Seasonal shift in epidemics of respiratory syncytial virus infection in Japan.

In Japan, respiratory syncytial virus (RSV) infection generally has occurred during autumn and winter. However, a possible change in the seasonal trend of RSV infection has been observed recently. The current study was conducted to determine whether the epidemic season of RSV infection in Japan has indeed changed significantly. We used expectation-based Poisson scan statistics to detect periods with high weekly reported RSV cases (epidemic cluster), and the epidemic clusters were detected between September and December in the 2012-2016 seasons while those were detected between July and October in the 2017-2019 seasons. Non-linear and linear ordinary least squares regression models were built to evaluate whether there is a difference in year trend in the epidemic seasonality, and the epidemic season was shifted to earlier in the year in 2017-2019 compared to that in 2012-2016. Although the reason for the shift is unclear, this information may help in clinical practice and public health.

Polymerase chain reaction primer sets for the detection of genetically diverse human sapoviruses.

Sapoviruses are increasingly being recognized as pathogens associated with gastroenteritis in humans. Human sapoviruses are currently assigned to 18 genotypes (GI.1-7, GII.1-8, GIV.1, and GV.1-2) based on the sequence of the region encoding the major structural protein. In this study, we evaluated 11 polymerase chain reaction (PCR) assays using published and newly designed/modified primers and showed that four PCR assays with different primer combinations amplified all of the tested human sapovirus genotypes using either synthetic DNA or cDNA prepared from human sapovirus-positive fecal specimens. These assays can be used as improved broadly reactive screening tests or as tools for molecular characterization of human sapoviruses.

GII.17 norovirus infections in outbreaks of acute nonbacterial gastroenteritis in Osaka City, Japan during two decades.

Norovirus (NoV) is a major cause of viral gastroenteritis, and GII.4 has been the predominant genotype worldwide since the mid-1990s. During the 2014 to 2015 winter, a rare genotype, NoV GII.17, emerged and became prevalent mainly in East Asia. Over the past two decades, NoV molecular surveillance in Osaka City, Japan, has revealed that NoV GII.17 was detected for the first time in February 2001 and that NoV GII.17-associated outbreaks remarkably increased during the 2014 to 2015 season, with higher incidence recorded in January to March 2015. Genetic analysis indicated that 28 GII.17 outbreak strains were closely related to the novel GII.P17-GII.17 variants represented by the Kawasaki308/2015/JP strain, similar to that in other regions. Statistical analysis showed that NoV GII.17 infections were more common in adults than GII.3 and GII.4 infections, suggesting that the affected adults most likely did not have antibodies against NoV GII.17 and the novel GII.17 variant had recently appeared. Regarding transmission, food was one of the most important factors involved in the spread of NoV GII.17 among adults; 61% of GII.17 outbreaks were foodborne, with oysters being the most common vehicle. Interplay between pathogens, hosts, and environmental factors was considered to be important in the 2014 to 2015 NoV GII.17 epidemic.

Broadly reactive real-time reverse transcription-polymerase chain reaction assay for the detection of human sapovirus genotypes.

Sapoviruses are associated with acute gastroenteritis. Human sapoviruses are classified into four distinct genogroups (GI, GII, GIV, and GV) based on their capsid gene sequences. A TaqMan probe-based real-time reverse transcription-polymerase chain reaction (RT-PCR) assay that detects the representative strains of these four genogroups is widely used for screening fecal specimens, shellfish, and environmental water samples. However, since the development of this test, more genetically diverse sapovirus strains have been reported, which are not detectable by the previously established assays. In this study, we report the development of a broader-range sapovirus real-time RT-PCR assay. The assay can detect 2.5 × 107 and 2.5 × 10 1 copies of sapovirus and therefore is as sensitive as the previous test. Analysis using clinical stool specimens or synthetic DNA revealed that the new system detected strains representative of all the 18 human sapovirus genotypes: GI.1-7, GII.1-8, GIV.1, and GV.1, 2. No cross-reactivity was observed against other representative common enteric viruses (norovirus, rotavirus, astrovirus, and adenovirus). This new assay will be useful as an improved, broadly reactive, and specific screening tool for human sapoviruses.

Impact of Coxsackievirus A6 emergence on hand, foot, and mouth disease epidemic in Osaka City, Japan.

Hand, foot, and mouth disease (HFMD) is an acute febrile illness characterized by fever; sore throat; and vesicular eruptions on the hands, feet, and oral mucosa. Until 2010, HFMD was predominantly associated with enterovirus (EV) A71 and coxsackievirus (CV) A16 in Japan. In 2011, CV-A6 emerged as a primary causative agent, causing the largest HFMD epidemic in Japan since 1981. Since then, CV-A6 has caused large HFMD epidemics every 2 years. The phylogenetic analysis of complete Viral Protein 1 (VP1) sequences revealed that most CV-A6 strains detected from 2011 to 2015 in Osaka City were classified into a different clade compared with CV-A6 strains detected from 1999 until 2009. The majority of CV-A6 strains detected in 2011 and most CV-A6 strains detected from 2013 to 2015 were mainly divided into two distinct genetic groups. Each epidemic strain carried unique amino acid substitutions in the presumed DE, EF, and GH loops of the VP1 protein that is exposed on the surface of the virion. There is a possibility that the appearance of substitutions on the surface of the virion and an accumulation of a susceptible population are significant factors in recent HFMD epidemics.

Epidemics of GI.2 sapovirus in gastroenteritis outbreaks during 2012-2013 in Osaka City, Japan.

Sapovirus (SaV) is a causative agent of gastroenteritis in humans in both sporadic cases and outbreaks. During the period from January 2005 to August 2014, SaV was detected in 30 (5.9%) of 510 gastroenteritis outbreaks in Osaka City, Japan using real-time RT-PCR. Seasonal distribution of SaV-associated outbreaks revealed an increase during the 2011-2012 season and the highest frequency of outbreaks during the 2012-2013 season. Genotyping analysis based on the capsid region demonstrated that the most common genotype was GI.2 (36.7%), in which the strains were closely related. The comparison of complete capsid gene sequences with 18 GI.2 strains (7 strains in this study and 11 from GenBank) between 1990 and 2013 showed that GI.2 strains were classified into at least three genetic clusters (1990-2000, 2004-2007, and 2008-2013) with chronologically unique amino acid residues and accumulation of mutations in the predicted P domain, suggesting the one of the causes of emergence and spread of GI.2 strains. This study will also be helpful for understanding the evolutionary mechanism of the SaV genome.

Detection and characterization of a human G9P[4] rotavirus strain in Japan.

In a surveillance system in Osaka City, Japan, 48 sporadic rotavirus A (RVA) infections were detected during 2008/2009-2011/2012 seasons. The G/P-genotypes of detected RVAs were G1P[8], G2P[4], G3P[8], G9P[4], and G9P[8]. Although G9P[4] is a rare genotype that had not been reported in Japan, it was the second most prevalent genotype, following G1P[8], and accounted for 35.3% of RVA cases in the 2011/2012 season. Further genotyping revealed that the G9P[4] strain had genotype 2 internal protein genes except for NSP3: G9-P[4]-I2-R2-C2-M2-A2-N2-T1-E2-H2. Among detected RVA strains, G9P[4] and some G9P[8] strains shared high nucleotide identity in VP7 and NSP3 genes. Phylogenetic and BLAST search analyses showed that the G9P[4] strain in Japan shared high nucleotide identity in genotype 2 genes with common G2P[4] strains circulating globally, but was distinct from other G9P[4] strains circulating worldwide. These results suggest that the G9P[4] strain in Japan may have emerged through an independent reassortment between G9P[8] and G2P[4]. Finally, the role of NSP3 protein in the circulating RVA from an amino acid comparison between T1- and T2-type NSP3 is discussed. These findings provide an important insight into less problematic combinations of circulating RVA genes derived from different genotypes.

Relationship between Severity of Aseptic Meningitis and Cerebrospinal Fluid Cytokine Levels.

BACKGROUND: Pediatricians sometimes see patients with severe aseptic meningitis and prolonged fever or severe headache, or both. This condition generally has a good prognosis and is usually treated with supportive therapy. However, there is neither guideline nor consensus for the treatment of patients with severe aseptic meningitis. Here, we investigated the relationship between disease severity and biomarkers. METHODS: The subjects were 32 children aged 0 to 14 years, 23 of whom had aseptic meningitis and 9 of whom were meningitis-free controls. Aseptic meningitis was retrospectively categorized into two subgroups, namely mumps meningitis (MM) and viral meningitis excluding that caused by mumps (EM). We defined a novel aseptic meningitis severity score (AMSS) from the signs and symptoms of aseptic meningitis and thus evaluated disease severity. We analyzed the profiles of cytokines in the patients' cerebrospinal fluid (CSF). RESULTS: The AMSS in MM was significantly higher than that in EM. IL-4, IL-6, IL-8, IL-10, and G-CSF levels in MM and EM CSF were higher than those in control CSF. IFN-γ levels were higher in MM than in controls (p<0.01). IL-10 and IFN-γ levels in MM were higher than those in EM. CONCLUSIONS: MM was more severe than EM. One likely reason is the higher CSF cytokine levels in MM. IFN-γ may be a potentially strong biomarker of MM severity. Our findings would help further understanding

Gastroenteritis outbreaks caused by a DS-1-like G1P[8] rotavirus strain, Japan, 2012-2013.

Rotavirus A (RVA) genotype G1P[8], a hallmark of the Wa-like strain, typically contains only genotype 1 genes. However, an unusual RVA G1P[8] with genotype 2 genes was recently detected in Japan. We determined the complete genomic constellation of this RVA. Our findings suggest that mixed RVAs may be more competitive than once thought.

Detection and genetic characterization of human enteric viruses in oyster-associated gastroenteritis outbreaks between 2001 and 2012 in Osaka City, Japan.

Enteric viruses are an important cause of viral food-borne disease. Shellfish, especially oysters, are well recognized as a source of food-borne diseases, and oyster-associated gastroenteritis outbreaks have on occasion become international occurrences. In this study, 286 fecal specimens from 88 oyster-associated gastroenteritis outbreaks were examined for the presence of 10 human enteric viruses using antigenic or genetic detection methods in order to determine the prevalence of these infections. All virus-positive patients were over 18 years old. The most common enteric virus in outbreaks (96.6%) and fecal specimens (68.9%) was norovirus (NoV), indicating a high prevalence of NoV infection associated with the consumption of raw or under-cooked oysters. Five other enteric viruses, aichiviruses, astroviruses, sapoviruses, enteroviruses (EVs), and rotavirus A, were detected in 30.7% of outbreaks. EV strains were characterized into three rare genotypes, coxsackievirus (CV) A1, A19, and EV76. No reports of CVA19 or EV76 have been made since 1981 in the Infectious Agents Surveillance Report by the National Infectious Diseases Surveillance Center, Japan. Their detection suggested that rare types of EVs are circulating in human populations inconspicuously and one of their transmission modes could be the consumption of contaminated oysters. Rapid identification of pathogens is important for the development of means for control and prevention. The results of the present study will be useful to establish an efficient approach for the identification of viral pathogens in oyster-associated gastroenteritis in adults.

Exploring the threshold for the start of respiratory syncytial virus infection epidemic season using sentinel surveillance data in Japan.

Introduction: An unusual seasonality of respiratory syncytial virus (RSV) infection in Japan is observed in recent years after 2017, becoming challenging to prepare for: a seasonal shift from autumn-winter to summer-autumn in 2017-2019, no major epidemic in 2020, and an unusually high number of cases reported in 2021. Methods: To early detect the start-timing of epidemic season, we explored the reference threshold for the start-timing of the epidemic period based on the number of cases per sentinel (CPS, a widely used indicator in Japanese surveillance system), using a relative operating characteristic curve analysis (with the epidemic period defined by effective reproduction number). Results: The reference values of Tokyo, Kanagawa, Osaka, and Aichi Prefectures were 0.41, 0.39, 0.42, and 0.24, respectively. Discussion: The reference CPS value could be a valuable indicator for detecting the RSV epidemic and may contribute to the planned introduction of monoclonal antibody against RSV to prevent severe outcomes.

Increase of GII.2 norovirus infections during the 2009-2010 season in Osaka City, Japan.

During the 2009-2010 season, a significant numerical increase of genotype GII.2 norovirus (NoV)-associated outbreaks was observed in Osaka City, Japan. The most common genotype in that season was GII.2 (44.6%), followed by GII.4 (39.2%). Mostly, GII.2 strains were associated with outbreaks in children and with person-to-person contact. The National Infectious Disease Surveillance Center reported that GII.2 NoV infections were widespread in Japan in that season. Comparative phylogenetic analysis of RNA-dependent RNA polymerase (RdRp) and capsid sequences revealed that this GII.2 epidemic resulted from two genetic strains. The first, GII.2p2 strains, had an identical genotype in the RdRp and capsid genes. GII.2p2 strains in the 2009-2010 season were a different genetic cluster from the strains of spring 2004, the previous epidemic of GII.2 NoV, but showed no unique amino acid change. The second, GII.2 chimera virus (GII.2p16), had GII.16 RdRp and GII.2 capsid genotypes, suggesting prior recombination at the junction of ORF1 and ORF2. GII.2p16 strains had four significant amino acid changes in the P2 subdomain, suggesting antigenic changes. Before the 2009-2010 season, GII.2 chimera viruses had been observed only sporadically. This spreading of GII.2p16 strains in the 2009-2010 season might be the first epidemic of GII.2 chimera virus. This study revealed that the NoV epidemic in the 2009-2010 season differed considerably from the prior season, when GII.4 was predominant. Furthermore, GII.2 strains persisted in human populations by drastic recombination and gradual accumulation of mutations, indicating a prevalent pattern of non-GII.4 genotypes with genetic evolution.

Human sapovirus classification based on complete capsid nucleotide sequences.

The genetically diverse sapoviruses (SaVs) are a significant cause of acute human gastroenteritis. Human SaV surveillance is becoming more critical, and a better understanding of the diversity and distribution of the viral genotypes is needed. In this study, we analyzed 106 complete human SaV capsid nucleotide sequences to provide a better understanding of their diversity. Based on those results, we propose a novel standardized classification scheme that meets the requirements of the International Calicivirus Scientific Committee. We believe the classification scheme and strains described here will be of value for the molecular characterization and classification of newly detected SaV genotypes and for comparing data worldwide.

Title: Factors associated with time lag between symptom onset and reporting in the first epidemic wave of COVID-19 in Osaka, Japan.

Objectives: Longer reporting lags after symptom onset reportedly exert a substantial impact on onward transmission, increasing outbreak probability. Our study investigated the risk factors associated with reporting lag. Methods: Using active epidemiological surveillance data for all symptomatic cases reported in Osaka Prefecture during the first wave of the coronavirus disease 2019 (COVID-19) epidemic (February 1-May 13, 2020), multivariable regression analyses were implemented to estimate the effects of exposure variables on reporting lag, by controlling for potential confounders. Results: Cases in their 30s showed a longer reporting lag than cases ≥ 80 years old. Cases who lived in areas with a high COVID-19 incidence demonstrated a longer reporting lag. Cases with a history of visiting a nightlife district also showed longer reporting lag than cases without such a history. Healthcare workers and cases with immunodeficiency both displayed shorter reporting lags than others. Conclusion: Identifying newly infected cases as soon as possible and increased testing capacity for all age groups, and for individuals with a history of visiting high infection-risk areas, represented important measures in shortening reporting lags in the first wave period. The evidence from this study may provide lessons for controlling future emerging diseases.

Enterovirus 68 in children with acute respiratory tract infections, Osaka, Japan.

Enterovirus 68 strains were detected in 14 specimens from children with respiratory tract infections and 1 specimen from a child with febrile convulsions during 2010 in Osaka, Japan. These strains had deletions in the 5' untranslated region and were genetically different from reported strains. This virus is associated with respiratory tract infections in Japan.

Molecular epidemiology of noroviruses detected in seasonal outbreaks of acute nonbacterial gastroenteritis in Osaka City, Japan, from 1996-1997 to 2008-2009.

In seasons from 1996-1997 through 2008-2009, noroviruses (NoVs) were detected in 505 outbreaks (71%) of nonbacterial gastroenteritis in Osaka City, Japan using molecular diagnosis with reverse transcription (RT)-PCR or real-time RT-PCR. The occurrences of NoV-associated outbreaks were related with the cold season during November-March (85.3%), and occasionally small epidemics of NoVs occurring during April-June were observed. Oyster-associated outbreaks were dominant transmission modes (25-61.1%) before the 2003-2004 season, and decreased (5-20.5%) from the 2003-2004 season, although outbreaks attributable to food-borne transmission (except for oysters) and person-to-person contact increased from the 2003-2004 season. The NoV strains were characterized into genotypes based on sequence analysis of partial capsid regions. Genotyping analyses identified at least 30 genotypes (12 in genogroup I [GI] and 18 in genogroup II [GII]) of NoV. The most common genotype was GII.4 (44.6%), followed in order by GII.3, GII.6, GII.2, and GII.5. The number of GII.4 NoVs increased greatly from the 2003-2004 season, eventually comprising a large share among the NoV- associated outbreaks (97.4%) of the 2006-2007 season. Occasional increased prevalence of genotypes other than GII.4 was observed during this study period. This study showed the appearance, spread, and disappearance of various genotypes and the change of NoV epidemic in a limited geographic region. Continuous NoV molecular surveillance is important for understanding NoV infections and for improving measures for their control and prevention.

Novel human reovirus isolated from children and its long-term circulation with reassortments.

Mammalian orthoreovirus (MRV), also known as reovirus, was discovered in the 1950s and became the first reported segmented double-stranded RNA virus. MRVs have since been found in a variety of animal species, including humans. However, reports on MRV infections are scarce due to the rarity of their symptomatic occurrence. In Japanese surveillance studies, MRVs have been detected as gastrointestinal pathogens since 1981, with a total of 135 records. In Osaka City, Japan, MRV was first isolated in 1994 from a child with meningitis, and then in 2005 and 2014 from children with gastroenteritis. Here, we conducted the first molecular characterization of human MRV isolates from Japan and identified a novel human reovirus strain belonging to MRV type 2, designated the MRV-2 Osaka strain. This strain, with all three isolates classified, is closely related to MRV-2 isolates from sewage in Taiwan and is relatively close to an MRV-2 isolate from a bat in China. Our data suggest that the MRV-2 Osaka strain, which has circulated amongst humans in Japan for at least two decades, has spread internationally.

Genetic analysis of the capsid gene of genotype GII.2 noroviruses.

Noroviruses (NoVs) are considered to be a major cause of acute nonbacterial gastroenteritis in humans. The NoV genus is genetically diverse, and genotype GII.4 has been most commonly identified worldwide in recent years. In this study we analyzed the complete capsid gene of NoV strains belonging to the less prevalent genotype GII.2. We compared a total of 36 complete capsid sequences of GII.2 sequences obtained from the GenBank (n = 5) and from outbreaks or sporadic cases that occurred in The Netherlands (n = 10) and in Osaka City, Japan (n = 21), between 1976 and 2005. Alignment of all capsid sequences did not show fixation of amino acid substitutions over time as an indication for genetic drift. In contrast, when strains previously recognized as recombinants were excluded from the alignment, genetic drift was observed. Substitutions were found at five informative sites (two in the P1 subdomain and three in the P2 subdomain), segregating strains into five genetic groups (1994 to 1997, 1999 to 2000, 2001 to 2003, 2004, and 2005). Only one amino acid position changed consistently between each group (position 345). Homology modeling of the GII.2 capsid protein showed that the five amino acids were located on the surface of the capsid and close to each other at the interface of two monomers. The data suggest that these changes were induced by selective pressure, driving virus evolution. Remarkably, this was observed only for nonrecombinant genomes, suggesting differences in behavior with recombinant strains.

An Epidemic of Hand, Foot, and Mouth Disease Caused by Coxsackievirus A6 in Osaka City, Japan, in 2017.

The second largest epidemic of hand, foot, and mouth disease since 1982 occurred in 2017, which involved 6,173 cases in Osaka City, Japan. The main causative agent was coxsackievirus A6 (CV-A6). Phylogenetic analysis revealed that the detected CV-A6 strains belonged to genetic groups A3 and A4 in clade A.

Dengue Virus in Traveler Returning to Japan from the Democratic Republic of the Congo, 2015.

Dengue fever (DF) is a mosquito-borne disease and a significant global public health problem. Although a few serological surveys in the literature suggest endemic DF in many parts of Africa, DF cases in these countries are generally underreported because of the lack of diagnostic testing and systematic surveillance; thus, little is known about the phylogenetic profile of circulating strains. In April 2015, DF was diagnosed in a Japanese national returning from the Democratic Republic of the Congo (DRC). Dengue virus 1 (DENV-1) RNA was detected in the patient's serum sample using real-time reverse transcription PCR. Phylogenetic analysis of the E gene revealed that the detected DENV-1 strain was classified as genotype V and was closely related, with 100% nucleotide identity, to the strain causing the 2013 DF epidemic in Angola, which is located directly south of the DRC. This is the first report to characterize the circulating DENV strain in the DRC, and the findings indicate that the DENV-1 strain causing the 2013 DF epidemic in Angola was also circulating in the DRC in 2015.