PMC:4504098 / 9244-11933
Annnotations
{"target":"https://pubannotation.org/docs/sourcedb/PMC/sourceid/4504098","sourcedb":"PMC","sourceid":"4504098","source_url":"https://www.ncbi.nlm.nih.gov/pmc/4504098","text":"We detected both serologic and molecular evidence of RESTV infection in Philippine bats. RESTV RNA in the oropharyngeal swab of three Miniopterus schreibersii clustered phylogenetically with the 2008 pig-derived sequences and the historic 1989 Philippine primate-derived sequence. Sequence from all three bats was identical, and aligned most closely with the 2008 pig isolate from a farm (Farm A) in Bulacan Province [14], less than 40 km from the bat sampling location. All sequenced products from bats had the single nucleotide change; all positive control and related material held at AAHL did not have the change. Limited variation is not surprising with an assay targeting a conserved region of the NP gene following recent introduction of infection a population. While the high Ct values from the qPCR indicate the assay is approaching the limits of detection with these samples, a number of factors support the veracity of the findings. At the laboratory level, the repeatability of positive findings using qPCR in both pooled and individual specimens, the repeatability of positive findings in archived duplicate specimens, the corroboration by conventional PCR, and the direct sequencing results. We detected RNA in archived duplicate samples of two of the three positive M. schreibersii; these duplicates were made in the Philippines, stored at the PAHC, and forwarded to AAHL and tested in a separate run 12 months after the first test. At the epidemiological level, the clustering of the positive samples in one species, in one cave, and in one sampling event is consistent with a current infection dynamic. The inability of the PAHC duplicate sample testing to fully corroborate the first results can be plausibly explained by reduced volume of the duplicate samples (less than half the original sample volume), meaning less extracted RNA in the duplicate assay. In addition, there was potential nucleic acid degradation as the duplicate samples had been stored for a considerably longer period than the original samples. The qPCR findings also suggest the possible presence of the virus in additional sympatric taxa (M. australis, Cynopterus brachyotis and Chaerephon plicata) and in additional locations (Puning Cave), but limits of detection issues and small sample volumes again precluded corroboration and sequencing. Limited blood volumes also constrained the use of additional assays such as cell culture and next generation sequencing. For context, around 85 % of the bats we screened weighed less than 100 g. In combination with our ethical decision not to destructively sample the bats, this meant that individual blood volumes were frequently much less than 100 μl.","tracks":[{"project":"Zoonoses_partialAnnotation","denotations":[{"id":"T2","span":{"begin":2145,"end":2166},"obj":"Species"},{"id":"T4","span":{"begin":2171,"end":2189},"obj":"Species"}],"attributes":[{"id":"A2","pred":"tao:has_database_id","subj":"T2","obj":"Tax:58060"},{"id":"A4","pred":"tao:has_database_id","subj":"T4","obj":"Tax:478698"},{"subj":"T2","pred":"source","obj":"Zoonoses_partialAnnotation"},{"subj":"T4","pred":"source","obj":"Zoonoses_partialAnnotation"}]},{"project":"2_test","denotations":[{"id":"26184657-19590002-143716470","span":{"begin":418,"end":420},"obj":"19590002"}],"attributes":[{"subj":"26184657-19590002-143716470","pred":"source","obj":"2_test"}]}],"config":{"attribute types":[{"pred":"source","value type":"selection","values":[{"id":"Zoonoses_partialAnnotation","color":"#ec93a5","default":true},{"id":"2_test","color":"#93bfec"}]}]}}