% of the costs of mosquito control are directly recouped through the value of avoided Ross River virus disease. In years when the risk of Ross River virus outbreaks is relatively low, due to below average rainfall, the costs of mosquito control exceed the value of avoided Ross River virus notifications—but in years where the risk of an epidemic of Ross River virus is high, effective mosquito control practices can avoid an epidemic of Ross River virus disease, and in this situation the financial value of avoided disease exceeds the costs of the mosquito control program.<br>Thesis (PhD Doctorate)<br>Doctor of Philosophy (PhD)<br>Griffith School of Environment<br>Science, Environment, Engineering and Technology<br>Full Text

[Truncated abstract] A functional ecosystem is increasingly being recognised as a requirement for health and well being of resident human populations. Clearing of native vegetation for agriculture has left 1.047 million hectares of south-west Western Australia affected by a severe form of environmental degradation, dryland salinity, characterised by secondary soil salinisation and waterlogging. This area may expand by a further 1.7-3.4 million hectares if current trends continue. Ecosystems in saline affected regions display many of the classic characteristics of Ecosystem Distress Syndrome (EDS). One outcome of EDS that has not yet been investigated in relation to dryland salinity is adverse human health implications. This thesis focuses on one such potential adverse health outcome: increased incidence of Ross River virus (RRV), the most common mosquito-borne disease in Australia. Spatial analysis of RRV notifications did not reveal a significant association with dryland salinity. To overcome inherent limitations with notification data, serological RRV antibody prevalence was also investigated, and again no significant association with dryland salinity was detected. However, the spatial scale imposed limited the sensitivity of both studies. ... This thesis represents the first attempt to prospectively investigate the influence of secondary soil salinity on mosquito-borne disease by combining entomological, environmental and epidemiological data. The evidence collected indicates that RRV disease incidence is not currently a significant population health priority in areas affected by dryland salinity despite the dominant presence of Ae. camptorhynchus. Potential limiting factors include; local climatic impact on the seasonal mosquito population dynamics; vertebrate host distribution and feeding behaviour of Ae. camptorhynchus; and the scarce and uneven human population distribution across the region. However, the potential for increased disease risk in dryland salinity affected areas to become apparent in the future cannot be discounted, particularly in light of the increasing extent predicted to develop over coming decades before any benefits of amelioration strategies are observed. Finally, it is important to note that both dryland salinity and salinity induced by irrigation are important forms of environmental degradation in arid and semi-arid worldwide, with a total population of over 400 million people. Potential health risks will of course vary widely across different regions depending on a range of factors specific to the local region and the complex interactions between them. It is therefore not possible to make broad generalisations. The need is highlighted for similar research in other regions and it is contended that an ecosystem health framework provides the necessary basis for such investigations.

The current situation of global warming has a serious impact on the control of arthropod-borne infectious diseases. Climate change has led to an increase in conducive breeding habitats for mosquitoes. This change in climate has contributed to the widespread distribution of mosquito-transmitted arthritogenic alphaviruses such as Ross River virus (RRV) and chikungunya virus (CHIKV). RRV is currently endemic to Australia and Papua New Guinea, while CHIKV causes global outbreaks. The recent CHIKV outbreak in the Americas has taken the world by surprise and has affected more than 1 million individuals as of early 2015. To date, there are no specific therapeutics strategies available for the treatment of alphaviral diseases, largely due to the ill-defined innate immune responses elicited by these viruses. This thesis describes new insights into the alphavirus-elicited immune response at the cellular and molecular levels using in vitro, in vivo and ex vivo experimental approaches. In these studies, we identified the differential role of macrophages in modulating the RRV disease progression through an atypical IL-10-dependent M1/M2 polarisation of inflammatory monocytes. RRV-induced differentiation of M1 macrophages triggered the pathogenic inflammatory processes giving rise to the onset of disease, while M2 macrophages were shown to play a protective role.<br>Thesis (PhD Doctorate)<br>Doctor of Philosophy (PhD)<br>Institute for Glycomics<br>Science, Environment, Engineering and Technology<br>Full Text

Mosquito-borne diseases are responsible for significant human morbidity and mortality throughout the world. Current vector control strategies have been impeded by mosquitoes acquiring resistance to insecticide. Therefore, development of new vector control strategies is urgently needed to complement current strategies. In this thesis, efforts have focused on characterizing the glycan-lectin interactions of Ross River virus (RRV; Togaviridae: Alphavirus) with their mosquito vectors. RRV is the most common arbovirus in Australia that causes clinical manifestations including arthralgia and myalgia. Many studies have shown the importance of viral surface glycans in mediating viral entry into host cells. Moreover, the viral surface glycans varies depending on the cells that they replicate in and this variation can affect the infectivity of virus. However, gaps remain in the role of viral glycans in virus host cell recognition. In Chapter 2, the surface glycans of RRV derived from two different cell lines, C6/36 (Ae. albopictus) and Vero (African green monkey kidney) were characterized using lectin array. Lectin array data revealed that RRV derived from two different cell lines exhibited similar glycan profiles. The glycan structures present on the surface of RRV are mannose, galactose, N-acetylglucosamine (GlcNAc) and N-acetylgalactosamine (GalNAc). To investigate the importance of these viral surface glycans in mediating viral entry into host cells, six lectins targeting these glycan structures were assessed for their ability to bind and block RRV entry into host cells. Of these lectins, two mannose-binding lectins, GNA and ConA, showed inhibition of RRV entry into C6/36 and Vero cells. These results suggest the potential use of mannose-binding lectins to block RRV transmission by mosquitoes. It is well known that cell surface glycans or lectins play important role in viral entry. Therefore, comprehensive surface glycan profiles and carbohydrate-binding characteristics of lectins from five mosquito cell lines were established in Chapter 3. Using lectin and glycan arrays, our results showed differences between the glycan structures and carbohydrate-binding characteristics of mosquito cell lines. In particular, complex-type glycans were detected on the cell surface of Ae. albopictus and An. gambiae. The presence of complex-type glycans as authentic constituents of insect glycans is still controversial and this is an important finding as complex-type glycans play diverse roles in regulation of biological functions. Zika virus (ZIKV) is primarily transmitted by the Aedes (Ae) mosquito, with Ae. aegypti and Ae. albopictus being the primary vectors. However, controversial findings on the potential of other mosquito species belonging to the genera of Anopheles (An) and Culex (Cx) have been reported. To date, the rationale underlying the specificity of ZIKV in infecting Aedes mosquitoes remain to be an unaddressed issue. In Chapter 4, we seek to characterize the susceptibility of seven cell lines derived from Ae, An and Cx mosquitoes towards ZIKV infection. Indeed, Ae cell lines were permissive to ZIKV infection and supported viral replication up to seven days post infection, while cells lines from An and Cx mosquitoes were unable to support replication. To specifically address if non-susceptible cell lines were due to the incompetence of ZIKV in establishing viral entry, a pseudoZIKV replicon system was utilized. Interestingly, while all Ae cell lines were highly susceptible to pseudoZIKV infection, the non-susceptible An. gambiae cell line (4a-3B) was also highly permissive to pseudoZIKV entry, in contrast to other An and Cx cell lines tested. Therefore, to identify the host factors involved in ZIKV replication in mosquito cells, RNA sequencing (RNAseq) analysis was performed on ZIKV-susceptible Ae and non-susceptible An cells after infection. Through comparative transcriptomics approach, we observed a differential regulation of attacin, an antimicrobial peptide (AMP) that may potentially play a critical role in modulating ZIKV replication in mosquito cells. Further investigations on the expression profile of different classes of AMPs including attacin, cecropin, defensin, diptericin and gambicin in Aedes; and attacin, cecropin, defensin and gambicin in Anopheles cells by qRT-PCR demonstrated that these AMPs were differentially regulated in ZIKV-susceptible and -resistant mosquito cells. These results suggest that the innate immunity may have a role to influence mosquito vector competence. Species specificity often relies on a specific interaction between a virus ligand and its host cell receptor. Therefore, expression levels of the receptor largely determine the tropisms of viruses. In Chapter 5, we constructed a representative cDNA library from the ZIKV-susceptible Ae. aegypti Aag-2 cell line using the SMART (Switching Mechanism At 5’ end of RNA Template) cDNA synthesis technology. This library will be a useful tool to provide genetic resources for many applications using the proposed strategy including identification of the cellular surface receptors for ZIKV viral entry and host factors required for sustained ZIKV replication in Aedes cells. Overall, this thesis provides in depth investigations into the glycan-lectin interactions between RRV and their mosquito vectors that may affect vector competence. Our study also identified the potential antiviral lectins that can block virus transmission. Finally, our study shed insights into the host factors involved in modulating ZIKV replication, providing a molecular platform for the future development of effective vector control and to evaluate the risk of emergence of a new vector for mosquito-transmitted viruses.<br>Thesis (PhD Doctorate)<br>Doctor of Philosophy (PhD)<br>Institute for Glycomics<br>Griffith Health<br>Full Text

Les huiles essentielles de citronnelle (Cymbopogon citratus), de géranium (Pelargonium graveolens) et de vétiver (Vetiveria zizanioides) sont utilisées partout dans le monde pour leur activité répulsive contre les principaux vecteurs (moustiques, tiques) de maladies infectieuses chez l'Homme (paludisme, chikungunya, …). L'application cutanée de ces produits naturels pour éviter le contact avec un vecteur n'avait pas été encore envisagée comme moyen de limiter les premiers stades de l’infection par l'agent pathogène transmis par le vecteur. Pour vérifier cette hypothèse, les travaux ont été consacrés à la mise en place d'un cadre structuré pour la réévaluation chimique et biologique des trois huiles essentielles sur le modèle du virus du Ross River (alphavirus) de la même famille que le virus du Chikungunya. La caractérisation chimique des huiles essentielles avec une technique de haute résolution (GC×GC/TOF-MS) a permis d'établir leur profil chémotypique précis. L'utilisation de marqueurs spécifiques (clones moléculaires du virus) a permis d'établir l'inhibition de la réplication virale en fonction des conditions d'application des huiles essentielles de géranium et citronnelle. Ces résultats suggèrent l'intérêt d’une huile essentielle répulsive dans les premiers stades d'une infection par un vecteur. À ce titre, l'étude comparative établit la haute valeur ajoutée de l'huile essentielle de géranium et oriente la recherche de nouveaux anti-infectieux naturels vers des complexes riches en monoterpènes<br>Essential oils of citronella (Cymbopogon citratus), geranium (Pelargonium graveolens) and vetiver (Vetiveria zizanioides) are used worldwide as topical repellent against the main vectors (mosquitoes, ticks) of human infectious diseases (Malaria, chikungunya, …). Skin treatment with these natural products, initially to avoid contact with the vector had not yet been considered as a way to disrupt the early stages of infection when the repelling action fails. To check this hypothesis, a structured framework has been performed for the chemical and biological re-evaluation of the three essential oils. The latter was tested against Ross River virus (alphavirus) that belongs to the same family of Chikungunya virus. Analysis of essential oils using a high-resolution technique (GC × GC / TOF-MS) resulted in a more accurate chemotypical profile of the local production. The use of specific markers (molecular clones of the virus, Saclay CEA) allowed to establish the inhibition of viral replication depending of the conditions of geranium and citronella essential oils application. These results suggest the great interest of an essential oil topical repellent in the early stages of a vector infection. The comparative study established the high value of geranium essential oil and gave future direction to the discovery of new anti-infectious solutions from monoterpenes-rich natural complexes

This PhD thesis combines four chapters on different fields of basic research and sets the focus on two circulating viruses of global concern, the orthomyxovirus influenza A virus (IAV) and the alphavirus Ross River virus (RRV). The first three chapters include swine influenza A virus (sIAV) surveillance for the detection and characterisation of IAV subtypes, an in vitro high throughput screening (HTS) on host micro RNAs (miRNAs) for the discovery of novel anti-IAV (H7N9) targets and their underlying mechanisms, and an approach to reduce disease pathogenesis in mice infected with H7N9 by targeting the pro-inflammatory factor CCL2. In a fourth chapter, drug repurposing with the interleukin-1 (IL-1) inhibitor anakinra was investigated to treat RRV-induced bone loss in mice. By combining these four chapters, a broad range of drug discovery is covered in this PhD thesis; Surveillance, HTS target discovery and the application of drug repurposing in animal models of viral diseases.<br>Thesis (PhD Doctorate)<br>Doctor of Philosophy (PhD)<br>Institute for Glycomics<br>Science, Environment, Engineering and Technology<br>Full Text

Arthritogenic alphaviruses such as Ross River virus (RRV) and chikungunya (CHIKV), Sindbis-like viruses (SINV), Barmah Forest virus (BFV), ) Mayaro virus (MAYV) and o’nyong-nyong virus (ONNV) are responsible for outbreaks of debilitating rheumatic joint disease during infection. CHIKV is typically notorious in central Africa, India, South-East Asia and Europe, while other alphaviruses such as RRV and BFV are endemic to Australia. In Australia, there are approximately 4000 cases of RRV reported annually, while the number of CHIKV cases continues to rise in epidemic regions. In 2014, CHIKV invaded several Caribbean islands in the Americas, with a current estimated 1.1 million autochthonous transmission cases as of 16th January 2015. During disease outbreak, more than 95% of patients experience intense persisting pain, generally in knees and the joints of extremeties. Despite the high occurrence rheumatic joint symptoms during alphavirus infection, there are limited studies on the skeletal pathologies due to the lack of routine clinical surveillance. Hence, very little is known about the impact of alphavirus infection on the bone remodelling process and warrants further investigation.<br>Thesis (PhD Doctorate)<br>Doctor of Philosophy (PhD)<br>Institute for Glycomics<br>Science, Environment, Engineering and Technology<br>Full Text

Understanding the epidemiological mechanisms that drive disease incidence and forecasting incidence are major goals in public health. This is particularly relevant for mosquito-borne pathogens that cause human disease burden. In terms of incidence, Australia’s most epidemiologically important mosquito-borne diseases is the Ross River virus (RRV, Togoviridae: Alphavirus). Ross River virus disease occurs widely across Australia with a significant burden to human health through epizootic spillover. Owing to its notifiable status, there is high-quality national health surveillance. In this thesis, my research focused on improving forecasting of RRV and understandings of the mechanisms driving patterns of RRV incidence. In Chapter 2, I describe research that I did on predictive modelling of RRV transmission in epidemic centres across Victoria determining environmental and meteorological factors associated with RRV disease incidence. I identified distinct similarities and differences in factors useful for predicting RRV incidence and outbreaks across Victoria, using readily available and inexpensive climate and meteorological data. The results highlighted new meteorological factors (i.e., rate of evapotranspiration) important in forecasting RRV incidence. The methodologies I used have formed the Ross River virus Outbreak Surveillance System for the Victorian Department of Health to monitor and provide intelligence for public health management action. The applied epidemiological implications from this study are processes and procedures to establish accessible predictive disease surveillance tools to aid public health management. This research documented how epidemiological studies can be integrated into public health systems. Having established an approach to construct a predictive surveillance system across multiple epidemic regions, in Chapter 3 I evaluated how the choice of the statistical model used in predictive modelling affects the accuracy of predictions. This was done to optimise methods for predicting RRV incidence and epidemics across Victoria and Western Australia. I determined that regional differences affect the choice of the statistical model used in establishing a predictive surveillance system. Current epidemiological studies forecasting RRV incidence often assume the best fit model for predicting RRV incidence is also the best for predicting RRV outbreaks, or vice versa. However, this assumption is rarely evaluated, and I found that the models used for predicting disease incidence may not be suitable for predicting disease outbreaks. Moreover, my research demonstrated that relative disease activity (i.e., greater number of notifications/incidence) does not lead to greater model predictive accuracy, and that, predictive accuracy is strongly influenced by model choice. Information gained from this research can be directly applied to improving and updating current RRV predictive surveillance systems. By using a common systematic approach to developing predictive disease surveillance systems, factors associated with disease can more reliably be assessed and compared across regions in relation to RRV epidemiology. In Chapter 4, I used distributed lag non-linear models to assess meteorological factors in non-linear exposure-response relationships for RRV incidence. My findings illustrated that the effect of meteorological factors on RRV incidence risk is non-linear with multiple delayed effects. I showed that there are optimal levels of exposure to meteorological factors that affect the risk of RRV incidence. My findings have the potential to allow public health personnel/departments to assess how future climate predictions may influence RRV disease activity across Australia with shifting climates. Currently predictive disease modelling of RRV often over or under predict disease activity, likely owing for complex dynamics not captured with linear predictor relationships. This research is foundational for exploring these methods as an approach to improve upon current predictive methods. In Chapter 5 I shifted my focus to improving upon mechanistic understanding of vector and host life-history traits and their importance in RRV pathogen transmission. Using empirically founded seasonally forced ordinary differential equation models, fitted to human RRV notification data, I investigated deterministic mechanisms of RRV transmission. I investigated mosquito vector, reservoir host, and transmission factors that shape RRV notifications among epidemic centres in Australia. I found that the combination of transmission mechanisms for RRV to be similar across sites supporting RRV as a multi-vector multi-host pathogen. My findings also support incidence of RRV as being largely underreported. By improving understanding of the fundamental transmission mechanisms of RRV my models have potential to aid current and future control strategies to limit the burden of human disease. Mosquito-borne diseases, such as Ross River virus disease, present an important issue for public health across Australia. The research presented in this thesis has enhanced forecasting and understanding of the mechanisms driving RRV disease incidence. The research contributions I have made help with applied epidemiological approaches in public health settings to improve population health management, such as through public health warnings associated with disease incidence, and with the control of mosquito vectors.

Introduction: This thesis presents research undertaken to assess the possibility of using data on climate, environment and the Ross River virus (RRV) vector and host species to develop early warning predictive models for RRV disease in New South Wales, Australia. Such models may then contribute to the anticipation, prevention and control of RRV disease. Background: Mosquito-borne diseases at the national and global levels are a substantial cause of morbidity and mortality. There are currently few vaccines available for mosquito-borne diseases (yellow fever and Japanese encephalitis); hence mosquito control, public health warnings and outbreak preparedness are the primary measures for preventing mosquito-borne diseases at the population level. On a longer time frame, the abatement of human-induce climate change and of aspects of land use and water management will also be important contributions to reducing the range and risks from these infectious diseases. For the more immediate measures to be successful, knowledge of when the next outbreak will occur within an appropriate response time is essential. As climate and environment have direct influences on components of the disease transmission cycle, they are suitable for inclusion in short-term and long-term predictive modelling of mosquito-borne diseases. RRV disease is the most widespread and most common mosquito-borne disease in Australia and causes considerable morbidity in the population. Over 86,500 notifications of RRV disease have been reported in Australia since 1991, an average of 4,300 notifications per year. RRV is maintained within a wide range of mosquitoes and vertebrate reservoir hosts. Climate and environment are known to influence the distribution and abundance of these RRV vectors and reservoir hosts, and thereby influence the probability of transmission to humans and the risk of clinical disease.

Pyrexia is an illness of unknown origin where people suffer unexplained fevers, headaches and fatigue. This study was undertaken to ascertain the prevalence of agents which may be associated with pyrexia in the Central Queensland region. Illnesses researched include Ross River Virus, Leptospirosis, Barmah Forest Virus, Brucellosis, Lyme Borrelosis and Streptococcal infections. A study was undertaken to ascertain the prevalence of agents which may be associated with pyrexia in the Central Queensland region. Five hundred and twenty four patient sera submitted to the laboratory from January 1991 to December 1992 for Ross River virus serology were also assayed by serological methods for evidence of exposure to Barmah Forest virus, flaviviruses, spotted fever group rickettsia, brucella, leptospirosis and lyme borrelosis. Titres of streptococcal antibodies and rheumatoid factor were also measured. Accompanying request forms were analysed for relevant clinical history and any additional pathology testing requests were noted. Ross River virus antibodies (IgG and/ or IgM) were found in 38.9% of patient sera. Males were significantly associated with the presence of antibodies (IgG and/or IgM) (P < 0.05) but females were 1.73 times as likely to have detectable IgM antibody at presentation. This suggests that females may be more symptomatic in the initial stages of infection and thus seek medical intervention. Flavivirus antibodies (IgG) were found in 19.79% of patient sera. One sample also contained IgM antibodies which reacted with both Kunjin and Alfuy viruses. Barmah Forest virus antibodies (IgG and/or IgM) were detected in 8% of patient sera. IgM antibody was detected in three patients, two of whom also had Ross River virus IgM antibody. Antibodies (IgG and/or IgM) to the spotted fever group of rickettsia were detected in 3.8% of patients. Lyme borrelosis antibodies (IgG) were detected in 1.7% of patients. A single case of brucellosis was diagnosed but no evidence of exposure to leptospiras was found. Raised streptococcal antibody titres were found in 27.1% of patient sera. Rheumatoid factor was detected in 4.696 of patient sera. A full blood count, erythrocyte sedimentation rate and rheumatoid factor determination were the most commonly requested additional pathology tests. Clinical history was provided on only 56.1% of request forms. These results have implications for future testing protocols. Diagnostic problems associated with serological testing on single serum samples were noted.

Background Climate variation has affected diverse physical and biological systems worldwide. Population health is one of the most important impacts of climate variation. Although the impact of climate variation on infectious diseases has been of significant concern recently, the relationship between climate variation and infectious diseases, including vector-borne diseases and enteric infections, needs greater clarification. Australia is grappling with developing politically acceptable responses to global warming. In China, few studies have been conducted to examine the effect of climate variation, including global warming, on population health. As residents of developing countries may suffer more from climate change compared with people living in more developed countries, this thesis has significance for both countries. Aims This study aims to contribute to a better understanding of the impact of climate variation on population health, and to provide scientific evidence for policy makers, researchers, public health practitioners and local communities in the development of public health strategies at an early stage, in order to prevent or reduce future risks associated with ongoing climate change. The objectives of this study include: (1) to quantify the association between climate variation and selected vectorborne diseases and enteric infections in different climatic regions in Australia and China; (2) to project the future burden of selected vector-borne diseases and enteric infections based on climate change scenarios in different climatic regions in Australia and China. Methods This ecological study has two components. The first uses time-series analyses to quantify the relationship between meteorological variables and infectious diseases, whereas the second projects the burden of selected infectious diseases using future climate and population scenarios. Temperate and subtropical climatic zones in both Australia and China were selected as the primary study areas, and a study of an Australian tropical region was also conducted. Study of Australia’s temperate zones was conducted in Adelaide, South Australia, as well as the Murray River region in that State. The study of China’s temperate zone was carried out in Jinan, Shandong Province. Subtropical studies were conducted in Baoan, Guangdong Province, China, and Brisbane in Queensland, whilst research for the tropics centred on Townsville, also in Queensland, Australia. The selected infectious diseases - one vector-borne disease and one enteric infection in each country - are Ross River Virus (RRV) infection and salmonellosis in Australia, and malaria and bacillary dysentery in China. Study periods vary from eight to sixteen years (depending upon the availability of data). Climate data, infectious disease surveillance data and demographic data were collected from local authorities. Data analyses conducted in the ecological studies include Spearman correlation analysis, time-series adjusted Poisson regression and the Seasonal Autoregressive Integrated Moving Average (SARIMA) model with consideration of lag effects, seasonality, long-term trends, and autocorrelation, on a weekly or monthly basis depending on data availability, and Hockey Sticky model to detect potential threshold temperatures. In the burden of disease component, analyses include the calculation of an indicator of the burden of disease - Years Lost due to Disabilities (YLDs) - and use scenario-based models to project YLDs for the selected diseases in 2030 and 2050 in Australia and 2020 and 2050 in China respectively. The projections consider both different scenarios of projected temperature and future population change. Results Relationship between climate variation and selected infectious diseases In all the study regions in Australia, maximum temperature, minimum temperature, rainfall and humidity are all significantly related to the number of RRV infections, with lag effects varying from 0 to 3 months. Additionally, high tides in the two seaside regions with tropical (Townsville) or subtropical (Brisbane) climates, and river flow in the temperate region (Murray River region), are related to the number of cases without any lag effects. A potential 1°C increase in maximum or minimum temperature may cause 4%~23% extra cases of RRV infection in the temperate region, 5~8% in the subtropical region, and 6%~15% in the tropical region. Maximum temperature, minimum temperature, humidity and air pressure are significantly related to malaria cases in the temperate city Jinan and subtropical city Baoan in China, with a lag effect range of 0 to 1 month. An association between rainfall and malaria cases was not detected in either region. A potential 1°C increase in maximum or minimum temperature may lead to 4%~15% extra malaria cases in the temperate region, and 12%-18% in the tropical region in China. Maximum temperature, minimum temperature, rainfall and humidity are all significantly related to the number of salmonellosis cases in the three study cities in Australia, with lag effects varying from 0 to 1 month. A potential 1°C increase in maximum or minimum temperature may cause 6%~19% extra salmonellosis cases in the temperate region (Adelaide), 5%~10% in the subtropical region (Brisbane), and 4%~15% in the tropical region (Townsville). The thresholds for the effects of maximum and minimum temperatures are 20ºC and 12ºC respectively in Adelaide. No threshold temperatures are detected in Townsville and Brisbane. Maximum temperature, minimum temperature, humidity, air pressure and rainfall are significantly related to bacillary dysentery cases in the temperate city Jinan and subtropical city Baoan in China, with the lag effect range of 0 to 2 months. A potential 1°C increase in maximum or minimum temperature may cause 7%~15% extra bacillary dysentery cases in the temperate region and 10% ~ 19% in the subtropical region in China. The thresholds for the effects of maximum and minimum temperatures on bacillary dysentery are 17ºC and 8ºC respectively in Jinan. No threshold temperatures are detected in Baoan. Projection of YLDs from target diseases In Australia, considering both climatic and population scenarios, if other factors remain constant, compared with the YLDs observed in 2000, the YLDs for salmonellosis might increase by up to 48% by 2030, and nearly double by 2050 in South Australia, while the YLDs might double by 2030 and increase by up to 143% by 2050 in Brisbane, Queensland. The YLDs for RRV infection might increase by up to 66% by 2030, and nearly double by 2050 in South Australia. They might increase by up to 61% by 2030 and double by 2050 in Brisbane, Queensland. In China, considering both climatic and population scenarios, if other factors remain constant, compared with the YLDs observed in 2000, the YLDs for bacillary dysentery might double by 2020 and triple by 2050 in both Jinan and Baoan. The YLDs for malaria might increase by up to 108% by 2020 and nearly triple by 2050 in Jinan, the temperate city, and increase by up to 144% by 2020 and nearly triple by 2050 in Baoan, the subtropical city. Conclusions 1. Both maximum and minimum temperatures are important in the transmission of vector-borne diseases in various climatic regions in both Australia and China. River flow or high tides may also play an important role in the transmission of such diseases. 2. Both maximum and minimum temperatures play an important role in the transmission of enteric infections in various climatic regions in both Australia and China, with a threshold temperature detected in the temperate regions but not in subtropical and tropical regions. 3. The effects of rainfall and relative humidity on selected infectious diseases vary in different study areas in Australia and China. 4. The burden of temperature-related infectious diseases may greatly increase in the future if there is no effective preventive intervention. Public health implications 1. Implication for health practice • Public health practitioners, together with relevant government organisations, should monitor trends in infectious diseases, as well as other relevant indexes, such as vectors, pathogens, and water and food safety. They should advise policy makers of the potential risks associated with climate change and develop public health strategies to prevent and reduce the impact of infectious disease associated with such change. • Doctors and other clinical practitioners should be prepared and supported in the provision of health care for any expected extra cases associated with climate variation and should play an important role in relevant health education on climate change. • Community participation is of significance to adapt to and mitigate the risk of climate change on population health. Community involvement helps to deliver programmes which more accurately target local needs. Therefore, community should be involved in the partnerships of climate change as early as possible. • Relevant education programs on the potential health impact of climate change should be conducted by government at all levels for different stakeholders, including industries, governments, communities, clinicians and researchers. • Advocacy for adapting to and mitigating climate change should be a longstanding public health activity. 2. Implication for researchers • The main task for researchers is to identify the independent contribution made by key climatic variables and whether there are exposure thresholds for infectious disease transmission. Further studies should include various infectious diseases in different climatic regions. • Developing countries and rural regions are more vulnerable to the impact of climate change so more research should be conducted for people living in those regions. • Studies using summary measures that combine prevalence of disease, quality of life and life expectancy, such as Disability Adjusted Life Years (DALYs), to assess the burden of disease due to climate change is necessary to assist in decision making. • More research should be conducted on the assessment of adaptive strategies and mitigation to future climate change. 3. Implication for policies • Public and preventive health strategies that consider local climatic conditions and their impact on vector and food borne diseases are important in reducing such impact due to climate change in the future. • The extra health burden that may be caused by future climate change may have a great impact on the currently overloaded public health system in both developed and developing countries. Long-term planning about health resource allocation, infrastructure establishment, and relevant response mechanisms should be developed at relevant government levels. • Effective prevention and intervention strategies will be possible only if the efforts of relevant sectors, including governments, communities, industries, research institutions, clinical professionals and individuals, have coordinated responses. • International and regional collaborations are necessary to address this global issue. In addition, strategies of an international dimension should be translated into regional and local actions. This is extremely important to developing countries such as China and India. • Sustainable development policies with consideration given to reducing green house gases and environmental degradation need immediate action which will benefit future generations. Health priorities should include the prevention of climate change.<br>http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1290777<br>Thesis(Ph.D.)-- School of Population Health and Clinical Practice, 2007