Rotavirus is the leading viral agent for pediatric gastroenteritis. However, the case-based surveillance for rotavirus is limited in China, and its circulation in the environment is not well investigated. From 2013 to 2014, rotavirus was detected in raw sewage samples of Jinan and Linyi by quantitative PCR (qPCR) and conventional reverse transcription PCR (RT-PCR). After sequenced and genotyped, sequences analysis was conducted. A total of 46 sewage samples were collected monthly for the detection of rotavirus, and rotavirus was positive in 43 samples (93.5 %, 43/46). By quantitative assessment, the concentrations of rotavirus in raw sewage ranged from 4.1 × 103 to 1.3 × 106 genome copies (GC)/L in Jinan, and from 1.5 × 103 to 3.0 × 105 GC/L in Linyi. A total of 318 sequences of 5 G-genotypes and 318 sequences of 5 P-genotypes were obtained. G9 (91.8 %, 292/318) and P[8] (56.0 %, 178/318) were the most common G- and P-genotype, respectively. Multiple transmission lineages were recognized in these genotypes. Interestingly, an intragenic recombination event between two G9 lineages was observed. This study provided the first report of comprehensive environmental surveillance for rotavirus in China. The results suggest that the concentration of rotavirus in raw sewage was high, and multiple rotavirus transmission lineages continuously co-circulated in Shandong.

Measles virus (MeV) genotype B3 is one globally significant circulating genotype. Here, we present a systematic description of long-term evolutionary characterizations of the MeV genotype B3’s hemagglutinin (H) gene in the elimination era. Our results show that the B3 H gene can be divided into two main sub-genotypes, and the highest intra-genotypic diversity was observed in 2004. MeV genotype B3’s H gene diverged in 1976; its overall nucleotide substitution rate is estimated to be 5.697 × 10⁻⁴ substitutions/site/year, and is slowing down. The amino acid substitution rate of genotype B3’s H gene is also decreasing, and the mean effective population size has been in a downward trend since 2000. Selection pressure analysis only recognized a few sites under positive selection, and the number of positive selection sites is getting smaller. All of these observations may reveal that genotype B3’s H gene is not under strong selection pressure, and is becoming increasingly conservative. MeV H-gene or whole-genome sequencing should be routine, so as to better elucidate the molecular epidemiology of MeV in the future.

Rotavirus group A (RVA) G1 is one leading genotype circulating in humans worldwide, and related molecular information from a global perspective is still limited. Here, we present a comprehensive description of the genetic characterizations and molecular evolution of the RVA G1 VP7 gene. Our results show that RVA G1 can be divided into two lineages and multiple sub-lineages with a relatively high genetic diversity. Vaccine strains are phylogenetic, closer to lineage I. The evolutionary rate of the RVA G1 VP7 gene is 8.869 × 10⁻⁴ substitutions/site/year, and its most recent common ancestor was in 1933. The RVA G1 VP7 gene shows a linear evolution at the nucleotide level and a linear accumulation of difference at the amino acid level. Sub-lineage replacement of G1 VP7 gene is also observed and the effective population size of the G1 VP7 gene has had great change in the past decades and has remained stable in recent years. Altogether, the RVA G1 VP7 gene constantly evolves and there is no clear evidence that the evolution of the RVA G1 VP7 gene was influenced by vaccines. Continuous surveillance is still indispensable to evaluate the molecular epidemiology of RVA, especially in the post-vaccination era.

Classic human astroviruses (HAstV) are major global viral agents for gastroenteritis, but the molecular characteristics of classic HAstVs are not well understood. Here, we presented the molecular evolution of all classic HAstV serotypes by the analysis of the capsid protein sequences. Our results show that classic HAstVs can be divided into four groups with the most recent common ancestor (TMRCA) of 749. The overall evolutionary rate of classic HAstVs on the capsid gene was 4.509 × 10⁻⁴ substitutions/site/year, and most of the serotypes present a clock-like evolution with an amino acid accumulation of mutations over time. The mean effective population size of classic HAstVs is in a downward trend, and some positive and more than 500 negative selection sites were determined. Taken together, these results reveal that classic HAstVs evolve at the intra-serotype level with high genetic heterogeneity and are driven by strong purifying selection. Long-term surveillance of classic HAstVs are needed to enrich the genomic data for further analysis.

Norovirus is the leading viral agent of gastroenteritis in humans. RNA-dependent RNA polymerase (RdRp) is essential in the replication of norovirus RNA. Here, we present a comprehensive evolutionary analysis of the norovirus GI RdRp gene. Our results show that the norovirus GI RdRp gene can be divided into three groups, and that the most recent common ancestor was 1484. The overall evolutionary rate of GI RdRp is 1.821 × 10⁻³ substitutions/site/year. Most of the amino acids of the GI RdRp gene were under negative selection, and only a few positively selected sites were recognized. Amino acid substitutions in the GI RdRp gene accumulated slowly over time. GI.P1, GI.P3 and GI.P6 owned the higher evolutionary rates. GI.P11 and GI.P13 had the faster accumulation rate of amino acid substitutions. GI.P2, GI.P3, GI.P4, GI.P6 and GI.P13 presented a strong linear evolution. These results reveal that the norovirus GI RdRp gene evolves conservatively, and that the molecular evolutionary characteristics of each P-genotype are diverse. Sequencing in RdRp and VP1 of norovirus should be advocated in the surveillance system to explore the effect of RdRp on norovirus activity.

Echovirus 14 (E-14) causes various clinical recognized syndromes, mostly with gastrointestinal syndrome and paralysis. The current study summarized the Shandong E-14 strains isolated from a 26-year acute flaccid paralysis (AFP) surveillance, and elucidated the characterization of phylogenetic and phylogeographic relationships of E-14 worldwide. As a predominant serotype circulating in AFP surveillance, phylogenetic analysis showed that E-14 exhibited both time and geographic subdivision worldwide. In order to know the evolutionary history and spatial temporal dynamics of E-14, evolutionary phylogeography was reconstructed using BEAST and SPREAD software based on the VP1 sequences. The time of the most recent common ancestor of E-14 was estimated around 85 years and evolved with 9.17 × 10−3 substitutions/site/year. Phylogeographic analysis suggested that two regional transmissions of E-14 were mainly detected, with one located between Europe and Africa countries and the other was in the Asia-Pacific region. Our study investigates the molecular evolution and phylogeographic of E-14, and brings new insight to the dispersal of E-14 worldwide. Regional transmission was mainly detected and Australia may be responsible for the spread of E-14 in recent years.