Thesis (PhD) - University of Canberra, 2007. / Includes bibliography (p.129-153).

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.

Thesis (Ph.D.)--University of Western Australia, 2006.

[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.

[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.

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. / 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.

An ecological study was undertaken on four populations of Krefft�s River Turtle Emydura krefftii inhabiting the Townsville Area of Tropical North Queensland. Two sites were located in the Ross River, which runs through the urban areas of Townsville, and two sites were in rural areas at Alligator Creek and Stuart Creek (known as the Townsville Creeks). Earlier studies of the populations in Ross River had determined that the turtles existed at an exceptionally high density, that is, they were superabundant, and so the Townsville Creek sites were chosen as low abundance sites for comparison. The first aim of this study was to determine if there had been any demographic consequences caused by the abundance of turtle populations of the Ross River. Secondly, the project aimed to determine if the impoundments in the Ross River had affected the freshwater turtle fauna. Specifically this study aimed to determine if there were any difference between the growth, size at maturity, sexual dimorphism, size distribution, and diet of Emydura krefftii inhabiting two very different populations. A mark-recapture program estimated the turtle population sizes at between 490 and 5350 turtles per hectare. Most populations exhibited a predominant female sex-bias over the sampling period. Growth rates were rapid in juveniles but slowed once sexual maturity was attained; in males, growth basically stopped at maturity, but in females, growth continued post-maturity, although at a slower rate. Sexual maturity was at 6-7 years of age for males, which corresponded to a carapace length of 150-160 mm, and 8-10 years of age for females, which corresponded to a carapace length of 185-240 mm. The turtles were omnivorous, although in the Ross River they ate more submerged vegetation (by percent amount and occurrence) than those of the Townsville Creeks. Turtles in Townsville Creeks ingested more windfall fruit and terrestrial insects.

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. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1290777 / Thesis(Ph.D.)-- School of Population Health and Clinical Practice, 2007

[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.

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.