Wastewater Analysis Indicates that Genetically Diverse Astroviruses, Including Strains Belonging to Novel Clades MLB and VA, Are Circulating within Japanese Populations.

Human astroviruses (HAstVs) are a common etiological agent of infantile gastroenteritis. Recent studies revealed that novel astrovirus (AstV) strains of the MLB clade (MLB-AstVs) and VA clade (VA-AstVs), which are genetically distinct from the classic HAstVs, are circulating in the human population. In the present study, we quantified classic HAstVs as well as carried out a genetic analysis of classic and novel HAstVs in wastewater in Japan. The concentration of classic HAstVs in the influent water samples ranged from 10(4) to 10(5) copies per liter, and the amount removed by wastewater treatment was determined to be 2.4 ± 0.3 log10. Four types of classic HAstV strains (HAstV types 1, 2, 5, and 4/8) as well as novel AstV strains belonging to the MLB-2, VA-1, and VA-2 clades were identified using reverse transcription-PCR (RT-PCR) assays, including assays newly developed for the detection of strains of the MLB and VA clades, followed by cloning and nucleotide sequencing. Our results suggest that genetically diverse AstV strains are circulating among the human population in Japan. The newly developed (semi)nested RT-PCR assays for these novel AstV clades are useful to identify and characterize the novel AstVs in environmental waters.