Increase in the number of new cases of ANCA-associated vasculitis in the COVID-19 vaccine era.

Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) is an autoimmune vasculitis characterized by the production of antibodies against ANCA, with unclear pathogenesis. With the ongoing COVID-19 pandemic, COVID-19 mRNA vaccination has been available in Japan since February 2021. Although autoimmune symptoms have been reported after COVID-19 vaccinations, there have been no clinical investigations regarding the relationship between COVID-19 mRNA vaccines and the pathogenesis of AAV. Thus, the present study aimed to investigate whether the administration of COVID-19 mRNA vaccines affects the development of AAV. The study identified patients with new-onset AAV who were MPO-ANCA or PR3-ANCA positive and met the entry criteria of the AAV EMA classification algorithm. The study compared the number of new AAV cases per year before and after the start of the COVID-19 mRNA vaccine program in Japan. The study found that the annual number of new cases of AAV in Japan's Nagasaki Prefecture increased by approximately 1.5-fold since the COVID-19 vaccine program was initiated, suggesting a possible link between the COVID-19 mRNA vaccines and the development of AAV. Although the study provides insight into the clinical evaluation and management of autoimmune symptoms following COVID-19 vaccination, further investigation of the possible association between COVID-19 mRNA vaccines and the pathogenesis of AAV is required.

Influenza A Virus Agnostic Receptor Tropism Revealed Using a Novel Biological System with Terminal Sialic Acid Knockout Cells.

Avian or human influenza A viruses bind preferentially to avian- or human-type sialic acid receptors, respectively, indicating that receptor tropism is an important factor for determining the viral host range. However, there are currently no reliable methods for analyzing receptor tropism biologically under physiological conditions. In this study, we established a novel system using MDCK cells with avian- or human-type sialic acid receptors and with both sialic acid receptors knocked out (KO). When we examined the replication of human and avian influenza viruses in these KO cells, we observed unique viral receptor tropism that could not be detected using a conventional solid-phase sialylglycan binding assay, which directly assesses physical binding between the virus and sialic acids. Furthermore, we serially passaged an engineered avian-derived H4N5 influenza virus, whose PB2 gene was deleted, in avian-type receptor KO cells stably expressing PB2 to select a mutant with enhanced replication in KO cells; however, its binding to human-type sialylglycan was undetectable using the solid-phase binding assay. These data indicate that a panel of sialic acid receptor KO cells could be a useful tool for determining the biological receptor tropism of influenza A viruses. Moreover, the PB2KO virus experimental system could help to safely and efficiently identify the mutations required for avian influenza viruses to adapt to human cells that could trigger a new influenza pandemic. IMPORTANCE The acquisition of mutations that allow avian influenza A virus hemagglutinins to recognize human-type receptors is mandatory for the transmission of avian viruses to humans, which could lead to a pandemic. In this study, we established a novel system using a set of genetically engineered MDCK cells with knocked out sialic acid receptors to biologically evaluate the receptor tropism for influenza A viruses. Using this system, we observed unique receptor tropism in several virus strains that was undetectable using conventional solid-phase binding assays that measure physical binding between the virus and artificially synthesized sialylglycans. This study contributes to elucidation of the relationship between the physical binding of virus and receptor and viral infectivity. Furthermore, the system using sialic acid knockout cells could provide a useful tool to explore the sialic acid-independent entry mechanism. In addition, our system could be safely used to identify mutations that could acquire human-type receptor tropism.

Cartography of SARS-CoV-2 variants based on the susceptibility to therapeutic monoclonal antibodies.

A comprehensive picture of a phenotypic relationship among severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants has been poorly studied. Here, this study presents cartography showing how the wild-type strain of SARS-CoV-2 and 14 variants are alike or different from the perspective of the susceptibility to 12 therapeutic monoclonal antibodies. The Alpha variant is close to the wild-type strain, whereas the Beta, Gamma, and Delta variants diverge from the wild-type. The map highlights the very unique property of the Omicron variant. Interestingly, sublineages of the Omicron variants, BA.1, BA.2, and BA.4/5, differ substantially in the cartography.

Age-Stratified Seroprevalence of SARS-CoV-2 Antibodies before and during the Vaccination Era, Japan, February 2020-March 2022.

Japan has reported a relatively small number of COVID-19 cases. Because not all infected persons receive diagnostic tests for COVID-19, the reported number must be lower than the actual number of infections. We assessed SARS-CoV-2 seroprevalence by analyzing >60,000 samples collected in Japan (Tokyo Metropolitan Area and Hokkaido Prefecture) during February 2020-March 2022. The results showed that ≈3.8% of the population had become seropositive by January 2021. The seroprevalence increased with the administration of vaccinations; however, among the elderly, seroprevalence was not as high as the vaccination rate. Among children, who were not eligible for vaccination, infection was spread during the epidemic waves caused by the SARS-CoV-2 Delta and Omicron variants. Nevertheless, seroprevalence for unvaccinated children <5 years of age was as low as 10% as of March 2022. Our study underscores the low incidence of SARS-CoV-2 infection in Japan and the effects of vaccination on immunity at the population level.

Analysis of research intensity on infectious disease by disease burden reveals which infectious diseases are neglected by researchers.

Infectious diseases are associated with considerable morbidity and mortality worldwide. Although human, financial, substantial, and time resources are limited, it is unknown whether such resources are used effectively in research to manage diseases. The correlation between the disability-adjusted life years to represent disease burden and number of publications as a surrogate for research activity was investigated to measure burden-adjusted research intensity for 52 infectious diseases at global and country levels. There was significantly low research intensity for paratyphoid fever and high intensity for influenza, HIV/acquired immunodeficiency syndrome, hepatitis C, and tuberculosis considering their disease burden. We identified the infectious diseases that have received the most attention from researchers and those that have been relatively disregarded. Interestingly, not all so-called neglected tropical diseases were subject to low burden-adjusted research intensity. Analysis of the intensity of infectious disease research at a country level revealed characteristic patterns. These findings provided a basis for further discussion of the more appropriate allocation of resources for research into infectious diseases.

[Comprehensive understanding of viral diseases by field, molecular, and theoretical studies].

Viral diseases are responsible for substantial morbidity and mortality and continue to be of great concern. To ensure better control of viral infections, I have been tackling the issue as a medical doctor, an academic researcher, and a public health officer. Especially, I have studied respiratory viruses, such as the influenza virus, from the perspectives of molecular virology, theoretical modeling, and field epidemiology. RNA biology and its involvement with viral life-cycle and pathogenicity are central topics of molecular study, while mathematical models of transmission dynamics and phylogenetics are major components of theoretical research. As a field epidemiologist, I work with public health authorities during viral disease outbreaks. I was deployed to West Africa for viral hemorrhagic fever outbreak responses as a WHO consultant, and I have served the Japanese Government as an advisor for COVID-19 countermeasures. I would like to integrate various approaches from clinical medicine to epidemiology, theoretical modeling, evolutionary biology, genetics, and molecular biology in my research. In that way, we could gain a more comprehensive understanding of viral diseases. I hope these findings will help ease the disease burden of viral infections around the world.

Familial Clusters of Coronavirus Disease in 10 Prefectures, Japan, February-May 2020.

The overall coronavirus disease secondary attack rate (SAR) in family members was 19.0% in 10 prefectures of Japan during February 22-May 31, 2020. The SAR was lower for primary cases diagnosed early, within 2 days after symptom onset. The SAR of asymptomatic primary cases was 11.8%.

Simulation study reveals factors that affect the predominance of SARS-CoV-2 novel variant.

The novel variants of the SARS-CoV-2 are a great global concern for the ongoing COVID-19 pandemic. However, how the novel variants predominate and replace existing strains remains elusive. In this study, I simulated the infection spread to investigate what kinds of viral, immunological, and epidemiological factors affect the predominance of SARS-CoV-2 novel variants. The results showed that the increase of the transmissibility of the novel variant substantially enhanced the predominance probability. In addition, the increasing trend of the infection spread, the large case number of the epidemic, and the ability of immune escape of the novel variant increased the predominance probability. A small number of cases and a decreasing trend of an entire epidemic, including not only the novel variant but also earlier strains, are especially important to reduce the chance of the predominance of the novel variant and delay the process. Good control of the COVID-19 epidemic could make the disease burden small and sequester the spread of the SARS-CoV-2 novel variants.

RNA Modifications in Genomic RNA of Influenza A Virus and the Relationship between RNA Modifications and Viral Infection.

Recent studies about the transcriptome-wide presence of RNA modifications have revealed their importance in many cellular functions. Nevertheless, information about RNA modifications in viral RNA is scarce, especially for negative-strand RNA viruses. Here we provide a catalog of RNA modifications including m1A, ac4C, m7G, inosine, and pseudouridine on RNA derived from an influenza A virus infected into A549 cells, as studied by RNA immunoprecipitation followed by deep-sequencing. Possible regions with RNA modifications were found in the negative-strand segments of viral genomic RNA. In addition, our analyses of previously published data revealed that the expression levels of the host factors for RNA modifications were affected by an infection with influenza A virus, and some of the host factors likely have a proviral effect. RNA modification is a novel aspect of host-virus interactions leading to the discovery of previously unrecognized viral pathogenicity mechanisms and has the potential to aid the development of novel antivirals.

[Epidemiology of Viral Hemorrhagic Fever in Africa].

A variety of viral hemorrhagic fevers such as Ebola virus disease exist in Africa and impose a great threat in public health due to their high fatality. It is considered to be difficult to eradicate the etiological agents of viral hemorrhagic fever because they have non-human natural hosts. Therefore, the importance of public health measures remains high in addition to the urgent need for the development of medicines for treatment and prevention. Furthermore, public health measures directly lead to the accumulation of epidemiological knowledge about the diseases. As an infectious disease consultant for the World Health Organization, I have been involved with public health activities including the development of clinical guidelines, the establishment of laboratory diagnostic systems, the training for infection, prevention and control, the planning of budget for outbreak response, and the analysis of epidemiological data. On the last point, I reported the situation of Ebola virus disease outbreak in Liberia, 2014-2015 and Lassa fever outbreak in Nigeria, 2018-2019 describing the risk factors, morbidity, and mortality of the diseases.

Clusters of Coronavirus Disease in Communities, Japan, January-April 2020.

We analyzed 3,184 cases of coronavirus disease in Japan and identified 61 case-clusters in healthcare and other care facilities, restaurants and bars, workplaces, and music events. We also identified 22 probable primary case-patients for the clusters; most were 20-39 years of age and presymptomatic or asymptomatic at virus transmission.

Association Between Preceding Viral Respiratory Infection and Subsequent Respiratory Illnesses Among Children: A Prospective Cohort Study in the Philippines.

Background: Acute respiratory infection (ARI) is of great concern in public health. It remains unclear whether viral infections can affect the host's susceptibility to subsequent ARIs. Methods: A prospective cohort study on ARIs of children below 5 years old was conducted in the Philippines from 2014 to 2016. The respiratory symptoms were recorded daily, and nasopharyngeal swabs were collected at both household and health facilities. The specimens were tested for respiratory viruses. We then determined whether viral etiology was associated with the severity of the present ARI and whether previous viral infections was associated with subsequent ARIs. Results: A total of 3851 children and 16337 ARI episodes were enrolled and recorded, respectively. Samples were collected from 24% of all ARI episodes; collection rate at the healthcare facilities was 95%. Enterovirus D68, rhinovirus species C, and respiratory syncytial virus were significantly associated with severe ARIs. The risk for subsequent ARIs was significantly enhanced after infections with adenovirus, influenza A virus, parainfluenza virus type 4, and rhinovirus species C. Conclusions: This study revealed that viral etiology plays a significant role in the severity of the present ARI and that viral infection affects the host's susceptibility to subsequent ARIs.

Epidemiologic and Clinical Features of Lassa Fever Outbreak in Nigeria, January 1-May 6, 2018.

Lassa fever (LF) is endemic to Nigeria, where the disease causes substantial rates of illness and death. In this article, we report an analysis of the epidemiologic and clinical aspects of the LF outbreak that occurred in Nigeria during January 1-May 6, 2018. A total of 1,893 cases were reported; 423 were laboratory-confirmed cases, among which 106 deaths were recorded (case-fatality rate 25.1%). Among all confirmed cases, 37 occurred in healthcare workers. The secondary attack rate among 5,001 contacts was 0.56%. Most (80.6%) confirmed cases were reported from 3 states (Edo, Ondo, and Ebonyi). Fatal outcomes were significantly associated with being elderly; no administration of ribavirin; and the presence of a cough, hemorrhaging, and unconsciousness. The findings in this study should lead to further LF research and provide guidance to those preparing to respond to future outbreaks.

Measures to control protracted large Lassa fever outbreak in Nigeria, 1 January to 28 April 2019.

Lassa fever cases have increased in Nigeria since 2016 with the highest number, 633 cases, reported in 2018. From 1 January to 28 April 2019, 554 laboratory-confirmed cases including 124 deaths were reported in 21 states in Nigeria. A public health emergency was declared on 22 January by the Nigeria Centre for Disease Control. We describe the various outbreak responses that have been implemented, including establishment of emergency thresholds and guidelines for case management.

Highly Efficient CRISPR/Cas9-Mediated Cloning and Functional Characterization of Gastric Cancer-Derived Epstein-Barr Virus Strains.

UNLABELLED: The Epstein-Barr virus (EBV) is etiologically linked to approximately 10% of gastric cancers, in which viral genomes are maintained as multicopy episomes. EBV-positive gastric cancer cells are incompetent for progeny virus production, making viral DNA cloning extremely difficult. Here we describe a highly efficient strategy for obtaining bacterial artificial chromosome (BAC) clones of EBV episomes by utilizing a CRISPR/Cas9-mediated strand break of the viral genome and subsequent homology-directed repair. EBV strains maintained in two gastric cancer cell lines (SNU719 and YCCEL1) were cloned, and their complete viral genome sequences were determined. Infectious viruses of gastric cancer cell-derived EBVs were reconstituted, and the viruses established stable latent infections in immortalized keratinocytes. While Ras oncoprotein overexpression caused massive vacuolar degeneration and cell death in control keratinocytes, EBV-infected keratinocytes survived in the presence of Ras expression. These results implicate EBV infection in predisposing epithelial cells to malignant transformation by inducing resistance to oncogene-induced cell death. IMPORTANCE: Recent progress in DNA-sequencing technology has accelerated EBV whole-genome sequencing, and the repertoire of sequenced EBV genomes is increasing progressively. Accordingly, the presence of EBV variant strains that may be relevant to EBV-associated diseases has begun to attract interest. Clearly, the determination of additional disease-associated viral genome sequences will facilitate the identification of any disease-specific EBV variants. We found that CRISPR/Cas9-mediated cleavage of EBV episomal DNA enabled the cloning of disease-associated viral strains with unprecedented efficiency. As a proof of concept, two gastric cancer cell-derived EBV strains were cloned, and the infection of epithelial cells with reconstituted viruses provided important clues about the mechanism of EBV-mediated epithelial carcinogenesis. This experimental system should contribute to establishing the relationship between viral genome variation and EBV-associated diseases.

Genetic Lineage and Reassortment of Influenza C Viruses Circulating between 1947 and 2014.

Since influenza C virus was first isolated in 1947, the virus has been only occasionally isolated by cell culture; there are only four strains for which complete genome sequences are registered. Here, we analyzed a total of 106 complete genomes, ranging from the first isolate from 1947 to recent isolates from 2014, to determine the genetic lineages of influenza C virus, the reassortment events, and the rates of nucleotide substitution. The results showed that there are six lineages, named C/Taylor, C/Mississippi, C/Aichi, C/Yamagata, C/Kanagawa, and C/Sao Paulo. They contain both antigenic and genetic lineages of the hemagglutinin-esterase (HE) gene, and the internal genes PB2, PB1, P3, NP, M, and NS are divided into two major lineages, a C/Mississippi/80-related lineage and a C/Yamagata/81-related lineage. Reassortment events were found over the entire period of 68 years. Several outbreaks of influenza C virus between 1990 and 2014 in Japan consisted of reassortant viruses, suggesting that the genomic constellation is related to influenza C virus epidemics. The nucleotide sequences were highly homologous to each other. The minimum percent identity between viruses ranged from 91.1% for the HE gene to 96.1% for the M gene, and the rate of nucleotide substitution for the HE gene was the highest, at 5.20 × 10(-4) substitutions/site/year. These results indicate that reassortment is an important factor that increases the genetic diversity of influenza C virus, resulting in its ability to prevail in humans. IMPORTANCE Influenza C virus is a pathogen that causes acute respiratory illness in children and results in hospitalization of infants. We previously demonstrated (Y. Matsuzaki et al., J Clin Virol 61:87-93, 2014, http://dx.doi.org/10.1016/j.jcv.2014.06.017) that periodic epidemics of this virus occurred in Japan between 1996 and 2014 and that replacement of the dominant antigenic group occurred every several years as a result of selection by herd immunity. However, the antigenicity of the HE glycoprotein is highly stable, and antigenic drift has not occurred for at least 30 years. Here, we analyzed a total of 106 complete genomes spanning 68 years for the first time, and we found that influenza C viruses are circulating worldwide while undergoing reassortment as well as selection by herd immunity, resulting in an increased ability to prevail in humans. The results presented in this study contribute to the understanding of the evolution, including reassortment events, underlying influenza C virus epidemics.

Differences in viral load among human respiratory syncytial virus genotypes in hospitalized children with severe acute respiratory infections in the Philippines.

BACKGROUND: Human respiratory syncytial virus (HRSV) is a leading viral etiologic agent of pediatric lower respiratory infections, including bronchiolitis and pneumonia. Two antigenic subgroups, HRSV-A and B, each contain several genotypes. While viral load may vary among HRSV genotypes and affect the clinical course of disease, data are scarce regarding the actual differences among genotypes. Therefore, this study estimated and compared viral load among NA1 and ON1 genotypes of HRSV-A and BA9 of HRSV-B. ON1 is a newly emerged genotype with a 72-nucleotide duplication in the G gene as observed previously with BA genotypes in HRSV-B. FINDINGS: Children <5 years of age with an initial diagnosis of severe or very severe pneumonia at a hospital in the Philippines from September 2012 to December 2013 were enrolled. HRSV genotypes were determined and the viral load measured from nasopharyngeal swabs (NPS). The viral load of HRSV genotype NA1 were significantly higher than those of ON1 and BA9. Regression analysis showed that both genotype NA1 and younger age were significantly associated with high HRSV viral load. CONCLUSIONS: The viral load of NA1 was higher than that of ON1 and BA9 in NPS samples. HRSV genotypes may be associated with HRSV viral load. The reasons and clinical impacts of these differences in viral load among HRSV genotypes require further evaluation.

Molecular epidemiology of enterovirus D68 from 2013 to 2014 in Philippines.

Enterovirus D68 (EV-D68) has been recognized as an important cause of acute respiratory infections. Here we report the molecular epidemiology of EV-D68 in Philippines from 2013 to 2014; we found cases in areas affected by Typhoon Haiyan and found new strains in the country.

Replication of many human viruses is refractory to inhibition by endogenous cellular microRNAs.

The issue of whether viruses are subject to restriction by endogenous microRNAs (miRNAs) and/or by virus-induced small interfering RNAs (siRNAs) in infected human somatic cells has been controversial. Here, we address this question in two ways. First, using deep sequencing, we demonstrate that infection of human cells by the RNA virus dengue virus (DENV) or West Nile virus (WNV) does not result in the production of any virus-derived siRNAs or viral miRNAs. Second, to more globally assess the potential of small regulatory RNAs to inhibit virus replication, we used gene editing to derive human cell lines that lack a functional Dicer enzyme and that therefore are unable to produce miRNAs or siRNAs. Infection of these cells with a wide range of viruses, including DENV, WNV, yellow fever virus, Sindbis virus, Venezuelan equine encephalitis virus, measles virus, influenza A virus, reovirus, vesicular stomatitis virus, human immunodeficiency virus type 1, or herpes simplex virus 1 (HSV-1), failed to reveal any enhancement in the replication of any of these viruses, although HSV-1, which encodes at least eight Dicer-dependent viral miRNAs, did replicate somewhat more slowly in the absence of Dicer. We conclude that most, and perhaps all, human viruses have evolved to be resistant to inhibition by endogenous human miRNAs during productive replication and that dependence on a cellular miRNA, as seen with hepatitis C virus, is rare. How viruses have evolved to avoid inhibition by endogenous cellular miRNAs, which are generally highly conserved during metazoan evolution, remains to be determined. Importance: Eukaryotic cells express a wide range of small regulatory RNAs, including miRNAs, that have the potential to inhibit the expression of mRNAs that show sequence complementarity. Indeed, previous work has suggested that endogenous miRNAs have the potential to inhibit viral gene expression and replication. Here, we demonstrate that the replication of a wide range of pathogenic viruses is not enhanced in human cells engineered to be unable to produce miRNAs, indicating that viruses have evolved to be resistant to inhibition by miRNAs. This result is important, as it implies that manipulation of miRNA levels is not likely to prove useful in inhibiting virus replication. It also focuses attention on the question of how viruses have evolved to resist inhibition by miRNAs and whether virus mutants that have lost this resistance might prove useful, for example, in the development of attenuated virus vaccines.

Seroprevalence and molecular characteristics of hepatitis E virus in household-raised pig population in the Philippines.

BACKGROUND: Hepatitis E virus (HEV) infection is a significant public health concern in Asia, and swine is an important source of sporadic HEV infection in human. However, no epidemiological data are available regarding HEV infection among the swine or human population in the Philippines. To assess the HEV infection status among pigs in rural areas, we investigated the molecular characteristics and seroprevalence of HEV among household-raised pigs in San Jose, Tarlac Province, the Philippines. RESULT: Serum and rectal swab samples were collected from 299 pigs aged 2-24 months from 155 households in four barangays (villages) between July 2010 and June 2011. Enzyme-linked immunosorbent assay (ELISA) revealed that 50.3% [95% confidence interval (CI) 44.5-56.2%] and 22.9% (95% CI 18.2-28.1%) of pigs tested positive for anti-HEV IgG and IgM, respectively. HEV RNA was detected in the feces of 22 pigs (7.4%, 95% CI 4.7-10.9%). A total of 103 households (66.5%, 95% CI 58.4-73.8%) had at least one pig that tested positive for anti-HEV IgG or IgM or HEV RNA. The prevalence of anti-HEV IgG and IgM in breeding pig (8-24 months) were higher than that in growing pigs (2-4 months) (p < 0.0001 and p = 0.008, respectively). HEV RNA was more frequently detected in 2-4-month-old pigs (9.2%, 95% CI 5.4-14.6%) than in ≥5-month-old pigs (4.8%, 95% CI 1.1-8.5%) without statistical significance (p = 0.142). HEV RNA showed 0-27.6% nucleotide difference at the partial ORF2 gene among the detected viruses, and a majority of them belonged to subtype 3a (20/22, 90.9%). CONCLUSION: We found a high prevalence of HEV antibodies in the household-raised pig population in rural areas of the Philippines, which indicates the potential risk of HEV infection among local residents. Only genotype 3 of HEV was observed, and genetically diverse strains of HEV were found to be circulating in pigs in this study.