Detection of enteroviruses, hepatitis A virus and rotaviruses in sewage by means of an immunomagnetic capture reverse transcription-PCR assay.
An immunomagnetic capture reverse transcription-PCR (IMC-RT-PCR) assay was evaluated to recover and detect enteric viruses in sewage and to remove PCR inhibitors. The procedure was applied along with a simple sample processing consisting of an initial separation of solids followed by polyethylen glycol precipitation and solvent extraction. This procedure reduced sample volumes by about 65-fold without eliminating RT-PCR inhibitors. Paramagnetic beads coupled to pooled human immunoglobulins were used to simultaneously capture poliovirus 1 (PV-1) and hepatitis A virus (HAV) from seeded sewage concentrates. The IMC was efficient in removing PCR inhibitors and in further reducing sample volumes by approximately 10-fold allowing the analysis of 6-7 ml of sewage sample per RT-PCR reaction. The detection limits of IMC-RT-PCR from seeded concentrates were 0.1-1 PFU for PV-1 and 1 MPNCU for HAV. The described procedure could be applied successfully for the detection of enteroviruses, HAV and rotaviruses in field sewage samples.
Casas
Nerea
N
oipsupae@vc.ehu.es
Suñén
Ester
E
D004770:Enterovirus / Q000235:genetics / Q000302:isolation & purification; D030041:Hepatitis A virus / Q000235:genetics / Q000302:isolation & purification; D018189:Immunomagnetic Separation / Q000379:methods; D017955:Poliovirus / Q000235:genetics / Q000302:isolation & purification; D012328:RNA Viruses / Q000235:genetics / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D012401:Rotavirus / Q000235:genetics / Q000302:isolation & purification; D012722:Sewage / Q000821:virology*
D012722:Sewage