Changing Patterns of Infectious Diseases Among Hospitalized Children in Hokkaido, Japan, in the Post-COVID-19 Era, July 2019 to June 2022.

BACKGROUND: Many reports have reported a reduction in respiratory infectious diseases and infectious gastroenteritis immediately after the coronavirus disease 2019 (COVID-19) pandemic, but data continuing into 2022 are very limited. We sought to understand the current situation of various infectious diseases among children in Japan as of July 2022 to improve public health in the post-COVID-19 era. METHODS: We collected data on children hospitalized with infectious diseases in 18 hospitals in Japan from July 2019 to June 2022. RESULTS: In total, 3417 patients were hospitalized during the study period. Respiratory syncytial virus decreased drastically after COVID-19 spread in early 2020, and few patients were hospitalized for it from April 2020 to March 2021. However, an unexpected out-of-season re-emergence of respiratory syncytial virus was observed in August 2021 (50 patients per week), particularly prominent among older children aged 3-6 years. A large epidemic of delayed norovirus gastroenteritis was observed in April 2021, suggesting that the nonpharmaceutical interventions for COVID-19 are less effective against norovirus. However, influenza, human metapneumovirus, Mycoplasma pneumoniae, and rotavirus gastroenteritis were rarely seen for more than 2 years. CONCLUSIONS: The incidence patterns of various infectious diseases in Japan have changed markedly since the beginning of the COVID-19 pandemic to the present. The epidemic pattern in the post-COVID-19 era is unpredictable and will require continued careful surveillance.

Incidence of and risk factors for suspected COVID-19 reinfection in Kyoto City: a population-based epidemiological study.

To determine the clinical characteristics of and risk factors for suspected reinfection with coronavirus 2019 (COVID-19). This was a retrospective cohort study using population-based notification records of residents in Kyoto City (1.4 M) with laboratory-confirmed COVID-19 infection between 1 March 2020 and 15 April 2022. Reinfection was defined by two or more positive COVID-19 test results ≧ 90 days apart. Demographic characteristics, the route and timing of infection and history of vaccination were analysed to identify risk factors for reinfection. Among the cohort of 107,475 patients, reinfection was identified in 0.66% (n = 709). The age group with the highest reinfection rate was 18-39 years (1.06%), followed by 40-59 years (0.58%). Compared to the medical and nursing professionals, individuals who worked in the construction and manufacturing industry (odds ratio [OR]: 2.86; 95% confidence interval [CI]: 1.66-4.92) and hospitality industry (OR: 2.05; 95% CI: 1.28-.31) were more likely to be reinfected. Symptomatic cases at initial infection, receiving more than 2 doses of vaccination and risk factors for severe infection at initial infection were protective factors against reinfection. Of the reinfected individuals, the reinfection route was unknown in 65%. Reinfection with COVID-19 is uncommon, with suspected reinfections more likely in adults, those with high exposure and unvaccinated individuals; the reinfection route was unknown in the majority of cases. This study confirmed the need to continue with self-protection efforts and to implement vaccination programs in high-risk populations.

Elucidation of the liver pathophysiology of COVID-19 patients using liver-on-a-chips.

SARS-CoV-2 induces severe organ damage not only in the lung but also in the liver, heart, kidney, and intestine. It is known that COVID-19 severity correlates with liver dysfunction, but few studies have investigated the liver pathophysiology in COVID-19 patients. Here, we elucidated liver pathophysiology in COVID-19 patients using organs-on-a-chip technology and clinical analyses. First, we developed liver-on-a-chip (LoC) which recapitulating hepatic functions around the intrahepatic bile duct and blood vessel. We found that hepatic dysfunctions, but not hepatobiliary diseases, were strongly induced by SARS-CoV-2 infection. Next, we evaluated the therapeutic effects of COVID-19 drugs to inhibit viral replication and recover hepatic dysfunctions, and found that the combination of anti-viral and immunosuppressive drugs (Remdesivir and Baricitinib) is effective to treat hepatic dysfunctions caused by SARS-CoV-2 infection. Finally, we analyzed the sera obtained from COVID-19 patients, and revealed that COVID-19 patients, who were positive for serum viral RNA, are likely to become severe and develop hepatic dysfunctions, as compared with COVID-19 patients who were negative for serum viral RNA. We succeeded in modeling the liver pathophysiology of COVID-19 patients using LoC technology and clinical samples.

Impaired CD4+ T cell response in older adults is associated with reduced immunogenicity and reactogenicity of mRNA COVID-19 vaccination.

Whether age-associated defects in T cells impact the immunogenicity and reactogenicity of mRNA vaccines remains unclear. Using a vaccinated cohort (n = 216), we demonstrated that older adults (aged ≥65 years) had fewer vaccine-induced spike-specific CD4+ T cells including CXCR3+ circulating follicular helper T cells and the TH1 subset of helper T cells after the first dose, which correlated with their lower peak IgG levels and fewer systemic adverse effects after the second dose, compared with younger adults. Moreover, spike-specific TH1 cells in older adults expressed higher levels of programmed cell death protein 1, a negative regulator of T cell activation, which was associated with low spike-specific CD8+ T cell responses. Thus, an inefficient CD4+ T cell response after the first dose may reduce the production of helper T cytokines, even after the second dose, thereby lowering humoral and cellular immunity and reducing systemic reactogenicity. Therefore, enhancing CD4+ T cell response following the first dose is key to improving vaccine efficacy in older adults.

Analytical and clinical performances of seven direct detection assays for SARS-CoV-2.

Background: Direct detection tests for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that bypass complicated nucleic acid/antigen purification steps are promising tools for the rapid diagnosis of coronavirus disease 2019 (COVID-19). Methods: To determine the analytical and clinical diagnostic performances of the direct detection assays, we compared 6 direct molecular detection assays, including two loop-mediated isothermal amplification (LAMP) assays and one lateral flow antigen assay, against the reference extraction-based RT-PCR assay using 183 respiratory samples (87 nasopharyngeal swabs, 51 saliva samples, and 45 sputum samples). Results: Analytical sensitivity analysis showed that the direct RT-PCR assay of Toyobo exhibited the lowest LOD of 1,000 copies/mL. Compared with the 80 positive and 103 negative samples based on the reference assay, the Toyobo assay had the highest positive percent agreement (PPA) of 96.3%, followed by the two direct RT-PCR assays of Takara and Shimadzu and one LAMP assay of Eiken (86.3-87.5%). The Fujirebio antigen assay had the lowest PPA of 44.7% among the assays tested. The negative percent agreement of these direct detection assays was 100%, except for the Eiken assay (96.3%). Conclusions: Large differences in PPA existed among the direct detection tests. Laboratories need to take these characteristics into consideration before implementing these assays.

SARS-CoV-2 disrupts respiratory vascular barriers by suppressing Claudin-5 expression.

In the initial process of coronavirus disease 2019 (COVID-19), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infects respiratory epithelial cells and then transfers to other organs the blood vessels. It is believed that SARS-CoV-2 can pass the vascular wall by altering the endothelial barrier using an unknown mechanism. In this study, we investigated the effect of SARS-CoV-2 on the endothelial barrier using an airway-on-a-chip that mimics respiratory organs and found that SARS-CoV-2 produced from infected epithelial cells disrupts the barrier by decreasing Claudin-5 (CLDN5), a tight junction protein, and disrupting vascular endothelial cadherin-mediated adherens junctions. Consistently, the gene and protein expression levels of CLDN5 in the lungs of a patient with COVID-19 were decreased. CLDN5 overexpression or Fluvastatin treatment rescued the SARS-CoV-2-induced respiratory endothelial barrier disruption. We concluded that the down-regulation of CLDN5 expression is a pivotal mechanism for SARS-CoV-2-induced endothelial barrier disruption in respiratory organs and that inducing CLDN5 expression is a therapeutic strategy against COVID-19.

High mortality and morbidity among vaccinated residents infected with the SARS-CoV-2 Omicron variant during an outbreak in a nursing home in Kyoto City, Japan.

BACKGROUND: Outbreaks of coronavirus disease 2019 (COVID-19) in long-term care facilities are associated with mortality, although vaccination have contributed to improvements. This study reports clinical impacts of a COVID-19 outbreak in a nursing home for elderly individuals in Kyoto City, Japan. METHODS: We performed epidemiologic and molecular investigations of the outbreak and characterized outcomes of the nursing home residents. RESULTS: During the outbreak period, a total of 31 residents (39.2%) and 26 staff members (49.1%) were infected with COVID-19. All residents and staff received two doses of a vaccine approximately 7 months prior. Ten residents with severe hypoxemia could not be transferred to a hospital due to a shortage of beds for COVID-19 patients. Within 90 days of the onset of the outbreak, 8 residents with COVID-19 (25.8%) died. A total of 48.4% of residents with COVID-19 developed 1 or more comorbidities. Viral genome analysis showed that the outbreak was caused by the Omicron BA.1.1.2 variant. CONCLUSIONS: Despite vaccination, high mortality and morbidity were observed in the COVID-19 outbreak due to the Omicron variant. Limiting medical care for residents with COVID-19 in facilities that experience ongoing outbreaks may be needed to reduce the risk of mortality among nursing home residents.

Heme oxygenase-1 as an important predictor of the severity of COVID-19.

BACKGROUND AND OBJECTIVE: A cytokine storm is caused by inflammatory cells, including pro-inflammatory macrophage phenotype (M1), and play a critical role in the pathogenesis of COVID-19, in which diffuse alveolar damage occurs in the lungs due to oxidative stress exposure. Heme oxygenase (HO)-1 is a stress-induced protein produced by the anti-inflammatory / anti-oxidative macrophage phenotype (M2), which also produces soluble CD163 (sCD163). In our study, we investigated and determined that serum HO-1 can be a predictive biomarker for assessing both the severity and the outcome of COVID-19 patients. METHOD: The serum concentrations of HO-1 and sCD163 of COVID-19 patients were measured on admission. The relationship between these biomarkers and other clinical parameters and outcomes were evaluated. RESULTS: Sixty-four COVID-19 patients (11 mild, 38 moderate, and 15 severe cases) were assessed. The serum HO-1 tended to increase (11.0 ng/mL vs. 24.3 ng/mL vs. 59.6 ng/mL with severity). Serum HO-1 correlated with serum lactate dehydrogenase (R = 0.422), C-reactive protein (R = 0.463), and the ground glass opacity (GGO) and consolidation score (R = 0.625) of chest computed tomography. The serum HO-1 showed a better area under the curve (AUC) for predicting ICU admission than the serum sCD163 (HO-1; 0.816 and sCD163; 0.743). In addition, composite parameters including serum HO-1 and the GGO and consolidation score showed a higher AUC for predicting ICU admission than the AUC of a single parameter. CONCLUSION: Clinically, serum HO-1, reflecting the activation of M2, could be a very useful marker for evaluating disease severity and predicting prognoses for COVID-19 patients. In addition, controlling activated M2 might be a preventative COVID-19 therapeutic target.

Transmissibility of SARS-CoV-2 B.1.1.214 and Alpha Variants during 4 COVID-19 Waves, Kyoto, Japan, January 2020-June 2021.

Household transmission is a primary source of SARS-CoV-2 spread. We used COVID-19 epidemiologic investigation data and viral genome analysis data collected in the city of Kyoto, Japan, during January 2020-June 2021 to evaluate the effects of different settings and viral strains on SARS-CoV-2 transmission. Epidemiologic investigations of 5,061 COVID-19 cases found that the most common category for close contact was within households (35.3%); this category also had the highest reverse transcription PCR positivity. The prevalent viral lineage shifted from B.1.1.214 in the third wave to the Alpha variant in the fourth wave. The proportion of secondary cases associated with households also increased from the third to fourth waves (27% vs. 29%). Among 564 contacts from 206 households, Alpha variant was significantly associated with household transmission (odds ratio 1.52, 95% CI 1.06-2.18) compared with B.1.1.214. Public health interventions targeting household contacts and specific variants could help control SARS-CoV-2 transmission.

Large-scale serosurveillance of COVID-19 in Japan: Acquisition of neutralizing antibodies for Delta but not for Omicron and requirement of booster vaccination to overcome the Omicron's outbreak.

Continuous appearance of SARS-CoV-2 variants and mass vaccination have been intricately influencing on the COVID-19 situation. To elucidate the current status in Japan, we analyzed totally 2,000 sera in August (n = 1,000) and December (n = 1,000) 2021 collected from individuals who underwent a health check-up. The anti-N seropositive rate were 2.1% and 3.9% in August and December 2021, respectively, demonstrating a Delta variant endemic during that time; it was approximately twofold higher than the rate based on the PCR-based diagnosis. The anti-S seropositive rate was 38.7% in August and it reached 90.8% in December, in concordance with the vaccination rate in Japan. In the December cohort, 78.7% of the sera showed neutralizing activity against the Delta variant, whereas that against the Omicron was much lower at 36.6%. These analyses revealed that effective immunity against the Delta variant was established in December 2021, however, prompt three-dose vaccination is needed to overcome Omicron's outbreak.

Comparison of six antibody assays and two combination assays for COVID-19.

INTRODUCTION: In this work, six SARS-CoV-2-specific antibody assays were evaluated, namely, two pan-immunoglobulin (pan-Ig) assays [Roche Elecsys Anti-SARS-CoV-2 (named "Elecsys" in this study) and the PerkinElmer SuperFlex™ Anti-SARS-CoV-2 Ab Assay (SuperFlex_Ab)], two IgM assays [SuperFlex™ Anti-SARS-CoV-2 IgM Assay (SuperFlex_IgM) and YHLO iFlash-SARS-CoV-2 IgM (iFlash_IgM)], and two IgG assays [SuperFlex™ Anti-SARS-CoV-2 IgG Assay (SuperFlex_IgG) and iFlash-SARS-CoV-2 IgG (iFlash_IgG)]. Combination assays of SuperFlex™ (SuperFlex_any) and iFlash (iFlash_any) were also evaluated. METHODS: A total of 438 residual serum samples from 54 COVID-19 patients in the COVID-19 group and 100 samples from individuals without evidence of SARS-CoV-2 infection in the negative control group were evaluated. RESULTS: In the early stage of COVID-19 infection, within 14 days of symptom onset, the seropositive rate was lower than that of the late stage 15 days after onset (65.4% vs 99.6%). In the total period, the pan-Ig and IgG assays had higher sensitivity (90.8-95.3%) than the IgM assays (36.5-40.7%). SuperFlex_Ab and SuperFlex_any had higher sensitivity than Elecsys and SuperFlex_IgG (p < 0.05). The specificity of all the assays was 100%, except for SuperFlex_IgM (99.0%). The concordance rate between each assay was higher (96.4-100%) in the late stage than in the early stage (77.4-98.1%). CONCLUSION: For the purpose of COVID-19 diagnosis, antibody testing should be performed 15 days after onset. For the purpose of epidemiological surveillance, highly sensitive assays should be used as much as possible, such as SuperFlex_Ab, iFlash_IgG and their combination. IgM assays were not suitable for these purposes.

Analysis of a city-wide COVID-19 prevention strategy for aged-care facilities during third and fifth waves of COVID-19 in Kyoto City, Kyoto, Japan.

BACKGROUND: During the third wave of the COVID-19 pandemic at the end of 2020, clusters occurred frequently in aged-care facilities (ACFs), which put pressure on the medical field in Japan. Based on this experience, Kyoto University and Kyoto City collaborated to promote a citywide COVID-19 prevention strategy to prevent the spread of COVID-19 within ACFs. The aim of this study was to clarify the effect of the prevention strategy among ACFs in Kyoto City during the third and fifth waves of the pandemic. METHODS: During the study period, the following measures were adopted as the prevention strategy in all ACFs: (1) active polymerase chain reaction (PCR) mass testing and facility-wide testing when a single case was identified, (2) implementation of strategies to prevent transmission within a facility, and (3) vaccination program for ACFs. RESULTS: Of the 1,144 facilities subjected to the mass testing, 71.0% participated in the whole program including active PCR testing. The remainder participated in the rest of the programs. The prevalence of ACF-related COVID-19 cases among total COVID-19 cases in Kyoto City decreased from 7.9% in the third wave to 4.1% in the fourth wave and 2.1% in the fifth wave. The incidence of clusters and proportion of severe elderly cases also decreased during the study period. CONCLUSIONS: A city-wide multidisciplinary effort including PCR mass testing and a vaccination program in cooperation with a university and local administrative office successfully reduced the clusters and transmission in ACFs in Kyoto City, Japan.

CRISPR-Cas3-based diagnostics for SARS-CoV-2 and influenza virus.

CRISPR-based diagnostics (CRISPR-dx), including the Cas12-based DETECTR and Cas13-based SHERLOCK Class 2 CRISPRs, have been used to detect the presence of DNA or RNA from pathogens, such as the 2009 pandemic influenza virus A (IAV) and the 2019 novel coronavirus SARS-CoV-2. Here, we describe the collateral single-stranded DNA cleavage with Class 1 type I CRISPR-Cas3 and highlight its potential for development as a Cas3-mediated rapid (within 40 min), low-cost, instrument-free detection method for SARS-CoV-2. This assay, which we call Cas3-Operated Nucleic Acid detectioN (CONAN), not only detects SARS-CoV-2 in clinical samples, but also offers specific detection of single-base-pair mutations in IAV variants. This tool allows rapid and accurate point-of-care testing for patients with suspected SARS-CoV-2 or drug-resistant IAV infections in hospitals.

Comparison of 12 Molecular Detection Assays for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2).

Molecular testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the mainstay for accurate diagnosis of the infection, but the diagnostic performances of available assays have not been defined. We compared 12 molecular diagnostic assays, including 8 commercial kits using 155 respiratory samples (65 nasopharyngeal swabs, 45 oropharyngeal swabs, and 45 sputum) collected at two Japanese hospitals. Sixty-eight samples were positive for more than one assay and one genetic locus, and were defined as true-positive samples. All the assays showed a specificity of 100% (95% CI, 95.8%-100%). The N2 assay kit of the US Centers for Disease Control and Prevention and the N2 assay of the Japanese National Institute of Infectious Disease (NIID) were the most sensitive assays with 100% sensitivity (95% CI, 94.7-100), followed by the Centers for Disease Control and Prevention N1 kit, E assay by Corman, and Japanese National Institute of Infectious Disease N2 assay multiplex with internal control reactions. These assays are reliable as first-line molecular assays in laboratories when combined with appropriate internal control reactions.