Nine approaches to recover viral RNA from environmental silty sediments were newly developed and compared to quantify RNA viruses in sediments using molecular methods. Four of the nine approaches employed direct procedures for extracting RNA from sediments (direct methods), and the remaining five approaches used indirect methods wherein viral particles were recovered before RNA extraction. A direct method using an SDS buffer with EDTA to lyse viral capsids in sediments, phenol-chloroform-isoamyl alcohol to extract RNA, isopropanol to concentrate RNA, and magnetic beads to purify RNA resulted in the highest rate of recovery (geometric mean of 11%, with a geometric standard deviation of 0.02; n ؍ 7) of poliovirus 1 (PV1) inoculated in an environmental sediment sample. The direct method exhibiting the highest rate of PV1 recovery was applied to environmental sediment samples. One hundred eight sediment samples were collected from the Takagi River, Miyagi, Japan, and its estuary from November 2007 to April 2009, and the genomic RNAs of enterovirus and human norovirus in these samples were quantified by reverse transcription (RT)-quantitative PCR (qPCR). The human norovirus genome was detected in one sample collected at the bay, although its concentration was below the quantification limit. Meanwhile, the enterovirus genome was detected in two samples at the river mouth and river at concentrations of 8.6 ؋ 10 2 and 2.4 ؋ 10 2 copies/g (wet weight), respectively. This is the first report to obtain quantitative data for a human pathogenic virus in a river and in estuarine sediments using RT-qPCR. [6,14], and rotavirus [6]) pathogens have been detected in environmental sediments. Whittington et al. reported previously that M. avium in sediments from a dam lake survived 12 to 26 weeks longer than it did in a water column (29). Moreover, the persistence of viral pathogens in environmental sediments has been reported. Smith et al. demonstrated previously that poliovirus 1 (PV1), coxsackieviruses B3 and A9, and echovirus 1 survived significantly longer when associated with marine sediments (23). A 3-log reduction in the infectivity of PV1 was observed in 14 days in seawater having marine sediments, whereas such a reduction was observed in 4 days in seawater without sediments (23). These results suggest that environmental sediments have a protective effect on pathogens (1, 20), and the association of pathogens with environmental sediments cannot be ignored when considering the fate of pathogens in water environments (22).
Bacterial (Mycobacterium aviumWhen storms, tides, or strong winds cause sediment resuspension, pathogens in sediments are also resuspended, resulting in high pathogen levels in the water column. Dorner et al. indicated previously the importance of the resuspension of microorganisms from stream sediments rather than land-based sources according to the hydrological simulation of Escherichia coli during storm events (4). The quantitative detection of pathogens in environmental sediments is thus crucial for ...