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Although epidemics of hand, foot, and mouth disease (HFMD) caused by enterovirus A71 (EV-A71) have occurred worldwide, the Asia-Pacific region has seen large sporadic outbreaks with many severe neurological cases. This suggests that the virulence of the circulating viruses fluctuates in each epidemic and that HFMD outbreaks with many severe cases occur when highly virulent viruses are circulating predominantly, which has not been experimentally verified. Here, we analyzed 32 clinically isolated strains obtained in Japan from 2002 to 2013, along with 27 Vietnamese strains obtained from 2015 to 2016 that we characterized previously using human SCARB2 transgenic mice. Phylogenetic analysis of the P1 region classified them into five clades belonging to subgenogroup B5 (B5-I to B5-V) and five clades belonging to subgenogroup C4 (C4-I to C4-V) according to the epidemic year and region. Interestingly, clades B5-I and B5-II were very virulent, while clades B5-III, B5-IV, and B5-V were less virulent. Clades C4-II, C4-III, C4- IV, and C4-V were virulent, while clade C4-I was not. The result experimentally showed for the first time that several clades with different virulence levels emerged one after another. The experimental virulence evaluation of circulating viruses using SCARB2 transgenic mice is helpful to assess potential risks of circulating viruses. These results also suggest that a minor nucleotide or amino acid substitution in the EV-A71 genome during circulation causes fluctuations in virulence. The data presented here may increase our understanding of the dynamics of viral virulence during epidemics. IMPORTANCE Outbreaks of hand, foot, and mouth disease (HFMD) with severe enterovirus A71 (EV-A71) cases have occurred repeatedly, mainly in Asia. In severe cases, central nervous system complications can lead to death, making it an infectious disease of importance to public health. An unanswered question about this disease is why outbreaks of HFMD with many severe cases sometimes occur. Here, we collected EV-A71 strains that were prevalent in Japan and Vietnam over the past 20 years and evaluated their virulence in a mouse model of EV-A71 infection. This method clearly revealed that viruses belonging to different clades have different virulence, indicating that the method is powerful to assess the potential risks of the circulating viruses. The results also suggested that factors in the virus genome cause an outbreak with many severe cases and that further studies facilitate the prediction of large epidemics of EV-A71 in the future. [ABSTRACT FROM AUTHOR]

Respiratory syncytial virus (RSV) antigen-detection kits are used in clinical settings in Japan. Here, we compared the sensitivities of 12 different kits that are commercially available in Japan. Notably, these kits were developed based on the principle of immunochromatography. The comparison was performed using six different test specimens：standard laboratory strains of RSV subgroups A and B, and clinical specimens of nasal aspirates from patients infected with RSV subgroups A or B. The specimens were serially diluted with medium at half log concentration, and the dilutions were applied to the kit tests. The minimum detection limit was determined for each kit and for every specimen by investigating the maximum dilution rate for acquiring positive result. The absolute minimum detection limits were calculated based on the viral RNA copy number of the specimens and the maximum dilution rate ranged from 107 to 109 copies/mL, depending on the specimens and kits. Thus, the sensitivity was revealed to differ among the viral specimens examined using each kit and among the kits used to examine each viral specimen, though the difference was not significant and revealed a maximum of only 10 times. The results were compared with the geometric mean values of the results for all 12 kits which were separately calculated for all 6 specimens. Consequently, some kits with lower or higher sensitivities to all the tested specimens were identified, suggesting that some general differences in sensitivity exist among commercially available RSV antigen-detection kits, although the differences are not significant. Of course, selecting kit with a higher sensitivity would be preferable, but no large differences in sensitivity among the available kits examined in the current study. Therefore, other factors such as cost-effectiveness, convenience, time required for testing, etc., can be taken into account when selecting a kit for use in clinical settings.