RESUMO
BACKGROUND: Alkhumra hemorrhagic fever virus (AHFV) is a newly described flavivirus first isolated in 1994-1995 from the Alkhumra district south of Jeddah, Saudi Arabia. Subsequently, the virus was also isolated from Makkah (2001-2003) and Najran (2008-2009), Saudi Arabia. METHODS: The full-length genome of an AHFV strain isolated from patients in Najran (referred to as AHFV/997/NJ/09/SA) was PCR amplified and sequenced, and compared with the sequences of 18 other AHFV strains previously isolated from Jeddah and Makkah, dengue virus (DENV), Kyasanur forest disease virus (KFDV), Langat virus, Omsk hemorrhagic fever virus (OHFV), and tick-borne encephalitis virus (TBEV). RESULTS: The RNA of the AHFV/997/NJ/09/SA strain was found to have 10,546 nucleotides encoding for a single 3,416-amino acid polyprotein, whereas the previously reported AHFV strains were composed of 10,685-10,749 nucleotides. The AHFV/997/NJ/09/SA strain showed about 99% homology with the previously reported AHFV strains. The KFDV, Langat virus, TBEV, and OHFV isolates formed a separate cluster with a variable homology. The most important variations were observed in the core protein and NS4a gene sequences of two AHFV isolates. CONCLUSION: The variation in the number of nucleotides and phylogenetic analysis with the other AHFV isolates could have resulted from recombination of circulating virus strains.
Assuntos
Vírus da Encefalite Transmitidos por Carrapatos/genética , Encefalite Transmitida por Carrapatos/virologia , Genoma Viral , RNA Viral/genética , Análise de Sequência de DNA , Análise por Conglomerados , Vírus da Encefalite Transmitidos por Carrapatos/isolamento & purificação , Humanos , Filogenia , Poliproteínas/genética , Arábia Saudita , Homologia de SequênciaRESUMO
BACKGROUND: Although Alkhumra haemorrhagic fever virus (AHFV) has been isolated from ticks, epidemiological data suggest that it is transmitted from livestock to humans by direct contact with animals or by mosquito bites, but not by ticks. This study was carried out to assess the ability of the virus to replicate in tick cells in vitro. METHODS: AHFV was inoculated into cell lines derived from the hard ticks Hyalomma anatolicum (HAE/CTVM9) and Rhipicephalus appendiculatus (RAE/CTVM1) and the soft tick Ornithodoros moubata (OME/CTVM24). Inoculated cells were directly examined every week for 4 weeks by real-time reverse transcription PCR and by IFAT using polyclonal antibodies. RESULTS: AHFV RNA was detected in all three inoculated tick cell lines throughout the 4-week observation period at levels up to almost twice that of the inoculum, but none of them exhibited a cytopathic effect. AHFV antigen could be detected in all three cell lines by IFAT. Titration of tick cell culture suspension in LLC-MK2 cells yielded AHFV titres of 10(6.6) 50% tissue culture infective dose (TCID50)/ml for OME/CTVM24 and 10(5.5) TCID50/ml for RAE/CTVM1 cells after 4 weeks of culturing; no viable virus was detected in HAE/CTVM9 cells. CONCLUSION: This is the first description of propagation of AHFV in tick cells.