The First Isolation and Whole Genome Sequencing of Murray Valley Encephalitis Virus from Cerebrospinal Fluid of a Patient with Encephalitis.

Murray Valley Encephalitis virus (MVEV) is a mosquito-borne Flavivirus. Clinical presentation is rare but severe, with a case fatality rate of 15⁻30%. Here we report a case of MVEV from the cerebrospinal fluid (CSF) of a patient in the Northern Territory in Australia. Initial diagnosis was performed using both MVEV-specific real-time, and Pan-Flavivirus conventional, Polymerase Chain Reaction (PCR), with confirmation by Sanger sequencing. Subsequent isolation, the first from CSF, was conducted in Vero cells and the observed cytopathic effect was confirmed by increasing viral titre in the real-time PCR. Isolation allowed for full genome sequencing using the Scriptseq V2 RNASeq library preparation kit. A consensus genome for VIDRL-MVE was generated and phylogenetic analysis identified it as Genotype 2. This is the first reported isolation, and full genome sequencing of MVEV from CSF. It is also the first time Genotype 2 has been identified in humans. As such, this case has significant implications for public health surveillance, epidemiology, and the understanding of MVEV evolution.

Fatal Infection with Murray Valley Encephalitis Virus Imported from Australia to Canada, 2011.

Murray Valley encephalitis virus (MVEV), a flavivirus belonging to the Japanese encephalitis serogroup, can cause severe clinical manifestations in humans. We report a fatal case of MVEV infection in a young woman who returned from Australia to Canada. The differential diagnosis for travel-associated encephalitis should include MVEV, particularly during outbreak years.

The Molecular Epidemiology and Evolution of Murray Valley Encephalitis Virus: Recent Emergence of Distinct Sub-lineages of the Dominant Genotype 1.

BACKGROUND: Recent increased activity of the mosquito-borne Murray Valley encephalitis virus (MVEV) in Australia has renewed concerns regarding its potential to spread and cause disease. METHODOLOGY/PRINCIPAL FINDINGS: To better understand the genetic relationships between earlier and more recent circulating strains, patterns of virus movement, as well as the molecular basis of MVEV evolution, complete pre-membrane (prM) and Envelope (Env) genes were sequenced from sixty-six MVEV strains from different regions of the Australasian region, isolated over a sixty year period (1951-2011). Phylogenetic analyses indicated that, of the four recognized genotypes, only G1 and G2 are contemporary. G1 viruses were dominant over the sampling period and found across the known geographic range of MVEV. Two distinct sub-lineages of G1 were observed (1A and 1B). Although G1B strains have been isolated from across mainland Australia, Australian G1A strains have not been detected outside northwest Australia. Similarly, G2 is comprised of only Western Australian isolates from mosquitoes, suggesting G1B and G2 viruses have geographic or ecological restrictions. No evidence of recombination was found and a single amino acid substitution in the Env protein (S332G) was found to be under positive selection, while several others were found to be under directional evolution. Evolutionary analyses indicated that extant genotypes of MVEV began to diverge from a common ancestor approximately 200 years ago. G2 was the first genotype to diverge, followed by G3 and G4, and finally G1, from which subtypes G1A and G1B diverged between 1964 and 1994. CONCLUSIONS/SIGNIFICANCE: The results of this study provides new insights into the genetic diversity and evolution of MVEV. The demonstration of co-circulation of all contemporary genetic lineages of MVEV in northwestern Australia, supports the contention that this region is the enzootic focus for this virus.

Isolation of arboviruses from mosquitoes (Diptera: Culicidae) collected from the Gulf Plains region of northwest Queensland, Australia.

As part of investigations into Japanese encephalitis (JE) virus and related flaviviruses in northern Australia, 153,529 mosquitoes were collected and processed for virus isolation from the Gulf Plains region of northwest Queensland. Collections from within 30 km of each of the townships of Croydon, Normanton and Karumba yielded 3,087 (2.0%), 66,009 (43.0%), and 84,433 (55.0%) mosquitoes, respectively, from which 16 viruses were isolated. Four isolates of Murray Valley encephalitis (MVE), two of Kunjin (KUN), three of Ross River (RR), and one of Sindbis (SIN) viruses were obtained from Culex sitiens subgroup mosquitoes. Molecular identification of the mosquito species composition of these virus positive pools revealed that most isolates were from pools containing mainly Culex annulirostris Skuse and low numbers of Culex palpalis (Taylor). Only three pools, one each of MVE, KUN, and RR, were from mosquitoes identified exclusively as Cx. annulirostris. Other viruses isolated include one Edge Hill virus from Ochlerotatus normanensis (Taylor), an isolate of SIN from Anopheles meraukensis Venhuis, two isolates of RR from Anopheles amictus Edwards, and single isolates of RR from Anopheles bancroftii Giles andAedes lineatopennis (Ludlow). The isolate of RR from Ae. lineatopennis was the first reported from this species. The public health implications of these isolations in the Gulf Plains region are discussed briefly.