Abstract:
:The role of environmental reservoirs in avian influenza virus (AIV) transmission has been investigated during AIV-associated outbreaks. To date, no method has been defined for detection of AIV from mud samples. A procedure using elution and polyethylene glycol (PEG) concentration steps was designed to detect AIV by RT-PCR from 42g of raw mud, corresponding to 30g of the solid fraction of mud. RNA was recovered with MagMAX AI/ND Viral RNA Isolation kit (Ambion, Austin, TX). Three elution buffers were studied and viral recoveries higher than 29% were yielded by elution with a 10% beef extract solution (pH 7). The overall method showed that, under some conditions, virus was not detectable in PEG samples, whereas viruses were detected in the elution fractions. PCR curves were improved significantly by running the amplification reaction with a mixture containing a PCR additive for inhibitor removal, such as T4 gene 32 protein (Gp32), although PCR inhibitors from mud were removed partially from PEG samples. A theoretical detection threshold of 5×10(5) RNA copies of H5N1 virus per 30g of solid mud could be obtained by elution. The overall method has proved successful for detecting H5N1 virus contamination of mud specimens collected during outbreak investigations of avian influenza in Cambodia.
journal_name
J Virol Methodsjournal_title
Journal of virological methodsauthors
Deboosere N,Horm SV,Delobel A,Gachet J,Buchy P,Vialette Mdoi
10.1016/j.jviromet.2011.10.013subject
Has Abstractpub_date
2012-01-01 00:00:00pages
148-53issue
1eissn
0166-0934issn
1879-0984pii
S0166-0934(11)00429-0journal_volume
179pub_type
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