Cacipacore virus as an emergent mosquito-borne Flavivirus.

INTRODUCTION:: Cacipacore virus (CPCV), a possible bird-associated flavivirus, has yet to be detected in mosquitoes. Our purpose is examining CPCV in mosquitoes from the Amazon region of Brazil. METHODS:: Approximately 3,253 Culicidae (grouped into 264 pools) were collected from the Amazon region during 2002-2006 and analyzed using a Flavivirus genus-specific reverse transcription- polymerase chain reaction followed by nested polymerase chain reaction assay and by nucleotide sequencing of amplicons. RESULTS:: Nucleotide sequences from five mosquito samples showed high similarity to the those of CPCV originally isolated in the Amazon region. CONCLUSIONS:: This is the first report of CPCV-infected mosquitoes which has implications on the arbovirus maintenance in nature and transmission to man.

Cacipacore virus as an emergent mosquito-borne Flavivirus

Abstract INTRODUCTION: Cacipacore virus (CPCV), a possible bird-associated flavivirus, has yet to be detected in mosquitoes. Our purpose is examining CPCV in mosquitoes from the Amazon region of Brazil. METHODS: Approximately 3,253 Culicidae (grouped into 264 pools) were collected from the Amazon region during 2002-2006 and analyzed using a Flavivirus genus-specific reverse transcription- polymerase chain reaction followed by nested polymerase chain reaction assay and by nucleotide sequencing of amplicons. RESULTS: Nucleotide sequences from five mosquito samples showed high similarity to the those of CPCV originally isolated in the Amazon region. CONCLUSIONS: This is the first report of CPCV-infected mosquitoes which has implications on the arbovirus maintenance in nature and transmission to man.

A real-time RT-PCR for rapid detection and quantification of mosquito-borne alphaviruses.

Mosquito-borne alphaviruses are widely distributed throughout the world, causing important human illnesses. Therefore, the development of methods to enable early diagnosis of infections by alphavirus is essential. We show here the development and evaluation of a quantitative real-time RT-PCR using genus-specific primers to the nsP1 viral gene of all mosquito-borne alphaviruses. The specificity and sensitivity of the assay were tested using seven alphaviruses and RNA transcribed from Venezuelan equine encephalitis virus. The detection limits of real-time RT-PCR ranged from 10 to 76 copies per ml. The melting temperature (TM) values for amplification of the alphavirus genomes were 83.05 °C and 85.28 °C. Interestingly, the assay suggested the possibility the arthritogenic alphaviruses with TM peaks of 84.83 to 85.28 °C and encephalitic alphaviruses of 83.34 °C to 84.68 °C could be discriminated both diseases. Real-time RT-PCR may prove very useful for the screening and preliminary diagnosis in outbreaks and surveillance studies as well as for measuring the viral load in pathogenesis studies.

A real-time reverse transcriptase polymerase chain reaction for detection and quantification of Vesiculovirus.

Vesiculoviruses (VSV) are zoonotic viruses that cause vesicular stomatitis disease in cattle, horses and pigs, as well as sporadic human cases of acute febrile illness. Therefore, diagnosis of VSV infections by reliable laboratory techniques is important to allow a proper case management and implementation of strategies for the containment of virus spread. We show here a sensitive and reproducible real-time reverse transcriptase polymerase chain reaction (RT-PCR) for detection and quantification of VSV. The assay was evaluated with arthropods and serum samples obtained from horses, cattle and patients with acute febrile disease. The real-time RT-PCR amplified the Piry, Carajas, Alagoas and Indiana Vesiculovirus at a melting temperature 81.02 ± 0.8ºC, and the sensitivity of assay was estimated in 10 RNA copies/mL to the Piry Vesiculovirus. The viral genome has been detected in samples of horses and cattle, but not detected in human sera or arthropods. Thus, this assay allows a preliminary differential diagnosis of VSV infections.

A real-time reverse transcriptase polymerase chain reaction for detection and quantification of Vesiculovirus

Vesiculoviruses (VSV) are zoonotic viruses that cause vesicular stomatitis disease in cattle, horses and pigs, as well as sporadic human cases of acute febrile illness. Therefore, diagnosis of VSV infections by reliable laboratory techniques is important to allow a proper case management and implementation of strategies for the containment of virus spread. We show here a sensitive and reproducible real-time reverse transcriptase polymerase chain reaction (RT-PCR) for detection and quantification of VSV. The assay was evaluated with arthropods and serum samples obtained from horses, cattle and patients with acute febrile disease. The real-time RT-PCR amplified the Piry, Carajas, Alagoas and Indiana Vesiculovirus at a melting temperature 81.02 ± 0.8ºC, and the sensitivity of assay was estimated in 10 RNA copies/mL to the Piry Vesiculovirus. The viral genome has been detected in samples of horses and cattle, but not detected in human sera or arthropods. Thus, this assay allows a preliminary differential diagnosis of VSV infections.

Detection of hantavirus in bats from remaining rain forest in São Paulo, Brazil.

BACKGROUND: The significant biodiversity found in Brazil is a potential for the emergence of new zoonoses. Study in some places of the world suggest of the presence to hantavirus in tissues of bats. Researches of hantavirus in wildlife, out rodents, are very scarce in Brazil. Therefore we decided to investigate in tissues of different species of wild animals captured in the same region where rodents were detected positive for this virus. The present work analyzed ninety-one animals (64 rodents, 19 opossums, and 8 bats) from a region of the Atlantic forest in Biritiba Mirin City, São Paulo State, Brazil. Lungs and kidneys were used for RNA extraction. FINDINGS: The samples were screened for evidence of hantavirus infection by SYBR-Green-based real-time RT-PCR. Sixteen samples positive were encountered among the wild rodents, bats, and opossums. The detection of hantavirus in the lungs and kidneys of three marsupial species (Micoureus paraguayanus, Monodelphis ihering, and Didelphis aurita) as well in two species of bats (Diphylla ecaudata and Anoura caudifer) is of significance because these new hosts could represent an important virus reservoirs. CONCLUSIONS: The analysis of nucleotide sequences of the partial S segment revealed that these genes were more related to the Araraquara virus strains. This work reinforces the importance of studying hantavirus in different animal species and performing a continued surveillance before this virus spreads in new hosts and generated serious problems in public health.