The recent emergence and spread of dengue hemorrhagic fever in the Americas have been a major source of concern. Efforts to control this disease are dependent on understanding the pathogenicity of dengue viruses and their transmission dynamics. Pathogenicity studies have been hampered by the lack of in vitro or in vivo models of severe dengue disease. Alternatively, molecular epidemiologic studies which associate certain dengue virus genetic types with severe dengue outbreaks may point to strains with increased pathogenicity. The comparison of nucleotide sequences (240 bp) from the E/NS1 gene region of the dengue virus genome has been shown to reflect evolutionary relationships and geographic origins of dengue virus strains. This approach was used to demonstrate an association between the introduction of two distinct genotypes of dengue type 2 virus and the appearance of dengue hemorrhagic fever in the Americas. Phylogenetic analyses suggest that these genotypes originated in Southeast Asia and that they displaced the native, American genotype in at least four countries. Vaccination and other control efforts should therefore be directed at decreasing the transmission of these "virulent" genotypes.

The vertebrate hosts of 672 blood-engorged Culex quinquefasciatus Say, collected in Harris County, Texas, during 2005, were identified by nucleotide sequencing PCR products of the cytochrome b gene. Analysis revealed that 39.1% had acquired blood from birds, 52.5% from mammals, and 8.3% were mixed avian and mammalian blood meals. Most frequent vertebrate hosts were dog (41.0%), mourning dove (18.3%), domestic cat (8.8%), white-winged dove (4.3%), house sparrow (3.2%), house finch (3.0%), gray catbird (3.0%), and American robin (2.5%). Results are interpreted in conjunction with concurrent avian and mosquito West Nile virus (WNV) surveillance activities in Harris County. We conclude that Cx. quinquefasciatus is an opportunistic feeder and principal mosquito vector of WNV in this metropolitan area; however, transmission by other mosquito species or by other modes of infection, such as ingestion, must account for the high WNV infection rates among local blue jays and American crows.

Mayaro and Una viruses (MAYV, UNAV) are mosquito-borne alphaviruses that may cause an acute febrile illness characterized by headache, retro-orbital pain, and rash that may progress to a severe and prolonged arthralgia. MAYV was first isolated in Trinidad in 1954, and UNAV was first identified in northern Brazil in 1959. Since then, numerous isolates of these agents have been made from humans, wild vertebrates, and mosquitoes in several countries in northern South America. Serological evidence suggests that these viruses are also present in portions of Central America. Because little is known about the natural transmission cycle of MAYV and virtually nothing is known about UNAV transmission, 63 isolates covering the known geographic and temporal ranges were used in phylogenetic analyses to aid in understanding the molecular epidemiology. Approximately 2 kb from the E1 and E2 glycoprotein genes and the complete 3' non-coding region were sequenced. Phylogenetic analyses of these sequences indicated that two distinct genotypes of MAYV exist with a distinct clade consisting exclusively of UNAV (previously designated as a subtype of MAYV). One MAYV genotype (genotype D) contains isolates from Trinidad and the northcentral portion of South America including Peru, French Guiana, Surinam, Brazil, and Bolivia. All of these isolates are highly conserved with a nucleotide divergence of < 6%. The second MAYV genotype (genotype L) contains isolates only from Brazil that are highly conserved (< 4% nucleotide divergence) but are quite distinct (15-19%) from the first genotype isolates. These analyses provide possible explanations for the natural ecology and transmission of MAYV and UNAV.

A new virus was isolated from three independent pools of Phlebotomus perniciosus sandflies (Diptera; Psychodidae) trapped in two regions of southeastern France, located 90 miles apart. Microscopic, antigenic and genetic analyses indicate that this novel virus belongs to the genus Phlebovirus in the family Bunyaviridae. The new virus is designated Massilia virus since the first isolate was obtained from sandflies collected in the suburban area of Marseille. The complete genome sequence was determined and used to compare the genetic and phylogenetic relationships of Massilia virus with other phleboviruses. Genetic and antigenic properties were employed to address whether or not Massilia virus should be considered a new species within the genus, or a member of a previously recognized species. Cerebrospinal fluid specimens, collected from local patients with central nervous system infections during the previous four-year period were tested for the presence of Massilia virus RNA, but gave negative results. In conclusion, Massilia virus is proposed as a member of the Sandfly fever Naples virus complex; its public health importance has yet to be determined.

As part of a comprehensive study on the ecology of arthropod-borne viruses in the Amazon Basin region of Peru, we assayed 539,694 mosquitoes captured in Loreto Department, Peru, for arboviruses. Mosquitoes were captured either by dry ice-baited miniature light traps or with aspirators while mosquitoes were landing on human collectors, identified to species, and later tested on Vero cells for virus. In total, 164 virus isolations were made and included members of the Alphavirus (eastern equine encephalomyelitis, Trocara, Una, Venezuelan equine encephalomyelitis, and western equine encephalomyelitis viruses), Flavivirus (Ilheus and St. Louis encephalitis), and Orthobunyavirus (Caraparu, Itaqui, Mirim, Murutucu, and Wyeomyia viruses) genera. In addition, several viruses distinct from the above-mentioned genera were identified to the serogroup level. Eastern equine encephalomyelitis virus was associated primarily with Culex pedroi Sirivanakarn & Belkin, whereas Venezuelan equine encephalomyelitis virus was associated primarily with Culex gnomatos Sallum, Huchings & Ferreira. Most isolations of Ilheus virus were made from Psorophora ferox (Von Humboldt). Although species of the Culex subgenus Melanoconion accounted for only 45% of the mosquitoes collected, 85% of the virus isolations were made from this subgenus. Knowledge of the viruses that are being transmitted in the Amazon Basin region of Peru will enable the development of more effective diagnostic assays, more efficient and rapid diagnoses of clinical illnesses caused by these pathogens, risk analysis for military/civilian operations, and development of potential disease control measures.

The genus Phlebovirus of the family Bunyaviridae consists of approximately 70 named viruses, currently assigned to nine serocomplexes (species) based on antigenic similarities. Sixteen other named viruses that show little serologic relationship to the nine recognized groups are also classified as tentative species in the genus. In an effort to develop a more precise classification system for phleboviruses, we are attempting to sequence most of the named viruses in the genus with the goal of clarifying their phylogenetic relationships. In this report, we describe the serologic and phylogenetic relationships of 13 viruses that were found to be members of the Candiru serocomplex; 6 of them cause disease in humans. Analysis of full genome sequences revealed branching inconsistencies that suggest five reassortment events, all involving the M segment, and thus appear to be natural reassortants. This high rate of reassortment illustrates the inaccuracy of a classification system based solely on antigenic relationships.

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Numerous tickborne viruses, including Dhori virus and foot-and-mouth disease virus, were isolated.