Estimating the effective reproduction number of COVID-19 from population-wide wastewater data: An application in Kagawa, Japan.

Although epidemiological surveillance of COVID-19 has been gradually downgraded globally, the transmission of COVID-19 continues. It is critical to quantify the transmission dynamics of COVID-19 using multiple datasets including wastewater virus concentration data. Herein, we propose a comprehensive method for estimating the effective reproduction number using wastewater data. The wastewater virus concentration data, which were collected twice a week, were analyzed using daily COVID-19 incidence data obtained from Takamatsu, Japan between January 2022 and September 2022. We estimated the shedding load distribution (SLD) as a function of time since the date of infection, using a model employing the delay distribution, which is assumed to follow a gamma distribution, multiplied by a scaling factor. We also examined models that accounted for the temporal smoothness of viral load measurement data. The model that smoothed temporal patterns of viral load was the best fit model (WAIC = 2795.8), which yielded a mean estimated distribution of SLD of 3.46 days (95% CrI: 3.01-3.95 days). Using this SLD, we reconstructed the daily incidence, which enabled computation of the effective reproduction number. Using the best fit posterior draws of parameters directly, or as a prior distribution for subsequent analyses, we first used a model that assumed temporal smoothness of viral load concentrations in wastewater, as well as infection counts by date of infection. In the subsequent approach, we examined models that also incorporated weekly reported case counts as a proxy for weekly incidence reporting. Both approaches enabled estimations of the epidemic curve as well as the effective reproduction number from twice-weekly wastewater viral load data. Adding weekly case count data reduced the uncertainty of the effective reproduction number. We conclude that wastewater data are still a valuable source of information for inferring the transmission dynamics of COVID-19, and that inferential performance is enhanced when those data are combined with weekly incidence data.

Survey of exposure to stranded dolphins in Japan to investigate an outbreak of suspected infection with highly pathogenic avian influenza (H5N1) clade 2.3.4.4(b) in humans.

Background: A highly pathogenic avian influenza (HPAI) A (H5N1) virus has been detected in domestic and wild animals worldwide. The incidence of HPAI infections in sea mammals has been increasing, as is the number of stranded marine mammals linked to H5N1 viral clade 2.3.4.4(b). In this study, we investigated a stranding event involving dolphins and human-dolphin contact, and investigated the potential risk of animal-to-human H5N1 transmission with a survey of exposure on the Tsurigasaki coast, Japan. Methods: We performed a non-random, convenient-sample-based, survey on Tsurigasaki beach where around 30 melon-headed whales were stranded on April 3, 2023. Face-to-face (n = 25) and telephone (n = 1) interviews among surfers took place on April 7 and 8. A nasal swab for quick antigen testing was taken from those who wished to be tested (n = 13), to detect infections with influenza A virus. Results: Although there was no confirmatory diagnosis of H5N1 in either humans or dolphins (while n = 3 dolphins were autopsied), we found that a large number of surfers had touched the dolphins with their bare hands while attempting to rescue them, and that some surfers were directly exposed to dolphin blood and body fluids in the ocean. Conclusions: The adequate communication of risk is required to minimize the threat of viral transmission at this particular human-animal interface. Administrative and legal responses to cross-species transmission, including guidelines via one health frameworks, a rapid evaluation process of ethical approval, and the systematic involvement of experts in infectious disease, must be urgently formulated.

Life-expectancy changes from 2019 to 22: A case study of Japan using provisional death count.

Many countries struggled with suppressing the incidence of COVID-19 caused by SARS-CoV-2 Omicron variant (B.1.1.529). As the epidemic size of COVID-19 in 2022 became bigger than earlier years in Japan, the present study aimed to estimate life expectancy at birth at the end of 2022, using provisional death datasets in Aichi and Fukui prefectures. We collected monthly death count from 2019 to the end of 2022, computing the period life table. While the life expectancy at birth in Aichi, 2019 was 84.6 years, it was very slightly extended to 84.7 years in 2020 and 2021, followed by a shortening for nearly 0.4 years in 2022. In Fukui, monotonous extension pattern was seen, i.e., 85.5 years in 2019, 85.6 in 2020, followed by 85.8 and 86.2 years in 2021 and 2022, respectively. Although decades-long trend of extending life expectancy at birth was partly discontinued from 2020 due to the pandemic at the national level, we have shown that the pandemic impact was still small in Japan by the end of 2022. First Omicron wave occurred shortly after primary series vaccination, and even real time booster program was underway during that wave. Different demographic consequences between Aichi and Fukui are explained by differential epidemic sizes prior to vaccination.

Comparison of phylogenetic and virulence factors between Escherichia coli isolated from biliary tract infections and uropathogenic Escherichia coli.

Escherichia coli is a gram-negative intestinal commensal that can also cause various infections, including urinary tract infections, biliary tract infections, neonatal meningitis, and septicemia. Although the characteristics of uropathogenic E. coli and the mechanisms of urinary tract infection have been well studied, the genetic distinctions among E. coli isolates from different types of infections have not yet been determined. This study compared the phylogenetic and virulence factors of E. coli isolates from bacteremic biliary tract infections with those from bacteremic urinary tract infections. The phylogenetic B2 group was the most prevalent in both pathogenic groups (68 % in biliary pathogenic isolates and 85 % in uropathogenic isolates), but the frequency pattern of the phylogenetic group was different. Half of the uropathogenic isolates belonged to ST95 and ST131 (51 %). Among the biliary pathogenic isolates, ST131 was the most prevalent, while the remaining half belonged to other STs outside the four major STs. The frequency of some virulence factors, such as papC, papG2, hlyA, tcpC, fyuA, kpsMT2, sat, and traT, was lower in the biliary pathogenic isolates than in the uropathogenic isolates. The frequency of phylogenetic groups and STs in MLST differed between E. coli isolates from bacteremic biliary tract infections and urinary tract infections. Additionally, some virulence factors, including adhesion and toxin gene groups, showed lower frequencies in the biliary pathogenic group than in the uropathogenic group. Studying the differences in E. coli pathovars from different infection sites is important for developing pathovar-specific targeted therapies such as vaccine therapy.

Complexation study of a 1,3-phenylene-bridged cyclic hexa-naphthalene with fullerenes C60 and C70 in solution and 1D-alignment of fullerenes in the crystals.

To investigate the host ability of a simple macrocycle, 1,3-phenylene-bridged naphthalene hexamer N6, we evaluated the complexation of N6 with fullerenes in toluene and in the crystals. The complexes in the solid-state demonstrate the one-dimensional alignment of fullerenes. The single-crystals of the C60@N6 composite have semiconductive properties revealed by photoconductivity measurements.

Life-expectancy changes during the COVID-19 pandemic from 2019-2021: estimates from Japan, a country with low pandemic impact.

Background: The COVID-19 pandemic had a low impact on Japan in 2020, but the size of the epidemic increased considerably there in 2021. This study made a statistical analysis of life expectancy changes up to the end of 2021 in Japan. Objective: We aimed to estimate changes in life expectancy from 2019 to 2021 associated with the COVID-19 pandemic. To do so, we decomposed the life expectancy changes from 2020-2021 into age-specific and cause of death-specific contributions. Methods: We used the absolute number of deaths by age and prefecture in Japan to calculate life expectancy from 2019-21 at both national and prefectural levels, and also examined the correlation between life expectancy gap and annual number of COVID-19 cases, total person-days spent in intensive care, and documented deaths due to COVID-19. We used the Arriaga decomposition method to decompose national life expectancy changes from 2020 to 2021 into age and cause of death components. Results: From 2019-2020, Japan's national level life expectancy across the entire population was extended by 0.24 years. From 2020-2021, it shortened by 0.15 years. The life expectancy shortened more among women (0.15 years) than men (0.12 years). There was significant heterogeneity in life expectancy changes from 2020-2021 by prefecture. It ranged from the maximum shortening of 0.57 years in Tottori prefecture to the maximum extension of 0.23 years in Fukui. The regression analysis revealed the negative correlation between the life expectancy change and burden of COVID-19 at prefectural level. The decomposition of life expectancy changes at birth from 2020-2021 showed that losses in life expectancy were largely attributable to the mortality of the population over 70 years old. Changes in life expectancy among infants and working-age adults mostly contributed to lengthening overall life expectancy. Among leading major causes of death, deaths due to neoplastic tumor and cardiovascular diseases contributed to shortening life expectancy, whereas respiratory diseases did not. Conclusion: The decades-long increasing trend in life expectancy was suspended by the COVID-19 pandemic. However, life expectancy changes from 2019-2020 and 2020-2021 were small in Japan. This may be attributable to the small epidemiological impact of COVID-19 during this time period, but nonetheless, the negative impact of COVID-19 on life expectancy was indicated in the present study. The chance of death accelerated in older people in 2021, but a smaller number of deaths than usual among infants and working age adults contributed to extended life expectancy, and the change in the cause of death structure under the COVID-19 pandemic also significantly contributed to shortening life expectancy.

Elucidation of composition of chlorine compounds in acidic sodium chlorite solution using ion chromatography.

With the spread of coronavirus infections, the demand for disinfectants, such as a sodium chlorite solution, has increased worldwide. Sodium chlorite solution is a food additive and is used in a wide range of applications. There is evidence that chlorous acid or sodium chlorite is effective against various bacteria, but the actual mechanism is not well understood. One reason for this is that the composition of chlorine-based compounds contained in sodium chlorite solutions has not been clearly elucidated. The composition can vary greatly with pH. In addition, the conventional iodometric titration method, the N,N-diethyl-p-phenylenediamine sulfate (DPD) method and the absorption photometric method cannot clarify the composition. In this study, we attempted to elucidate the composition of a sodium chlorite solution using absorption spectrophotometry and ion chromatography (IC). IC is excellent for qualitative and quantitative analysis of trace ions. Through this, we aimed to develop an evaluation method that allows anyone to easily determine the bactericidal power of sodium chlorite. We found that commercially available sodium chlorite solution is 80% pure, with the remaining 20% potentially containing sodium hypochlorite solution. In addition, when sodium chlorite solution became acidified, its absorption spectrum exhibited a peak at 365 nm. Sodium chlorite solution is normally alkaline, and it cannot be measured by the DPD method, which is only applicable under acidic conditions. The presence of a peak at 365 nm indicates that the acidic sodium chlorite solution contains species with oxidizing power. On the other hand, the IC analysis showed a gradual decrease in chlorite ions in the acidic sodium chlorite solution. These results indicate that chlorite ions may not react with this DPD reagent, and other oxidizing species may be present in the acidic sodium chlorite solution. In summary, when a sodium chlorite solution becomes acidic, chlorine-based oxidizing species produce an absorption peak at 365 nm. Sodium hypochlorite and sodium chlorite solutions have completely different IC peak profiles. Although there are still many problems to be solved, we believe that the use of IC will facilitate the elucidation of the composition of sodium chlorite solution and its sterilization mechanism.

Night-time population consistently explains the transmission dynamics of coronavirus disease 2019 in three megacities in Japan.

Background: Mobility data are crucial for understanding the dynamics of coronavirus disease 2019 (COVID-19), but the consistency of the usefulness of these data over time has been questioned. The present study aimed to reveal the relationship between the transmissibility of COVID-19 in Tokyo, Osaka, and Aichi prefectures and the daily night-time population in metropolitan areas belonging to each prefecture. Methods: In Japan, the de facto population estimated from GPS-based location data from mobile phone users is regularly monitored by Ministry of Health, Labor, and Welfare and other health departments. Combined with this data, we conducted a time series linear regression analysis to explore the relationship between daily reported case counts of COVID-19 in Tokyo, Osaka, and Aichi, and night-time de facto population in downtown areas estimated from mobile phone location data, from February 2020 to May 2022. As an approximation of the effective reproduction number, the weekly ratio of cases was used. Models using night-time population with lags ranging from 7 to 14 days were tested. In time-varying regression analysis, the night-time population level and the daily change in night-time population level were included as explanatory variables. In the fixed-effect regression analysis, the inclusion of either the night-time population level or daily change, or both, as explanatory variables was tested, and autocorrelation was adjusted by introducing first-order autoregressive error of residuals. In both regression analyses, the lag of night-time population used in best fit models was determined using the information criterion. Results: In the time-varying regression analysis, night-time population level tended to show positive to neutral effects on COVID-19 transmission, whereas the daily change of night-time population showed neutral to negative effects. The fixed-effect regression analysis revealed that for Tokyo and Osaka, regression models with 8-day-lagged night-time population level and daily change were the best fit, whereas in Aichi, the model using only the 9-day-lagged night-time population level was the best fit using the widely applicable information criterion. For all regions, the best-fit model suggested a positive relationship between night-time population and transmissibility, which was maintained over time. Conclusion: Our results revealed that, regardless of the period of interest, a positive relationship between night-time population levels and COVID-19 dynamics was observed. The introduction of vaccinations and major outbreaks of Omicron BA. Two subvariants in Japan did not dramatically change the relationship between night-time population and COVID-19 dynamics in three megacities in Japan. Monitoring the night-time population continues to be crucial for understanding and forecasting the short-term future of COVID-19 incidence.

Protection against SARS-CoV-2 BA.4 and BA.5 subvariants via vaccination and natural infection: A modeling study.

With continuing emergence of new SARS-CoV-2 variants, understanding the proportion of the population protected against infection is crucial for public health risk assessment and decision-making and so that the general public can take preventive measures. We aimed to estimate the protection against symptomatic illness caused by SARS-CoV-2 Omicron variants BA.4 and BA.5 elicited by vaccination against and natural infection with other SARS-CoV-2 Omicron subvariants. We used a logistic model to define the protection rate against symptomatic infection caused by BA.1 and BA.2 as a function of neutralizing antibody titer values. Applying the quantified relationships to BA.4 and BA.5 using two different methods, the estimated protection rate against BA.4 and BA.5 was 11.3% (95% confidence interval [CI]: 0.01-25.4) (method 1) and 12.9% (95% CI: 8.8-18.0) (method 2) at 6 months after a second dose of BNT162b2 vaccine, 44.3% (95% CI: 20.0-59.3) (method 1) and 47.3% (95% CI: 34.1-60.6) (method 2) at 2 weeks after a third BNT162b2 dose, and 52.3% (95% CI: 25.1-69.2) (method 1) and 54.9% (95% CI: 37.6-71.4) (method 2) during the convalescent phase after infection with BA.1 and BA.2, respectively. Our study indicates that the protection rate against BA.4 and BA.5 are significantly lower compared with those against previous variants and may lead to substantial morbidity, and overall estimates were consistent with empirical reports. Our simple yet practical models enable prompt assessment of public health impacts posed by new SARS-CoV-2 variants using small sample-size neutralization titer data to support public health decisions in urgent situations.

A Cold-Blooded Tiptoer: Nonresolving Cellulitis in an Immunocompromised Patient.

Mycobacterium haemophilum is a nontuberculous mycobacteria (NTM) with a predilection for skin and soft tissue infection (SSTI) in the immunocompromised host. We report a case of disseminated M haemophilum infection initially presenting as a nonresolving subacute cellulitis of bilateral lower extremities. Genetic sequencing was used for final identification, while a commercially available polymerase chain reaction test returned a false-positive result for Mycobacterium intracellulare. Consequently, we highlight the importance of M haemophilum as a major differential diagnosis of SSTI in the immunocompromised host and the need for careful interpretation of rapid diagnostic tests.

Clinical and Microbiological Characteristics of Recurrent Escherichia coli Bacteremia.

The causative agents of recurrent Escherichia coli bacteremia can be genetically identical or discordant, but the differences between them remain unclear. This study aimed to explore these differences, with regard to their clinical and microbiological features. Patients were recruited from a Japanese tertiary teaching hospital based on blood culture data and the incidence of recurrent E. coli bacteremia. We compared the patients' clinical and microbiological characteristics between the two groups (those with identical or discordant E. coli bacteremia) divided by the result of enterobacterial repetitive intergenic consensus PCR. Among 70 pairs of recurrent E. coli bacteremia strains, 49 pairs (70%) were genetically identical. Patients with genetically identical or discordant E. coli bacteremia were more likely to have renal failure or neoplasms, respectively. The virulence factor (VF) scores of genetically identical E. coli strains were significantly higher than those of genetically discordant strains, with the prevalence of eight VF genes being significantly higher in genetically identical E. coli strains. No significant differences were found between the two groups regarding antimicrobial susceptibility and biofilm formation potential. This study showed that genetically identical E. coli bacteremia strains have more VF genes than genetically discordant strains in recurrent E. coli bacteremia. IMPORTANCE Escherichia coli causes bloodstream infection, although not all strains are pathogenic to humans. In some cases, this infection reoccurs, and several reports have described the clinical characteristics and/or molecular microbiology of recurrent Escherichia coli bacteremia. However, these studies focused on patients with specific characteristics, and they included cases caused by microorganisms other than Escherichia coli. Hence, little is known about the pathogenicity of Escherichia coli isolated from the recurrent one. The significance of our study is in evaluating the largest cohorts to date, as no cohort studies have been conducted on this topic.

Cancer-microenvironment triggered self-assembling therapy with molecular blocks.

Drug delivery systems (DDS) have been studied in an effort to reduce side effects by increasing the accumulation of anticancer drugs in cancer cells. However, the transport efficiency is still low due to the blocking by surrounding stromal tissues and the multiple intracellular drug transportation processes required to get the drug to a target cytosol. Thus, improving the efficiency of cancer therapy is still a major challenge. Here, a drug-free cancer microenvironment-targeting therapy using molecular blocks (MBs) is demonstrated, which is designed for efficient blood circulation and penetration through the stromal tissues as either a single molecule or a few molecules. When the MBs moved to a cancer microenvironment by the enhanced permeability and retention effect, they formed a self-assembled aggregate on the cancer cell surfaces in response to the weak acid (pH ∼ 6.5) condition leading to subsequent cancer cell death by membrane disruption. This strategy avoids multiple intracellular transportation processes and also stimulates cell membrane disruption by self-assembly of the MB via hydrophobic interactions. Deoxycholic acid (DCA) was selected as a cancer microenvironment-responsive unit because its pKa = 6.6. The DCA conjugated 4-arm poly(ethylene glycol) (4-MB) showed self-assembly phenomena on cancer cell membranes and subsequently significant cytotoxicity was clearly observed. Moreover, they clearly showed efficient accumulation in the tumor and the effective suppression of tumor growth in in vivo experiments. This MB therapy will be a new strategy for addressing the current issues of DDS.

The prevalence of the iutA and ibeA genes in Escherichia coli isolates from severe and non-severe patients with bacteremic acute biliary tract infection is significantly different.

BACKGROUND: Although Escherichia coli is the most frequently isolated microorganism in acute biliary tract infections with bacteremia, data regarding its virulence are limited. RESULTS: Information on cases of bacteremia in acute biliary tract infection in a retrospective study was collected from 2013 to 2015 at a tertiary care hospital in Japan. Factors related to the severity of infection were investigated, including patient background, phylogenetic typing, and virulence factors of E. coli, such as adhesion, invasion, toxins, and iron acquisition. In total, 72 E. coli strains were identified in 71 cases, most of which primarily belonged to the B2 phylogroup (68.1%). The presence of the iutA gene (77.3% in the non-severe group, 46.4% in the severe group, P = 0.011) and the ibeA gene (9.1% in the non-severe group, and 35.7% in the severe group, P = 0.012) was significantly associated with the severity of infection. Among the patient characteristics, diabetes mellitus with organ involvement and alkaline phosphatase were different in the severe and non-severe groups. CONCLUSIONS: We showed that bacteremic E. coli strains from acute biliary tract infections belonged to the virulent (B2) phylogroup. The prevalence of the iutA and ibeA genes between the two groups of bacteremia severity was significantly different.

First Reported Human Case of Spondylodiscitis by Staphylococcus condimenti: A Case Report and Literature Review.

Staphylococcus condimenti is a Gram-positive coccus that was first isolated from soy sauce mash. Only four cases of human S. condimenti infections have been reported to date. We herein report the first case of spondylodiscitis caused by S. condimenti. A 72-year-old Japanese man complaining of lower back pain and numbness in his legs was diagnosed with spondylodiscitis. A computed tomography (CT)-guided biopsy was performed. A culture of the intravertebral disc aspirate yielded S. condimenti. The result was confirmed using gene sequencing methods. The patient was successfully treated without relapse. This case shows that S. condimenti can be pathogenic and cause invasive infection.

Genetic diversity and epidemiology of accessory gene regulator loci in Clostridioides difficile.

Quorum sensing is known to regulate bacterial virulence, and the accessory gene regulator (agr) loci is one of the genetic loci responsible for its regulation. Recent reports examining Clostridioides difficile show that two agr loci, agr1 and agr2, regulate toxin production, but the diversity of agr loci and their epidemiology is unknown. In our study, in silico analysis was performed to research genetic diversity of agr, and C. difficile isolates from clinical samples underwent multilocus sequence typing (MLST) and PCR analysis of agr loci. To reveal the distribution of agr among different strains, phylogenetic analysis was also performed. In our in silico analysis, two different subtypes, named agr2R and agr2M, were found in agr2, which were previously reported. PCR analysis of 133 C . difficile isolates showed that 131 strains had agr1, 61 strains had agr2R, and 26 strains had agr2M; agr2R was mainly found in clade 1 or clade 2 organisms, whereas agr2M was only found in clade 4. With rare exception, agr1-negative sequence types (STs) belonged to clade C-Ⅰ and C-Ⅲ, and one clade 4 strain had agr2R. Our study revealed subtypes of agr2 not previously recognized, and the distribution of several agr loci in C. difficile . These findings provide a foundation for further functional and clinical research of the agr loci.

Vibrio fluvialis Liver Abscess and Bacteremia in a Sashimi Lover: A Case Report and Review of the Literature.

Vibrio fluvialis is a foodborne pathogen known to cause a cholera-like gastroenteritis syndrome. Here we report the first case of V. fluvialis liver abscess and bacteremia presumed to be from sashimi, a Japanese raw seafood delicacy. We also provide a literature review of reported cases of V. fluvialis extra-intestinal diseases including bacteremia.

The First Case Report of Acute Symptomatic HEV Genotype 4 Infection in an HIV-positive Patient in Japan.

Hepatitis E virus (HEV) is a common cause of acute hepatitis. Four major genotypes of HEV have been studied, with genotype 4 being the predominant genotype across Asia. We herein describe the case of a 50-year-old man with a history of human immunodeficiency virus (HIV) infection who was admitted with acute transaminitis. Serum anti-HEV-IgA and HEV-RNA were detected at the time of presentation and further testing revealed HEV genotype 4. To the best of our knowledge, this represents the first clinical case report of acute symptomatic HEV genotype 4 infection in an HIV-positive patient in Japan.

Phenology-dependent cold exposure and thermal performance of Ostrinia nubilalis ecotypes.

BACKGROUND: Understanding adaptation involves establishing connections between selective agents and beneficial population responses. However, relatively little attention has been paid to seasonal adaptation, in part, because it requires complex and integrative knowledge about seasonally fluctuating environmental factors, the effects of variable phenology on exposure to those factors, and evidence for temporal specialization. In the European corn borer moth, Ostrinia nubilalis, sympatric pheromone strains exploit the same host plant (Zea mays) but may genetically differ in phenology and be reproductively "isolated by time." Z strain populations in eastern North America have been shown to have a prolonged larval diapause and produce one annual mating flight (July), whereas E strain populations complete an earlier (June) and a later (August) mating flight by shortening diapause duration. Here, we find evidence consistent with seasonal "adaptation by time" between these ecotypes. RESULTS: We use 12 years of field observation of adult seasonal abundance to estimate phenology of ecotype life cycles and to quantify life-stage specific climatic conditions. We find that the observed reduction of diapause duration in the E strain leads their non-diapausing, active life stages to experience a ~ 4 °C colder environment compared to the equivalent life stages in the Z strain. For a representative pair of populations under controlled laboratory conditions, we compare life-stage specific cold tolerance and find non-diapausing, active life stages in the E strain have as much as a 60% greater capacity to survive rapid cold shock. Enhanced cold hardiness appears unrelated to life-stage specific changes in the temperature at which tissues freeze. CONCLUSIONS: Our results suggest that isolation by time and adaptation by time may both contribute to population divergence, and they argue for expanded study in this species of allochronic populations in nature experiencing the full spectrum of seasonal environments. Cyclical selective pressures are inherent properties of seasonal habitats. Diverse fluctuating selective agents across each year (temperature, predation, competition, precipitation, etc.) may therefore be underappreciated drivers of biological diversity.

Epidemiology and virulence-associated genes of Clostridioides difficile isolates and factors associated with toxin EIA results at a university hospital in Japan.

INTRODUCTION: Clostridioides difficile is one of the most important nosocomial pathogens; however, reports regarding its clinical and molecular characteristics from Japan are scarce. AIMS: We studied the multilocus sequence typing (MLST)-based epidemiology and virulence-associated genes of isolates and the clinical backgrounds of patients from whom the isolates had been recovered. METHODS: A total of 105 stool samples tested in a C. difficile toxin enzyme immune assay (EIA) were analysed at the University of Tokyo Hospital from March 2013 to July 2014. PCR for MLST and the virulence-associated genes tcdA, tcdB, cdtA, cdtB and tcdC was performed on C. difficile isolates meeting our inclusion criteria following retrospective review of medical records. EIA-positive and EIA-negative groups with toxigenic strains underwent clinical and molecular background comparison. RESULTS: The toxigenic strains ST17, ST81, ST2, ST54, ST8, ST3, ST37 and ST53 and the non-toxigenic strains ST109, ST15 and ST100 were frequently recovered. The prevalence rate of tcdA-negative ST81 and ST37, endemic in China and Korea, was higher (11.4%) than that reported in North America and Europe, and hypervirulent ST1(RT027) and ST11(RT078) strains that occur in North America and Europe were not recovered. The linkage between the EIA results and cdt A/B positivity, tcdC deletion, or tcdA variation was absent among toxigenic strains. Compared with the 38 EIA-negative cases, the 36 EIA-positive cases showed that the patients in EIA-positive cases were older and more frequently had chronic kidney disease, as well as a history of beta-lactam use and proton pump inhibitor therapy. CONCLUSION: In Japan, the prevalence rates for tcdA-negative strains are high, whereas the cdtA/B-positive strains are rare. EIA positivity is linked to older age, chronic kidney disease and the use of beta-lactams and proton pump inhibitors.