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More than 75 arboviruses (arthropod-borne viruses) have been identified in Australia. While Alfuy virus (ALFV), Barmah Forest virus (BFV), Edge Hill virus (EHV), Kokobera virus (KOKV), Murray Valley encephalitis virus (MVEV), Sindbis virus (SINV), Ross River virus (RRV), Stratford virus (STRV), and West Nile virus strain Kunjin (KUNV) have been associated with human infection, there remains a paucity of data regarding their respective transmission cycles and any potential nonhuman vertebrate hosts. It is likely that these viruses are maintained in zoonotic cycles involving native animals rather than solely by human-to-human transmission. A serosurvey (n = 100) was undertaken to determine the prevalence of neutralizing antibodies against a panel of Australian arboviruses in western gray kangaroos (Macropus fuliginosus) obtained from 11 locations in the midwest to southwest of Western Australia. Neutralizing antibodies against RRV were detected in 25%, against BFV in 14%, and antibodies to both viruses in 34% of serum samples. The prevalence of antibodies against these two viruses was the same in males and females, but higher in adult than in subadult kangaroos (p

We report a case of Barmah Forest virus infection in a child from Central Province, Papua New Guinea, who had no previous travel history. Genomic characterization of the virus showed divergent origin compared with viruses previously detected, supporting the hypothesis that the range of Barmah Forest virus extends beyond Australia.

An arbovirus surveillance military exercise was conducted to assess the risk of Ross River virus (RRV) and Barmah Forest virus (BFV) in the Australian Defence Force (ADF) Wide Bay training area (WBTA), northeastern Australia, in April 2018. Of the 5,540 female mosquitoes collected, 3,702 were screened for RRV and BFV by quantitative reverse transcription–polymerase chain reaction in a field laboratory. One pool of Verrallina funerea was positive for RRV and 8 pools (7 pools of Aedes vigilax and 1 pool of Culex annulirostris) were positive for BFV. Phylogenetic analysis of the complete nucleotide sequence of the E2 protein subgrouped both RRV and BFV with viruses previously isolated from human infections, indicating the potential risk of RRV and BFV infection to ADF personnel while training in WBTA. This is the 1st time that both RRV and BFV have been detected in a military training area.

Barmah Forest virus (BFV) is a medically important mosquito‑borne alphavirus endemic to Australia. Symptomatic disease can be a major cause of morbidity, associated with fever, rash, and debilitating arthralgia. BFV disease is similar to that caused by Ross River virus (RRV), the other major Australian alphavirus. Currently, just four BFV whole‑genome sequences are available with no genome‑scale phylogeny in existence to robustly characterise genetic diversity. Thirty novel genome sequences were derived for this study, for a final 34-taxon dataset sampled over a 44 year period. Three distinct BFV genotypes were characterised (G1&ndash, 3) that have circulated in Australia and Papua New Guinea (PNG). Evidence of spatio‑temporal co‑circulation of G2 and G3 within regions of Australia was noted, including in the South West region of Western Australia (WA) during the first reported disease outbreaks in the state&rsquo, s history. Compared with RRV, the BFV population appeared more stable with less frequent emergence of novel lineages. Preliminary in vitro assessment of RRV and BFV replication kinetics found that RRV replicates at a significantly faster rate and to a higher, more persistent titre compared with BFV, perhaps indicating mosquitoes may be infectious with RRV for longer than with BFV. This investigation resolved a greater diversity of BFV, and a greater understanding of the evolutionary dynamics and history was attained.

Arboviruses (ARthropods BOrne VIRUSES) are disease-causing viruses transmitted through the bite of hematophagous arthropods, such as mosquitoes and ticks. Among these, the alphavirus, genus of the Togaviridae family, is considered the most arthritogenic species, responsible for diseases such as chikungunya fever (CHIK), O'nyong-nyong virus fever, Ross River virus disease, Barmah Forest virus disease, Sindbis virus disease, and Mayaro fever. These arboviral diseases, especially CHIK, have impacted public health in recent decades, leading to devastating epidemics, particularly in developing countries, due to their high potential for chronicity, functional impairment, and great impact on the quality of life. In a similar way, chronic musculoskeletal symptoms have been described in all alphavirus infections. However, CHIK is the best studied. The purpose of this article is to review physiopathology, clinical manifestations, diagnosis, and treatment of alphaviruses, focusing on CHIK and chronic evolution of musculoskeletal symptoms.

In the state of Western Australia, Australia, active surveillance is conducted for mosquitoborne viruses of major human health significance: alphaviruses Ross River virus (RRV) and Barmah Forest virus (BFV) and flaviviruses Murray Valley encephalitis virus (MVEV) and West Nile virus (subtype Kunjin virus; KUNV). These flaviviruses are endemic and epidemic to the northern and central areas of Australia, where surveillance involves year-round testing for seroconversions in sentinel chickens and virus isolation from mosquito pools collected annually. More frequent mosquito collection is prevented by the logistical difficulties of accessing remote areas. Commonly isolated arboviruses include the flaviviruses MVEV (and subtype Alfuy virus), KUNV, Kokobera virus (KOKV), and Edge Hill virus (EHV) and the alphaviruses RRV, BFV, and Sindbis virus. This system occasionally detects viruses that cannot be identified as known viruses, such as Stretch Lagoon virus, an orbivirus isolated in 2002. We describe the detection and characterization of a novel flavivirus named Fitzroy River virus (FRV), isolated from mosquitoes collected in northern Western Australia, and seroepidemiologic evidence of human or animal infection.

Infections with commonly occurring Australian arthropod-borne arboviruses such as Ross River virus (RRV) and Barmah Forest virus (BFV) are diagnosed routinely by pathology laboratories in Australia. Others, such as Murray Valley encephalitis (MVEV) and Kunjin (KUNV) virus infections may be diagnosed by specialist reference laboratories. Although Alfuy (ALFV), Edge Hill (EHV), Kokobera (KOKV), Sindbis (SINV), and Stratford (STRV) viruses are known to infect humans in Australia, all are considered ‘neglected.’ The aetiologies of approximately half of all cases of undifferentiated febrile illnesses (UFI) in Australia are unknown and it is possible that some of these are caused by the neglected arboviruses. The aims of this study were to determine the seroprevalence of antibodies against several neglected Australian arboviruses among residents of Queensland, north-east Australia, and to ascertain whether any are associated with UFI. One hundred age- and sex-stratified human plasma samples from blood donors in Queensland were tested to determine the prevalence of neutralising antibodies against ALFV, BFV, EHV, KOKV, KUNV, MVEV, RRV, SINV, and STRV. The seroconversion rates for RRV and BFV infections were 1.3 and 0.3% per annum, respectively. The prevalence of antibodies against ALFV was too low to enable estimates of annual infection rates to be determined, but the values obtained for other neglected viruses, EHV (0.1%), KOKV (0.05%), and STRV (0.05%), indicated that the numbers of clinical infections occurring with these agents are likely to be extremely small. This was borne out by the observation that only 5.7% of a panel of 492 acute phase sera from UFI patients contained IgM against any of these arboviruses, as detected by an indirect immunofluorescence assay. While none of these neglected arboviruses appear to be a cause of a significant number of UFIs in Australia at this time, each has the potential to emerge as a significant human pathogen if there are changes to their ecological niches.

This report describes the epidemiology of mosquito-borne diseases of public health importance in Australia during the 2014–15 season (1 July 2014 to 30 June 2015) and includes data from human notifications, sentinel chicken, vector and virus surveillance programs. The National Notifiable Diseases Surveillance System received notifications for 12,849 cases of disease transmitted by mosquitoes during the 2014–15 season. The Australasian alphaviruses Barmah Forest virus and Ross River virus accounted for 83% (n=10,723) of notifications. However, over-diagnosis and possible false positive diagnostic test results for these two infections mean that the true burden of infection is likely overestimated, and as a consequence, revised case definitions were implemented from 1 January 2016. There were 151 notifications of imported chikungunya virus infection. There were 74 notifications of dengue virus infection acquired in Australia and 1,592 cases acquired overseas, with an additional 34 cases for which the place of acquisition was unknown. Imported cases of dengue were most frequently acquired in Indonesia (66%). There were 7 notifications of Zika virus infection. No cases of locally-acquired malaria were notified during the 2014–15 season, though there were 259 notifications of overseas-acquired malaria and one notification for which no information on the place of acquisition was supplied. Imported cases of malaria were most frequently acquired in southern and eastern Africa (23%) and Pacific Island countries (20%). In 2014–15, arbovirus and mosquito surveillance programs were conducted in most of the states and territories. Surveillance for exotic mosquitoes at international ports of entry continues to be a vital part of preventing the establishment of vectors of mosquito-borne diseases such as dengue to new areas of Australia. In 2014-15, there was a sharp increase in the number of exotic mosquitoes detected at the Australian border, with 36 separate exotic mosquito detections made, representing a 280% increase from the 2013-14 period where there were 13 exotic mosquito detections. Commun Dis Intell 2016;40(3):e401–436.

Effective arbovirus surveillance is essential to ensure the implementation of control strategies, such as mosquito suppression, vaccination, or dissemination of public warnings. Traditional strategies employed for arbovirus surveillance, such as detection of virus or virus-specific antibodies in sentinel animals, or detection of virus in hematophagous arthropods, have limitations as an early-warning system. A system was recently developed that involves collecting mosquitoes in CO2-baited traps, where the insects expectorate virus on sugar-baited nucleic acid preservation cards. The cards are then submitted for virus detection using molecular assays. We report the application of this system for detecting flaviviruses and alphaviruses in wild mosquito populations in northern Australia. This study was the first to employ nonpowered passive box traps (PBTs) that were designed to house cards baited with honey as the sugar source. Overall, 20/144 (13.9%) of PBTs from different weeks contained at least one virus-positive card. West Nile virus Kunjin subtype (WNVKUN), Ross River virus (RRV), and Barmah Forest virus (BFV) were detected, being identified in 13/20, 5/20, and 2/20 of positive PBTs, respectively. Importantly, sentinel chickens deployed to detect flavivirus activity did not seroconvert at two Northern Territory sites where four PBTs yielded WNVKUN. Sufficient WNVKUN and RRV RNA was expectorated onto some of the honey-soaked cards to provide a template for gene sequencing, enhancing the utility of the sugar-bait surveillance system for investigating the ecology, emergence, and movement of arboviruses.

Review question/objective The objective of the proposed systematic review is to critically analyze currently available research studies and present the best available evidence related to the effectiveness of disease management interventions on health-related quality of life (HRQOL) of patients with established arthritogenic alphavirus infections. The primary review question is: What is the effectiveness of clinical manifestations management interventions on HRQOL outcomes among patients with established arthritogenic alphavirus infections? The secondary review questions are: What is the effectiveness of early diagnosis of disease on HRQOL outcomes among patients with established arthritogenic alphavirus infections? What is the effectiveness of arthritogenic alphavirus infections education on HRQOL outcomes among patients with established arthritogenic alphavirus infections? Inclusion criteria Types of participants This systematic review will include patients with established arthritogenic alphavirus infections of all ages and stages of infection. Arthritogenic alphavirus infections will include infection from the following alphaviruses: 1) Chikungunya virus 2) Ross River virus 3) Barmah Forest virus 4) O'Nyong-nyong virus 5) Mayaro virus 6) Sindbis virus 7) Semliki Forest virus. Types of intervention(s)/phenomena of interest The interventions of interest are those utilized in the management of arthritogenic Alphavirus infections, confined to the scope of the primary and secondary questions: 1) Management interventions for clinical manifestations 2) Early diagnosis of disease 3) Disease education (including psycho-education). Types of outcomes To be included, studies will measure self-reported objective and subjective measures of HRQOL as the primary or secondary end points. Specific outcomes of interest in this review include: 1) Overall HRQOL measured using HRQOL assessment tools 2) HRQOL domains measured using HRQOL assessment tools, not limited to: a) Physical functioning b) Cognitive functioning c) Social functioning d) Role functioning e) Emotional functioning f) Pain g) Anxiety h) Depression i) Fatigue j) Sleep k) General health perspective. 3) Other domains not covered by HRQOL assessment tools, usually relating to disease-specific and treatment-specific symptoms. Adverse outcomes of interest will include: a) Any harm associated with the management of arthritogenic alphavirus infections b) Decrease in overall HRQOL or HRQOL domain c) Recurrence of arthritogenic alphavirus infections or the onset of secondary disease.

The most common causes of human infection from the arboviruses that are endemic in Australia are the arthritogenic alphaviruses: Ross River virus (RRV) and Barmah Forest virus (BFV). The most serious infections are caused by the neurotropic flaviviruses, Murray Valley encephalitis virus (MVEV) and the Kunjin subtype of West Nile virus. The greatest individual risk of arbovirus infection occurs in tropical/subtropical northern Australia because of the warm, wet summer conditions from December to June, where conventional arbovirus surveillance is difficult due to a combination of low population density, large distances between population centers, poor roads, and seasonal flooding. Furthermore, virus detection requires samples to be sent to Perth up to 2,000 km away for definitive analysis, causing delays of days to weeks before test results are available and public health interventions can be started. We deployed a portable molecular biology laboratory for remote field detection of endemic arboviruses in northern Queensland, then in tropical Western Australia and detected BFV, MVEV, and RRV RNA by polymerase chain reaction (PCR) assays of extracts from mosquitoes trapped in Queensland. We then used a field-portable compact real-time thermocycler for the samples collected in the Kimberley region of Western Australia. Real-time field PCR assays enabled concurrent endemic arbovirus distribution mapping in outback Queensland and Western Australia. Our deployable laboratory method provides a concept of operations for future remote area arbovirus surveillance.

Ross River virus (RRV), Barmah Forest virus (BFV), and dengue are three common mosquito-borne diseases in Australia that display notable seasonal patterns. Although all three diseases have been modeled on localized scales, no previous study has used harmonic models to compare seasonality of mosquito-borne diseases on a continent-wide scale. We fit Poisson harmonic regression models to surveillance data on RRV, BFV, and dengue (from 1993, 1995 and 1991, respectively, through 2015) incorporating seasonal, trend, and climate (temperature and rainfall) parameters. The models captured an average of 50–65% variability of the data. Disease incidence for all three diseases generally peaked in January or February, but peak timing was most variable for dengue. The most significant predictor parameters were trend and inter-annual periodicity for BFV, intra-annual periodicity for RRV, and trend for dengue. We found that a Temperature Suitability Index (TSI), designed to reclassify climate data relative to optimal conditions for vector establishment, could be applied to this context. Finally, we extrapolated our models to estimate the impact of a false-positive BFV epidemic in 2013. Creating these models and comparing variations in periodicities may provide insight into historical outbreaks as well as future patterns of mosquito-borne diseases.

Australia is a climatically diverse country varying from a tropical climate in the north to arid central desert and grassland regions, and to temperate climates in the south. There are many viral infections found in Australia that are common to developed countries worldwide, but this article will focus on those that pose a special risk for travellers to Australia, especially the mosquito-borne viruses. The commonest are the members of the alphavirus genus, particularly Ross River virus and Barmah Forest virus, which cause predominantly arthralgia with or without fever or rash. Less frequent but more serious illness is seen with the encephalitic flaviviruses, Murray Valley encephalitis virus, and the Kunjin strain of West Nile virus. In addition dengue occurs intermittently on the northern part of Queensland, and in recent years Japanese encephalitis virus has been found in the Torres Strait Islands and the tip of far north Queensland. Also of interest, but with a much lower risk, are the bat-borne viruses, Hendra virus and Australian bat lyssavirus, that have caused a small number of human infections. However, it is important to remember that most tourists pass through other countries in the Asia/Pacific region on their way to and from Australia and may therefore have acquired infections prior to or after leaving Australia.

The two most reported mosquito-borne diseases in Queensland, a northern state of Australia, are Ross River virus (RRV) disease and Barmah Forest virus (BFV) disease. Both diseases are endemic in Queensland and have similar clinical symptoms and comparable transmission cycles involving a complex inter-relationship between human hosts, various mosquito vectors, and a range of nonhuman vertebrate hosts, including marsupial mammals that are unique to the Australasian region. Although these viruses are thought to share similar vectors and vertebrate hosts, RRV is four times more prevalent than BFV in Queensland.We performed a retrospective analysis of BFV and RRV human disease notification data collected from 1995 to 2007 in Queensland to ascertain whether there were differences in the incidence patterns of RRV and BFV disease. In particular, we compared the temporal incidence and spatial distribution of both diseases and considered the relationship between their disease dynamics. We also investigated whether a peak in BFV incidence during spring was indicative of the following RRV and BFV transmission season incidence levels.Although there were large differences in the notification rates of the two diseases, they had similar annual temporal patterns, but there were regional variations between the length and magnitude of the transmission seasons. During periods of increased disease activity, however, there was no association between the dynamics of the two diseases.The results from this study suggest that while RRV and BFV share similar mosquito vectors, there are significant differences in the ecology of these viruses that result in different epidemic patterns of disease incidence. Further investigation is required into the ecology of each virus to determine which factors are important in promoting RRV and BFV disease outbreaks.

Associations between mosquito abundance and Ross River virus (RRV) and Barmah Forest virus (BFV) disease are demonstrated for the Gippsland Lakes region of eastern Victoria, Australia. Significant ...

Between October 2012 and October 2013, unprecedented high numbers of Barmah Forest virus (BFV) disease cases were reported in the Northern Territory (NT). An investigation was launched by the NT Department of Health in cooperation with the Department of Primary Industry and Fisheries and the Department of Land Resource Management to investigate possible causes for this phenomenon. The investigation included virus isolations from mosquitoes collected in Darwin urban areas, BFV antibody testing in peri-urban small mammals and a human BFV disease case series investigation of recent cases. No BFV was isolated from the 4641 mosquitoes tested, none of the mammals tested positive for BFV antibodies, and the high BFV disease case numbers did not correlate with the relatively low mosquito vector numbers trapped in 2012-2013. It was estimated that up to 89% of the 79 human cases investigated did not have an acute arboviral illness and therefore had tested falsely positive. An Alere PanBio BFV immunoglobulin M enzyme-linked immunosorbent assay test kit is generally used to test for BFV, with the BFV disease case definition based on immunoglobulin M positives only. Other jurisdictions in Australia also reported high numbers of BFV disease cases, with the majority of the cases suspected to be false positives. Therefore, current testing methods need to be revised to reflect the true numbers of BFV disease cases occurring in Australia and to provide correct diagnoses for patients.

Host feeding patterns of mosquitoes were assessed through the identification of 865 blood meals collected from Brisbane during 2000-2001. Under natural conditions, mosquito feeding (including that of Culex annulirostris, Aedes vigilax, and Aedes notoscriptus) was primarily on dogs (37.4%), but also on birds (18.4%), horses (16.8%), brushtail possums (13.3%), humans (11.6%), and cats, flying foxes, and macropods, depending on site. From 1997 to 1999, sera (N=1706) were collected from dogs, cats, horses, flying foxes, and brushtail possums in the Brisbane area and were analyzed by microneutralization assay for antibodies to Ross River virus (RRV) and Barmah Forest virus (BFV). For RRV, all vertebrate species tested had been naturally infected, and seroprevalence varied from 10.5% to 25.5%, whereas for BFV, rates varied between 0% and 11.3%. Brushtail possums were often infected in the field, with 17.6% and 10.7% of wild individuals having antibodies to RRV and BFV, respectively. Horses and flying foxes also had a relatively high prevalence of antibodies to RRV. This study, therefore, provides data to indicate that brushtail possums play a role in the urban transmission of RRV in Brisbane and that horses, when they occur, also fill the same role.

Spatially and temporally accurate information about infectious mosquito distribution allows for pre-emptivepublic health interventions that can reduce the burden of mosquito-borne infections on human populations.However, the labile nature of arboviruses, the low prevalence of infection in mosquitoes, the expensive laborcosts for mosquito identification and sorting, and the specialized equipment required for arbovirus testing canobstruct arbovirus surveillance efforts. The recently developed techniques of testing mosquito expectorate usinghoney-baited nucleic acid preservation cards or sugar bait stations allows a sensitive method of testing forinfectious, rather than infected, mosquito vectors. Here we report the results from the first large-scale incorporationof honey-baited cards into an existing mosquito surveillance program. During 4 months of the peakvirus season ( January–April, 2014) for a total of 577 trap nights, we set CO2-baited encephalitis vector survey(EVS) light traps at 88 locations in South Australia. The collection container for the EVS trap was modified toallow for the placement of a honey-baited nucleic acid preservation card (FTATM card) inside. After collection,mosquitoes were maintained in a humid environment and allowed access to the cards for 1 week. Cards werethen analyzed for common endemic Australian arboviruses using a nested RT-PCR. Eighteen virus detections,including 11 Ross River virus, four Barmah Forest virus, and three Stratford virus (not previously reported fromSouth Australia) were obtained. Our findings suggest that adding FTA cards to an existing mosquito surveillanceprogram is a rapid and efficient way of detecting infectious mosquitoes with high spatial resolution. Refereed/Peer-reviewed

Verrallina funerea (Theobald) (Diptera: Culicidae) is a brackish water mosquito species found most commonly in Indonesia, Papua New Guinea, and the northeastern coastal regions of Australia. Aspects of the vector competence of this species for Barmah Forest virus (family Togaviridae, genus Alphavirus, BFV) and Ross River virus (family Togaviridae, genus Alphavirus, RRV), two medically important arboviruses in Australia, were investigated. Laboratory-reared Ve. funerea were moderately susceptible to experimental infection with BFV (median cell culture infectious dose of 10(3.6) per mosquito) and were capable of transmission to suckling mice (52% after a 9-12-d extrinsic incubation period). Maximum salivary gland infections for BFV (65%) and RRV (50%) were observed 8 and 10 d postinfection, respectively. To examine any regional differences in vector competence, field populations (separated by up to 200 km) of Ve. funerea and Aedes vigilax (Skuse) from southeastern Queensland and northern New South Wales were fed BFV and RRV, and subsequent infection rates were compared. For both viruses, no statistically significant variations in body, disseminated, or salivary gland infection rates were found in either Ve. funerea or Ae. vigilax. The results from this study indicate that Ve. funerea may have an important role as an amplification vector during outbreaks of both viruses and that local government authorities should rapidly treat brackish water habitats to control this species during periods of increased disease activity.

Barmah Forest virus (BFV) disease is the second most common mosquito-borne disease in Australia. Although the majority of notifications are received from Queensland, little is known about the distribution of the disease within the state, or the important mosquito vectors and nonhuman vertebrate hosts. We conducted a retrospective statistical analysis of the notifications received from Queensland residents from 1993 to 2003 to establish long-term local incidence rates and to identify disease outbreaks. In total, 4,544 notifications were received over the 10-yr period. Disease reporting peaked in autumn, although the peak transmission season encompassed both summer and autumn. Long-term standardized incidence rates for summer/autumn and winter/spring varied across the state, showing positive spatial autocorrelation in both 6-mo periods. Although 15 instances of increased disease activity were identified, only one major disease outbreak affecting eight contiguous local government areas was detected in summer/autumn 2002/2003. This outbreak contained 297 cases, 115 more than would be expected over this period. The factors important to this outbreak are unknown and require further investigation. Although the incidence rates for BFV disease are lower than Ross River virus disease, the most reported mosquito-borne disease in Australia, several factors indicate that this virus should be considered an important public health risk in Queensland. These include consistent endemic transmission, apparent underreporting of the disease, and the potential for outbreaks in major population centers.

Our paper presents an assessment of research and operational development in relation to medically important mosquito-borne disease, mainly the arboviruses Ross River, Barmah Forest, Murray Valley encephalitis, Japanese encephalitis, Kunjin and dengue, but also with respect to malaria. Since 1972, there have been considerable gains in research output, organisational structure, communication, surveillance including quarantine inspection and operational control. This has been due to the 1974 epidemic of Murray Valley encephalitis virus extending into temperate Australia, increasing occurrence of Ross River and the dengue viruses, the discovery of Barmah Forest virus as a disease entity in 1988, and the introduction of Japanese encephalitis in 1995. Because many of the outputs involve methodologies of global import, this has resulted in an unprecedented upsurge in publications of international standard.

SUMMARY A series of five patients presented with eruptions beginning on the face. In each case the facial changes were characterized by urticated erythema with minimal epidermal change. The patients also demonstrated more widespread vesiculopapular, macular or purpuric eruptions. At presentation most patients were asymptomatic; however, several subsequently developed constitutional symptoms. Each of these patients was reactive for Barmah Forest virus immunoglobulin (Ig)M, and on repeat testing four were reactive for Barmah Forest virus IgG.

Ross River virus and Barmah Forest virus are Australian arboviruses of the Alphavirus genus. Features of alphavirus infection include an increased permeability of cells to monovalent cations followed by virion budding. Virally encoded ion channels are thought to have a role in these processes. In this paper, the 6K proteins of Ross River virus and Barmah Forest virus are shown to form cation-selective ion channels in planar lipid bilayers. Using a novel purification method, bacterially expressed 6K proteins were inserted into bilayers with a defined orientation (i.e. N-terminal cis, C-terminal trans). Channel activity was reversibly inhibited by antibodies to the N and C termini of 6K protein added to the cis and trans baths, respectively. Channel conductances varied from 40-800 picosiemens, suggesting that the protein is able to form channels with a range of possible oligomerization states.

In 2005, established populations of Aedes albopictus (Skuse) were discovered in the Torres Strait, the region that separates Papua New Guinea from northern Australia. This increased the potential for this species to be introduced to mainland Australia. Because it is an arbovirus vector elsewhere, we undertook laboratory-based infection and transmission experiments to determine the potential for Ae. albopictus from the Torres Strait to become infected with and transmit the four major Australian endemic arboviruses--Murray Valley encephalitis virus, West Nile virus Kunjin strain (WNV(KUN)), Ross River virus (RRV), and Barmah Forest virus--as well as the exotic Japanese encephalitis virus. Ae. albopictus is susceptible to infection with all viruses, with infection rates ranging between 8% for WNV(KUN) and 71% for RRV. Transmission rates of approximately 25% were observed for RRV and Barmah Forest virus, but these were17% for Murray Valley encephalitis virus, WNV(KUN), and Japanese encephalitis virus. Given its relative vector competence for alphaviruses, we also examined the replication kinetics and extrinsic incubation periods required for transmission of RRV and chikungunya virus. Despite lower body titers, more mosquitoes reared and maintained at 28 degrees C became infected with and transmitted the virus than those reared and maintained at 22 degrees C. The minimum time between Ae. albopictus consuming an infected bloodmeal and transmitting chikungunya virus was 2 d at 28 degrees C and 4 d at 22 degrees C, and for RRV, it was 4 d, irrespective of the temperature. Given its opportunistic feeding habits and aggressive biting behavior, the establishment of Ae. albopictus on the Australian mainland could have a considerable impact on alphavirus transmission.

SUMMARYRoss River virus (RRV) is a fascinating, important arbovirus that is endemic and enzootic in Australia and Papua New Guinea and was epidemic in the South Pacific in 1979 and 1980. Infection with RRV may cause disease in humans, typically presenting as peripheral polyarthralgia or arthritis, sometimes with fever and rash. RRV disease notifications in Australia average 5,000 per year. The first well-described outbreak occurred in 1928. During World War II there were more outbreaks, and the name epidemic polyarthritis was applied. During a 1956 outbreak, epidemic polyarthritis was linked serologically to a group A arbovirus (Alphavirus). The virus was subsequently isolated from Aedes vigilax mosquitoes in 1963 and then from epidemic polyarthritis patients. We review the literature on the evolutionary biology of RRV, immune response to infection, pathogenesis, serologic diagnosis, disease manifestations, the extraordinary variety of vertebrate hosts, mosquito vectors, and transmission cycles, antibody prevalence, epidemiology of asymptomatic and symptomatic human infection, infection risks, and public health impact. RRV arthritis is due to joint infection, and treatment is currently based on empirical anti-inflammatory regimens. Further research on pathogenesis may improve understanding of the natural history of this disease and lead to new treatment strategies. The burden of morbidity is considerable, and the virus could spread to other countries. To justify and design preventive programs, we need accurate data on economic costs and better understanding of transmission and behavioral and environmental risks.

Aedes aegypti (L.), Culex sitiens Weidemann, Culex annulirostris Skuse, and Culex quinquefasciatus Say mosquitoes colonized at the Queensland Institute of Medical Research, Brisbane Australia, were fed on blood containing Barmah Forest virus (BF). Only Cx. annulirostris was susceptible to infection, with a median cell culture infectious dose (CCID50) of 103.36 per mosquito. Ae. aegypti and Cx. quinquefasciatus were infected experimentally, but at rates of

In 1995, the largest recorded outbreak of human disease resulting from infection with the mosquito transmitted alphavirus Barmah Forest (BF) virus occurred along the south coast of New South Wales, Australia. The virus was first isolated in early January from mosquitoes collected at Batemans Bay and predisposed the recognition of 135 human clinical cases. The cases of BF virus were identified initially from Batemans Bay during late January, and the majority (30%) of all cases came from this town. After 5 wk, all major centers on the south coast had clinical patients. Aedes vigilax (Skuse) were especially abundant at Batemans Bay, with levels up to 8 times greater than normal. This species yielded 111 isolates and appeared to be the major vector of BF virus. Attempts to examine if BF virus was maintained in the field by vertical transmission within Ae. vigilax populations were unsuccessful; no evidence of vertical transmission with BF virus, nor any other arbovirus, was found in >17,000 adults emerging from field-collected larvae from the region following peak virus activity. In addition to BF virus, other viruses were recovered from field-collected adult mosquitoes, including Ross River (10 isolates), Edge Hill (21), and Stratford (10). Ae. vigilax again yielded the majority of these viral isolates. The BF virus outbreak appeared to be associated with several factors. A lack of recent BF virus activity in the region provided a highly susceptible human population, and unusual weather conditions of above average rainfall coupled with high tides resulted in extraordinarily large populations of Ae. vigilax.

Aedes notoscriptus (Skuse) mosquitoes colonized from Brisbane, Queensland, Australia, were fed on blood suspensions containing either Barmah Forest (BF) virus, dengue virus type 1 (DEN-1), dengue virus type 2 (DEN-2), dengue virus type 3 (DEN-3), or dengue virus type 4 (DEN-4). Ae. notoscriptus was shown to be moderately susceptible to oral infection with BF virus, ID50 = 10(3.3) CCID50 per mosquito, with titers in infected mosquitoes peaking 10 d after infection. BF virus transmission occurred between days 5 and 12, with an average rate of 45% (range, 40-50%). For DEN 1-4, Ae. notoscriptus showed low level susceptibility to oral infection, with maximum head and thorax squash infection rates of 8, 5, 13, and 3%, respectively. Rates of infection were notably higher in a north Queensland strain of Aedes aegypti (L.) used as a positive control, with maximum thorax squash infection rates of 58, 68, 78, and 60% for DEN 1-4, respectively. On the basis of these data, it is concluded that Ae. notoscriptus could act as an efficient vector of BF in urban environments, but it is unlikely to be an important vector of DEN.

Barmah Forest virus (BFV) and Ross River virus (RRV) are mosquito-borne viruses with similar vectors and environmental requirements. They cause diseases characterised by arthralgia, arthritis and myalgia, often accompanied by fever and rash. Arthritis is more common and more prominent in RRV disease and rash is more common and florid with BFV infection, although the diseases cannot be reliably distinguished by their clinical symptoms. Diagnosis is based on serological tests and a definite diagnosis of recent infection requires the demonstration of rising titres of IgG. Arthralgia, myalgia and lethargy may continue for at least six months in up to half of patients with RRV, but in only about 10% of patients with BFV. Both diseases are managed symptomatically.

Of the mosquito-borne arboviruses, the encephalitic Murray Valley encephalitis and Kunjin viruses are a major public health concern, but the arthritides Ross River and Barmah Forest viruses are more important in a public health sense, being responsible for a far greater number of infections. Reported cases of Ross River totalled approximately 30,000 during 1991-1996; there have been several widely separated outbreaks of Barmah Forest in recent years and case reports are increasing annually. Surveillance programmes have increased our understanding of the geographic regions, climatic conditions and vector factors associated with viruses. Virus activity is widespread but is often localised, is driven primarily by mosquito abundance and various species are involved; host factors are involved also, but are not well understood. Typically, mosquito populations are governed by availability of habitat and environmental conditions. Models of climate change predict increases in rainfall, tides and temperature for parts of Australia, and such changes have the potential to increase the risk of arbovirus transmission by increasing the distribution and abundance of vectors, and duration of mosquito and arbovirus seasons. However, the amplitude of climate change is uncertain and the ecology of arbovirus transmission is complex. It is likely that some areas will have increases in arbovirus activity and human infection with predicted climate change, but risk of increased transmission will vary with locality, vector, host and human factors.

Barmah Forest virus (BFV) is a mosquito borne (+) ssRNA alphavirus found only in Australia. It causes rash, myalgia and arthralgia in humans and is usually diagnosed serologically. We developed a real-time PCR assay to detect BFV in an effort to improve diagnosis early in the course of infection. The limit of detection was 16 genome equivalents with a specificity of 100%. Fifty five serum samples from BFV-infected patients were tested by the PCR. 52 of 53 antibody-positive samples were PCR negative. Two culture-positive (neutralizing antibody negative) samples were positive on first round PCR, while one sample (IgM and neutralizing antibody strongly positive, IgG negative) was positive on second round PCR, suggesting that viral RNA is detectable and transiently present in early infection. PCR can provide results faster than culture, is capable of high throughput and by sequencing the PCR product strain variants can be characterized.

Background Arthropod-borne viruses are important emerging pathogens world-wide. Viruses transmitted by mosquitoes, such as dengue, yellow fever, and Japanese encephalitis viruses, infect hundreds of millions of people and animals each year. Global surveillance of these viruses in mosquito vectors using molecular based assays is critical for prevention and control of the associated diseases. Here, we report an oligonucleotide DNA microarray design, termed ArboChip5.1, for multi-gene detection and identification of mosquito-borne RNA viruses from the genera Flavivirus (family Flaviviridae), Alphavirus (Togaviridae), Orthobunyavirus (Bunyaviridae), and Phlebovirus (Bunyaviridae). Methodology/Principal Findings The assay utilizes targeted PCR amplification of three genes from each virus genus for electrochemical detection on a portable, field-tested microarray platform. Fifty-two viruses propagated in cell-culture were used to evaluate the specificity of the PCR primer sets and the ArboChip5.1 microarray capture probes. The microarray detected all of the tested viruses and differentiated between many closely related viruses such as members of the dengue, Japanese encephalitis, and Semliki Forest virus clades. Laboratory infected mosquitoes were used to simulate field samples and to determine the limits of detection. Additionally, we identified dengue virus type 3, Japanese encephalitis virus, Tembusu virus, Culex flavivirus, and a Quang Binh-like virus from mosquitoes collected in Thailand in 2011 and 2012. Conclusions/Significance We demonstrated that the described assay can be utilized in a comprehensive field surveillance program by the broad-range amplification and specific identification of arboviruses from infected mosquitoes. Furthermore, the microarray platform can be deployed in the field and viral RNA extraction to data analysis can occur in as little as 12 h. The information derived from the ArboChip5.1 microarray can help to establish public health priorities, detect disease outbreaks, and evaluate control programs., Author Summary Approximately half of the world's population is at risk of viral, mosquito-borne illness such as dengue, yellow fever, Japanese encephalitis, Rift Valley fever, and chikungunya. In the past, these viruses have been regarded as pathogens of the tropics; however, they are emerging as global causes of illness. Very few effective drugs and vaccines have been developed for mosquito-borne viral infections and even less are available to people in resource-limited countries. An important aspect of disease prevention is mosquito surveillance to determine geographical range and seasonal prevalence of the associated viruses. However, there are hundreds of viruses transmitted by mosquitoes that are pathogenic to humans and animals. Using a portable microarray, we developed an assay with the ability to detect most of the known medically important viruses transmitted by mosquitoes. This assay was designed for use in conjunction with broad-range screening tools as a cost effective, rapid method to determine the identity of viruses from infected mosquitoes. To our knowledge, this is the most comprehensive assay to date for field surveillance of mosquito-borne viruses.

Objectives To describe the first reported outbreak of Barmah Forest (BF) virus disease in the south-west of Western Australia. Design Case series correlated with results of arbovirus surveillance. All patients with clinically suspected Ross River (RR) virus infection were serologically tested for antibodies to BF and RR viruses. Home address and date of presentation of patients with serologically confirmed recent infection were recorded. Mosquitoes collected from the districts before and during the BF virus outbreak were identified to species level and tested for virus. Results Twenty-two cases of BF disease were reported from the region between August 1992 and March 1994. Most occurred in the Peel region in the spring and early summer of 1993. Eighteen isolates of BF virus were obtained from three different species of mosquito trapped between January and October 1993. Fifteen were from mosquitoes in the Peel region and a single isolate was from the Perth metropolitan area. No isolates were obtained from the region before 1993. RR virus was not isolated from mosquitoes trapped in the region during the BF virus outbreak. Conclusions Most BF infections were acquired in the Peel region during spring and early summer of 1993. Aedes camptorhynchus mosquitoes were probably the main vectors. The lack of isolations from mosquitoes before 1993 suggests that the virus may have only recently been introduced (or reintroduced) to the region. It was transmitted under conditions that were apparently not conducive to transmission of RR virus.

Mosquito-transmitted alphaviruses causing human rheumatic disease are globally distributed and include chikungunya virus, Ross River virus, Barmah Forest virus, Sindbis virus, o'nyong-nyong virus and Mayaro virus. These viruses cause endemic disease and, occasionally, large epidemics; for instance, the 2004-2011 chikungunya epidemic resulted in 1.4-6.5 million cases, with imported cases reported in nearly 40 countries. The disease is usually self-limiting and characterized by acute and chronic symmetrical peripheral polyarthralgia-polyarthritis, with acute disease usually including fever, myalgia and/or rash. Arthropathy can be debilitating, usually lasts weeks to months and can be protracted; although adequate attention to differential diagnoses is recommended. The latest chikungunya virus epidemic was also associated with some severe disease manifestations and mortality, primarily in elderly patients with comorbidities and the young. Chronic alphaviral rheumatic disease probably arises from inflammatory responses stimulated by the virus persisting in joint tissues, despite robust antiviral immune responses. Serodiagnosis by ELISA is the standard; although international standardization is often lacking. Treatment usually involves simple analgesics and/or NSAIDs, which can provide relief, but better drug treatments are clearly needed. However, the small market size and/or the unpredictable and rapid nature of epidemics present major hurdles for development and deployment of new alphavirus-specific interventions.

New Zealand (NZ) historically has been free of arboviral activity with the exception of Whataroa virus (Togaviridae: Alphavirus), which is established in bird populations and is transmitted by local mosquitoes. This naive situation is threatened by global warming, invasive mosquitoes, and tourism. To determine the threat of selected medically important arboviruses to NZ, vector competence assays were conducted using field collected endemic and introduced mosquito species. Four alphaviruses (Togaviridae): Barmah Forest virus, Chikungunya virus, Ross River virus, and Sindbis virus, and five flaviviruses (Flaviviridae): Dengue virus 2, Japanese encephalitis virus, Murray Valley encephalitis virus, West Nile virus, and Yellow fever virus were evaluated. Results indicate some NZ mosquito species are highly competent vectors of selected arboviruses, particularly alphaviruses, and may pose a threat were one of these arboviruses introduced at a time when the vector was prevalent and the climatic conditions favorable for virus transmission.

Worldwide, mosquito-borne alphaviruses are a major cause of infectious arthritis-like disease, but this has not yet significantly impacted the Americas (1). The arthritogenic alphaviruses include Ross River virus (RRV), chikungunya virus (CHIKV), Sindbis-like viruses (SINV), Barmah Forest virus (BFV), Mayaro virus (MAYV), and o’nyong-nyong virus (ONNV), which are typically found from Europe to central Africa, southern Asia, and in portions of northern South America (2). Infection with these viruses begins as a febrile illness, which in a high percentage of cases, can progress to rheumatic disease, which has been described as polyyarthralgia and/or polyarthritis and can be chronic, flaring, and potentially debilitating (2). In PNAS, Chen et al. (3) present evidence that joint pain and associated symptoms of arthritogenic alphavirus infection (in this case a mouse model of RRV) are strikingly similar to a classical arthritis manifestation, exhibiting cytokine induction profiles reminiscent of rheumatoid arthritis and disruption of osteoblast/osteoclast homeostasis, leading to bone loss.

To the Editor: Buruli ulcer (BU) is a destructive skin disease caused by the toxin-producing environmental pathogen Mycobacterium ulcerans. Since the 1980s, BU has emerged as a major public health problem in rural West and Central Africa (1), where some researchers have suggested a role for aquatic insects as either reservoirs or vectors of M. ulcerans (2,3). However, this hypothesis remains unproven (4). In contrast to the emerging BU–endemic areas in tropical rural West Africa, the climate of the Australian state of Victoria is temperate, yet locally acquired BU also has increased there in recent years (5). In addition, notifications have varied markedly from year to year for reasons not yet explained. During the investigation of a new outbreak of BU in Victoria, we demonstrated that M. ulcerans is detectable by PCR in mosquitoes and that being bitten by mosquitoes increases the odds of being diagnosed with BU (6,7). However, M. ulcerans–positive mosquitoes might reflect only the presence of M. ulcerans in the local environment and play no role in transmission. To further investigate links between BU and mosquitoes, we compared patterns of notifications of BU with other notifiable diseases in Victoria. In particular, we were interested in any association between BU and the locally transmitted vector-borne alphaviruses Ross River virus (RRV) and Barmah Forest virus (BFV). Areas of BU and RRV/BFV endemicity overlap geographically, but areas with RRV and BFV are more extensive and include inland river systems where BU has not so far been reported. Notification data for RRV, BFV, and other notifiable infections in Victoria are publicly available (8). Although BU was not made notifiable until January 2004 (before which notification was voluntary), since early 2000, most diagnoses were confirmed by culture or PCR at the Victorian Infectious Diseases Reference Laboratory, from which we obtained data for this report. Our analysis showed that in the last 7 years (2002–2008), BU notifications correlated with combined RRV/BFV notifications (r2 = 0.52, p = 0.06) (Figure). During the same period, no correlation was observed with tuberculosis, the other important mycobacterial disease in Victoria (r2 = 0.12, p = 0.43); legionellosis, caused by a nonvectored water-associated pathogen (r2 = 0.04, p = 0.66); or any other notifiable infectious disease (data not shown). Figure Numbers of cases per 100,000 inhabitants for selected notifiable diseases, Victoria, Australia, 2000–2008. Buruli ulcer is shown on the right y axis, other diseases on the left y axis. RRV, Ross river virus; BFV, Barmah Forest virus; TB, tuberculosis. ... Although the environmental reservoir and mode of transmission of M. ulcerans remain unknown, mosquitoes are well known for transmitting RRV and BFV to humans, and year-to-year variation in incidence of these vector-borne viral infections is linked to changes in mosquito numbers (9,10). We are not implying that M. ulcerans, RRV, and BFV are transmitted simultaneously from the same reservoir species to the same humans or by the same mosquitoes. Also, environmental conditions that promote outbreaks of RRV/BFV infection might promote BU outbreaks without any other connection. However, we believe the correlation we have identified between BU and other mosquito-borne diseases is striking and further strengthens the link between mosquitoes and the transmission of M. ulcerans in Victoria.

To determine the presence of arboviruses in mosquito populations from major urban areas of eastern Australia, a total of 67,825 mosquitoes, representing ∼60 species, was collected and tested from Cairns, Brisbane, and Sydney between January 2005 and April 2008. Mosquito pools were screened by inoculation onto mosquito cell cultures and the detection of viral antigen using a panel of flavivirus and alphavirus monoclonal antibodies in an enzyme-linked immunosorbent assay. Suspect positive samples were confirmed using virus-specific real-time reverse transcriptasepolymerase chain reaction assays. No flaviviruses were detected, but 2 alphaviruses were isolated from mosquito pools collected from Cairns, with 1 Barmah Forest virus isolate from a pool of 100 Aedes vigilax and 1 Ross River virus isolate from a pool of 83 Verrallina carmenti. In addition, a single Aedes alternans collected from Sydney yielded an isolate most similar to Stretch Lagoon virus, a newly described virus from the genus Orbivirus. These results suggest that during the study, arboviruses were circulating at a low level in the areas sampled. The findings from this study will promote public health awareness of the risk of arboviruses in urban areas, leading to more informative public health campaigns to safeguard the Australian public. Refereed/Peer-reviewed

The purpose of the present article is to present a review of the Ross River virus (RRV) and Barmah Forest virus (BFV) literature in relation to potential implications for future disease in tropical northern Australia. Ross River virus infection is the most common and most widespread arboviral disease in Australia, with an average of 4,800 national notifications annually. Of recent concern is the sudden rise in BFV infections; the 2005–2006 summer marked the largest BFV epidemic on record in Australia, with 1,895 notifications. Although not life-threatening, infection with either virus can cause arthritis, myalgia, and fatigue for 6 months or longer, resulting in substantial morbidity and economic impact. The geographic distribution of mosquito species and their seasonal activity is determined in large part by temperature and rainfall. Predictive models can be useful tools in providing early warning systems for epidemics of RRV and BFV infection. Various models have been developed to predict RRV outbreaks, but these appear to be mostly only regionally valid, being dependent on local ecological factors. Difficulties have arisen in developing useful models for the tropical northern parts of Australia, and to date no models have been developed for the Northern Territory. Only one model has been developed for predicting BFV infections using climate and tide variables. It is predicted that the exacerbation of current greenhouse conditions will result in longer periods of high mosquito activity in the tropical regions where RRV and BFV are already common. In addition, the endemic locations may expand further within temperate regions, and epidemics may become more frequent in those areas. Further development of predictive models should benefit public health planning by providing early warning systems of RRV and BFV infection outbreaks in different geographical locations.

Togaviruses are major causes of human disease internationally. They include the alphaviruses, which are distributed worldwide and cause illness in many hundreds of thousands of people each year. Clinical illnesses caused by togaviruses range from mild fever to severe diseases, such as encephalitis and hemorrhagic fever. One of the commonest clinical manifestations of alphavirus infections is fever accompanied by rash and/or arthritis. The alphaviruses causing these illnesses are all closely related and include Chikungunya virus, O’nyong-nyong virus, Ross River virus, Barmah Forest virus, Semliki Forest virus, Sindbis virus (including Ockelbo), and Mayaro virus. In addition, the burden of acute infection and the occasional cases of severe illness or chronic illness with joint pain is a common consequence of infection. This article reviews the epidemiology and ecology of these viruses, the diseases they cause, and their pathogenesis, diagnosis of infection, and treatment.

Barmah Forest virus is a mosquito-borne agent (alphavirus) reported to cause both clinical and subclinical infections in New South Wales. This report describes 29 cases of clinical Barmah Forest virus infection diagnosed between July 1988 and March 1989 (21 from Queensland, six from New South Wales and two from Victoria) and provides evidence of extensive subclinical infection with this virus (0.23% of the population per annum) throughout Queensland. It also includes a description of the first isolation of Barmah Forest virus from a patient. Data obtained in the course of the study suggest that Barmah Forest virus infections may not be diagnosed correctly in many instances because of the similarity of the symptoms of this disease to those of epidemic polyarthritis and the small number of laboratories providing the necessary serological services.

Clinical infection with Barmah Forest virus (BFV) is becoming increasingly recognised with serological testing. We report the first case of glomerulonephritis after BFV infection. The patient required diuretic and antihypertensive therapy, but made an almost complete recovery. BFV infection should be considered in the differential diagnosis of glomerulonephritis.

Arthritogenic alphaviruses are globally distributed mosquito-borne RNA viruses causing epidemics of polyarthritis/arthralgia, with disease emerging or reemerging and increasingly being reported in travelers. This article summarizes the current knowledge of these diseases, focusing on recent developments in the understanding of Ross River virus disease. RECENT FINDINGS: Alphaviral arthritides have often been blamed for protracted chronic illnesses. However, validated quality-of-life questionnaires and exhaustive searches for differential diagnoses showed that Ross River virus disease, although severe at onset, progressively resolved over 3 to 6 months. Many patients did experience long-term disease lasting more than 12 months, but in nearly all cases this was due to other conditions, primarily unrelated rheumatic conditions or depression. There is no indication that alphaviral arthritides predispose to other conditions; thus, patients whose Ross River virus disease has actually resolved may be underdiagnosed for other conditions. Ross River virus polyarthritis probably arises from inflammation associated with productive viral infections in synovial macrophages, which persist despite neutralizing antibodies and antiviral cytokine responses. Persistence may be facilitated by downregulation of cytokine responses by virus-antibody complexes binding to Fc receptors and induction of interleukin-10. How virus escapes neutralizing antibodies remains unclear but may involve phagocytosis of apoptotic virus-infected cells and infection of the phagocyte via the phagosome. SUMMARY: Diagnosis of alphaviral arthritides is complicated by nonspecific symptoms and the lack of commercial serodiagnostic kits, except for Ross River and Barmah Forest virus infections in Australia. Differential diagnoses should be actively pursued, especially if symptoms persist. Treatment with nonsteroidal anti-inflammatory drugs appears largely effective, with no evidence of long-term sequelae or relapse.

Brushtail possums, Trichosurus vulpecula Kerr, were experimentally infected with Ross River (RR) or Barmah Forest (BF) virus by Aedes vigilax (Skuse) mosquitoes. Eight of 10 animals exposed to RR virus developed neutralizing antibody, and 3 possums developed high viremia for48 hr after infection, sufficient to infect recipient mosquitoes. Two of 10 animals exposed to BF virus developed neutralizing antibody. Both infected possums maintained detectable neutralizing antibody to BF for at least 45 days after infection (log neutralization index2.0 at 45 days). Eight possums did not develop neutralizing antibody to BF despite exposure to infected mosquitoes. These results suggest that T. vulpecula may potentially act as a reservoir species for RR in urban areas. However, T. vulpecula infected with BF do not develop viremia sufficient to infect mosquitoes and are unlikely to be important hosts for BF.

▪ Abstract Ross River virus is the most common mosquito-borne pathogen in Australia, and approximately 5000 human cases are reported annually. The infection is not fatal, but there is considerable morbidity associated with a debilitating polyarthritis that is the major symptom. The virus is annually active in most regions of Australia, but exists as strains that vary in virulence. Native macropods are thought to be the natural vertebrate hosts, although horses and humans may be involved during epidemic activity, and vertical transmission of the virus occurs in mosquitoes. Different mosquito species are involved as vectors in various regions and in different seasonal and environmental conditions. In coastal areas the saltmarsh mosquitoes Aedes camptorhynchus and Ae. vigilax are the most important vectors in southern and northern regions, respectively, whereas in inland areas Culex annulirostris is the most important vector, although various Aedes species can be involved depending on region and conditions, and the epidemiology of the disease and vector control imperatives vary with circumstance concomitantly.

Mosquito-borne arboviruses are an important public health issue in Australia. The alphaviruses Ross River and Barmah Forest virus are widespread and active annually, and cause debilitating polyarthritis. The flaviviruses Murray Valley encephalitis, Kunjin and Japanese encephalitis virus are restricted in distribution and activity but may cause life-threatening illness, and dengue viruses are active in some areas.

Aedes vigilax (Skuse) mosquitoes colonized from Townsville, Queensland, Australia, were fed on blood containing Barmah Forest virus (BF) isolated from the same species. The colony was susceptible to infection, with an ID50 of 10(2.6) CCID50 per mosquito. Infection and transmission rates for mosquitoes fed 10(3.5) CCID50 virus per mosquito varied from 58 to 100% and 36 to 100%, respectively, between days 3 and 13 after infection. Titers in infected mosquitoes were high by 5 d after infection and did not vary substantially thereafter. Virus transmission to suckling mice occurred from 3 d after infection. This work establishes that Ae. vigilax is a competent vector of BF.

An increase in the incidence of polyarthritis caused by Barmah Forest (BF) virus, and its recent emergence into Western Australia, prompted a study of the molecular epidemiology of this Australian mosquito-borne alphavirus. The nucleotide sequence of a 500-basepair region of the 3' end of the envelope (E2) gene of the prototype BF virus strain (BH2193) was compared with other members of the alphavirus genus, and to a panel of isolates of BF virus collected more for than 20 years from different geographic regions of Australia. The BF virus was shown to be genetically distinct from other members of the alphavirus genus. A high degree of sequence homology (98-100%) was found between the BF isolates, with no evidence of geographic or temporal divergence. This nucleotide homogeneity was similar to that observed with other Australian mosquito-borne viruses with avian vertebrate hosts, such as Sindbis, Murray Valley, and Kunjin viruses, but it contrasts to the heterogeneity reported for Ross River virus, an alphavirus with mammalian vertebrate hosts.