Molecular and serologic methods usable for the biological diagnosis of Coltivirus infection are reported. We designed a multiplex reverse transcription-polymerase chain reaction system that allowed ...the simultaneous and specific amplification of three genomic segments from as little as 0.01 plaque-forming units. Another system in the S2 viral segment permitted the differential diagnosis of American and European viral isolates. We also discuss some improvements of previous ELISAs, and the results obtained with paired sera from Colorado tick fever (CTF) virus-infected individuals. Western blot analysis was developed that allowed the detection of antibodies to a 38-kD viral protein in all tested sera. It also enabled the detection of anti-CTF virus antibodies in ELISA-negative sera. Specific IgM antibodies against a synthetic viral peptide could be detected in sera at the acute stage of the infection. Together, these results should permit the diagnosis of Coltivirus infection at any stage of the pathology.
H Attoui, RN Charrel, F Billoir, JF Cantaloube, P de Micco and X de Lamballerie
Laboratoire de Virologie Moleculaire, Tropicale et Transfusionnelle, Faculte de Medecine de Marseille, Universite de la ...Mediterranee, Marseille, France.
In this study, the basis for the classification of virus isolates grouped
within the genus Coltivirus, family Reoviridae, is discussed. Sequences of
dsRNA segments from American (segments 9-12), European (segment 12) and
Asian (segments 7-12) isolates were characterized and polythetic criteria
were defined for their taxonomic classification. These criteria (including
sequence analysis) permitted the different species to be distinguished and
classified into two groups. In both groups, subgroups were defined
according to the degree of homology between the genomic sequences. American
and European isolates are classified within group A, which includes
subgroups A1 (Colorado tick fever virus species) and A2 (Eyach virus
species). Asian isolates are classified in group B, which includes
subgroups B1 (JKT-7075 virus species) and B2 (JKT-6423 virus species). The
proteins encoded by the sequenced genomic segments were analysed. This
allowed the identification of dsRNA binding domains in the proteins encoded
by segment 8 of subgroup B1 isolates and segment 12 of subgroup B2
isolates. A conserved pattern of amino acids in segment 7 of group B
isolates matched sequences found in the catalytic domains of protein
kinases.
An upgrade of the near detector of the T2K long baseline neutrino oscillation experiment is currently being conducted. This upgrade will include two new Time Projection Chambers, each equipped with ...16 charge readout resistive Micromegas modules. A procedure to validate the performance of the detectors at different stages of production has been developed and implemented to ensure a proper and reliable operation of the detectors once installed. A dedicated X-ray test bench is used to characterize the detectors by scanning each pad individually and to precisely measure the uniformity of the gain and the deposited energy resolution over the pad plane. An energy resolution of about 10% is obtained. A detailed physical model has been developed to describe the charge dispersion phenomena in the resistive Micromegas anode. The detailed physical description includes initial ionization, electron drift, diffusion effects and the readout electronics effects. The model provides an excellent characterization of the charge spreading of the experimental measurements and allowed the simultaneous extraction of gain and RC information of the modules.
The group-B of genus
Coltivirus encompasses isolates from humans, ticks or mosquitoes collected in Indonesia and China. Subgroup-B1 includes the strain JKT/dsR-7075 and subgroup-B2 strains ...JKT/dsR-6423, JKT/dsR-6969, JKT/dsR–7043 and the Banna virus. Data are described for the PCR-based diagnosis of infection by group B coltiviruses. Sets of primers were designed from the sequences of the 7th, 9th and 12th viral segments and RT–PCR assays were developed. Consensus primers permitted the detection of all known isolates of subgroup 1 or 2. Viral strains could be characterised further using primers specific for type B2a or B2b, or based on the length of the amplification products. All primers gave negative results when using RNAs from Orbiviruses or Group-A coltiviruses. These primers permitted the detection of Group-B coltiviruses-RNA in infected mouse blood at the acute stage of the disease. Accordingly, they could be used for the diagnosis of infection in humans.
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