On-site reverse transcription-quantitative polymerase chain reaction detection of rotaviruses concentrated from environmental water samples using methacrylate monolithic supports.
Rotaviruses are the leading cause of gastroenteritis in children and they exist widely in water environments. Ingestion of 10-100 viral particles is enough to initiate disease, what calls for extremely sensitive detection methods. In this study we have confirmed the validity of a recently published method for rotavirus concentration and detection based on the combination of methacrylate monoliths and real-time reverse transcription-quantitative PCR (RT-qPCR). The method was used to concentrate rotaviruses from different tap water and environmental water samples collected in Slovenia within years 2007 and 2009. The performance of virus concentration using monolithic supports was improved in comparison to the one of tangential ultrafiltration upon application of both methods on a range of environmental samples. Several samples were successfully concentrated on-site after successful adaptation of the method to field requirements. In such on-site format, the combination of concentration using CIM and detection using RT-qPCR detected as low as 30 rotavirus particles/ml, spiked in an environmental water sample.
Department of Biotechnology and Systems Biology, National Institute of Biology, Ljubljana, Slovenia. ion.gutierrez@nib.si
Ion
I
Gutiérrez-Aguirre
Andrej
A
Steyer
Marko
M
Banjac
Petra
P
Kramberger
Mateja
M
Poljšak-Prijatelj
Maja
M
Ravnikar
D002853:Chromatography, Liquid / Q000379:methods; D008689:Methacrylates / Q000737:chemistry*; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D045483:Rivers / Q000821:virology; D012401:Rotavirus / Q000302:isolation & purification*; D012965:Sodium Chloride; D014773:Virology / Q000379:methods*; D014871:Water Microbiology; D014881:Water Supply / Q000032:analysis
D008689:Methacrylates; D012965:Sodium Chloride