Detection of rotaviruses in water by gene probes

Gene probes and other molecular techniques are a novel approach for the detection of enteric viruses in water. Gene probe hybridization to genomic viral RNA has been reported to detect 2.5 × 10⁵ physical particles of SA-11 rotavirus (Dimitrov et al., 1985), 10⁴ physical particles of hepatitis A virus (Jiang et al., 1986), 500-1000 PFU of SA-11 (De Leon, 1989), and 500-1000 PFU of coxsackie B3 (Shieh et al., 1990, submitted). When determinations of gene probre sensitivity are made in environmental concentrates a 100-fold decrease in signal may be observed (De Leon, 1989). Probe sensitivity in environmental samples can be enhanced with other molecular techniques. Sephadex G-200 spun columns (De Leon, 1989) or Centricon ultrafiltration membranes (Sobsey, 1990) can remove proteins and other interfering factors from water concentrates. The passage of environmental concentrates through Sephadex G-200 spun columns can result in a 10 to 1000-fold increase in detection of viruses (De Leon, 1989). Target nucleic acid amplification is the most feasible alternative for achieving the sensitivity limits needed for environmental samples. Cell culture (Shieh et al., 1990), rotavirus incorporated transcriptase (De Leon, 1989) and Polymerase Chain Reaction (Shieh et al., 1990; Sobsey and De Leon, 1990) have been used to amplify viral sequences in environmental samples. In this study, the rotavirus incorporated transcriptase was used to amplify rotaviral mRNA in environmental samples prior to detection by gene probes.