A molecular epidemiological and transmission analysis of Clostridioides difficile using draft whole-genome sequencing in a single hospital.

BACKGROUND: The nosocomial transmission of toxin-producing Clostridioides difficile is a significant concern in infection control. C. difficile, which resides in human intestines, poses a risk of transmission, especially when patients are in close contact with medical staff. METHODS: To investigate the nosocomial transmission of C. difficile in a single center, we analyzed the genetic relationships of the bacteria. This was done using draft whole-genome sequencing (WGS) and examining single nucleotide polymorphisms (SNPs) in core-genome, alongside data regarding the patient's hospital wards and room changes. Our retrospective analysis covered 38 strains, each isolated from a different patient, between April 2014 and January 2015. RESULTS: We identified 38 strains that were divided into 11 sequence types (STs). ST81 was the most prevalent (n = 11), followed by ST183 (n = 10) and ST17 (n = 7). A cluster of strains that indicated suspected nosocomial transmission (SNT) was identified through SNP analysis. The draft WGS identified five clusters, with 16 of 38 strains belonging to these clusters. There were two clusters for ST81 (ST81-SNT-1 and ST81-SNT-2), two for ST183 (ST183-SNT-1 and ST183-SNT-2), and one for ST17 (ST17-SNT-1). ST183-SNT-1 was the largest SNT cluster, encompassing five patients who were associated with Wards A, B, and K. The most frequent room changer was a patient labeled Pt08, who changed rooms seven times in Ward B. Patients Pt36 and Pt10, who were also in Ward B, had multiple admissions and discharges during the study period. CONCLUSIONS: Additional culture tests and SNP analysis of C. difficile using draft WGS revealed silent transmission within the wards, particularly in cases involving frequent room changes and repeated admissions and discharges. Monitoring C. difficile transmission using WGS-based analysis could serve as a valuable marker in infection control management.

Pseudomonas otitidis bacteremia in an immunocompromised patient with cellulitis: case report and literature review.

BACKGROUND: Pseudomonas otitidis belongs to the genus Pseudomonas and causes various infections, including ear, skin, and soft tissue infections. P. otitidis has a unique susceptibility profile, being susceptible to penicillins and cephalosporins but resistant to carbapenems, due to the production of the metallo-ß-lactamase called POM-1. This revealed genetic similarities with Pseudomonas aeruginosa, which can sometimes lead to misidentification. CASE PRESENTATION: We report the case of a 70-year-old Japanese male who developed cellulitis and bacteremia during chemotherapy for multiple myeloma. He was initially treated with meropenem, but blood culture later revealed gram-negative bacilli identified as P. otitidis using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Carbapenem resistance was predicted from previous reports; therefore, we switched to dual therapy with levofloxacin and cefepime, and favorable treatment results were obtained. CONCLUSION: This is the first reported case of P. otitidis cellulitis and bacteremia in an immunocompromised patient. Carbapenems are typically used in immunocompromised patients and P. otitidis is often resistant to it. However, its biochemical properties are similar to those of Pseudomonas aeruginosa; therefore, its accurate identification is critical. In the present study, we rapidly identified P. otitidis using MALDI-TOF MS and switched from carbapenems to an appropriate antimicrobial therapy, resulting in a successful outcome.

Oral fidaxomicin versus vancomycin for the treatment of Clostridioides difficile infection: A systematic review and meta-analysis of randomized controlled trials.

BACKGROUND: Fidaxomicin (FDX) has received considerable attention as a novel therapeutic alternative agent to vancomycin (VCM) for Clostridioides difficile infection (CDI). However, the superiority and efficacy profile of FDX are not sufficiently determined by high-quality evidence. This study aimed to clarify the superiority of FDX for CDI treatment through a systematic review and meta-analysis. METHODS: We conducted a meta-analysis of randomized controlled trials (RCTs) which evaluated the efficacy and safety of FDX and VCM in patients with CDI. Electronic databases (PubMed, Cochrane Library, Web of Science, and Clinicaltrials.gov) were searched for studies published until October 15, 2021. The primary endpoint was global cure. The secondary endpoints were clinical cure, recurrence, and adverse event. Risk ratios (RRs), risk differences (RDs), and 95% confidence intervals were calculated using Mantel-Haenszel random-effects model. The risk of bias was assessed using Cochrane Handbook for Systematic Reviews of Interventions and Assessment Criteria. RESULTS: Six RCTs were included in this meta-analysis. Compared to VCM, FDX was associated with significantly higher global cure rates (RR = 1.18, P < 0.00001; RD = 0.11, 95% CI = 0.07-0.16). In addition, clinical cure rates were comparable between FDX and VCM (P = 0.31). FDX was associated with significantly lower recurrence rates compared to VCM (RR = 0.59, P < 0.0001). In addition, adverse event rates were not significantly different between the drugs (P = 0.41). CONCLUSION: FDX achieves significantly higher global cure rates and lower recurrence rates and is comparable to VCM in clinical cure rates and adverse event rates in patients with CDI. Collectively, FDX is superior to VCM as a therapeutic agent for CDI.

Clinical validation of quantitative SARS-CoV-2 antigen assays to estimate SARS-CoV-2 viral loads in nasopharyngeal swabs.

BACKGROUND: Expansion of the testing capacity for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an important issue to mitigate the pandemic of coronavirus disease-2019 (COVID-19) caused by this virus. Recently, a sensitive quantitative antigen test (SQT), Lumipulse® SARS-CoV-2 Ag, was developed. It is a fully automated chemiluminescent enzyme immunoassay system for SARS-CoV-2. METHODS: In this study, the analytical performance of SQT was examined using clinical specimens from nasopharyngeal swabs using reverse transcription polymerase chain reaction (RT-PCR) as a control. RESULTS: Receiver operating characteristic analysis of 24 SARS-CoV-2-positive and 524 -negative patients showed an area under the curve of 0.957 ± 0.063. Using a cut-off value of 1.34 pg/ml, the sensitivity was 91.7%, the specificity was 98.5%, and the overall rate of agreement was 98.2%. In the distribution of negative cases, the 99.5 percentile value was 1.03 pg/ml. There was a high correlation between the viral load calculated using the cycle threshold value of RT-PCR and the concentration of antigen. The tendency for the antigen concentration to decrease with time after disease onset correlated with that of the viral load. CONCLUSIONS: Presented results indicate that SQT is highly concordant with RT-PCR and should be useful for the diagnosis of COVID-19 in any clinical setting. Therefore, this fully automated kit will contribute to the expansion of the testing capability for SARS-CoV-2.

Immunochromatographic test for the detection of SARS-CoV-2 in saliva.

We evaluated the rapid immunochromatographic test for severe acute respiratory coronavirus 2 (SARS-CoV-2) antigen detection using 16 saliva specimens collected from 6 COVID-19 hospitalized patients, and detected N-antigen in 4 of 7 RT-PCR positive specimens. This POCT detected SARS-CoV-2 antigen in saliva and would be useful for COVID-19 diagnosis.

Streptococcal toxic shock syndrome caused by ß-hemolytic streptococci: Clinical features and cytokine and chemokine analyses of 15 cases.

OBJECTIVES: ß-Hemolytic streptococci occasionally cause severe infections such as necrotizing fasciitis and streptococcal toxic shock syndrome (STSS). Here, we conducted a prospective study to investigate the production of cytokines and chemokines in patients with STSS to explore its pathogenesis in survivors and fatal cases. METHODS: From January 2013 through August 2015, all culture results from normally sterile sites were prospectively followed and screened for STSS. Clinical characteristics of the patients with STSS were evaluated and compared between survivors and fatal cases. Serum samples were collected on admission for quantification of various cytokines and chemokines. Bacterial strains were categorized by Lancefield grouping and analyzed for the emm type, and presence of speA, speB, speC, and speF. RESULTS: Fifteen patients received diagnosis of STSS. The median age of the patients was 60-year-old, and the mortality rate was 40% despite intensive treatment. Nine strains were categorized as group A, two belonged to group G, and four to group B. Group A contained various emm genotypes. Unexpectedly, potent proinflammatory cytokine levels such as TNF-α and IL-1ß were not significantly elevated, and comparison with surviving patients showed that IL-6, IL-8, and MCP-1 levels were significantly decreased and creatine kinase level was significantly elevated in fatally ill cases. CONCLUSION: Our results indicate that reduced production of proinflammatory cytokines and chemokines may be involved in STSS pathogenesis and critical for prognosis of patients with STSS.

Oral vancomycin versus metronidazole for the treatment of Clostridioides difficile infection: Meta-analysis of randomized controlled trials.

At present, vancomycin (VCM) and metronidazole (MNZ) are used for the first-line standard treatment of Clostridioides difficile infection (CDI). However, their differential use has not been sufficiently investigated. In this study, a meta-analysis on differences in the efficacy for CDI between VCM and MNZ was performed. Reports of randomized controlled studies using VCM or MNZ to treat CDI were surveyed. Meta-analysis was performed using the Mantel-Haenszel method and random-effects model, and the risk ratio and 95% confidence interval were calculated. Excluding overlapping reports, 1043 reports were extracted and 5 randomized controlled studies were extracted. There was no difference in therapeutic effects for CDI between VCM and MNZ (RR = 1.08, 95% CI (0.99-1.17), p = 0.09, I2 = 37%). On subgroup analysis by the severity, there was no difference in the clinical effects for CDI between VCM and MNZ in non-severe cases (risk ratio: 1.09, 95% confidence interval: 1.00-1.19, p = 0.06), but the clinical effects of VCM were significantly higher than those of MNZ in severe cases (risk ratio: 1.19, 95% confidence interval: 1.02-1.39, p = 0.03). No significant difference was noted in the recurrence rate, incidence of adverse event, time to exhibit therapeutic effects, or judgment of the bacteriological effects. As the therapeutic effects of VCM were superior in severe CDI cases, VCM should be considered first in severe cases.

Worldwide Lineages of Clinical Pneumococci in a Japanese Teaching Hospital Identified by DiversiLab System.

Pneumococcal Molecular Epidemiology Network (PMEN) clones are representatives of worldwide-spreading pathogens. DiversiLab system, a repetitive PCR system, has been proposed as a less labor-and time-intensive genotyping platform alternative to conventional methods. However, the utility and analysis parameters of DiversiLab for identifying worldwide lineages was not established. To evaluate and optimize the performance of DiversiLab for identifying worldwide pneumococcal lineages, we examined 245 consecutive isolates of clinical Streptococcus pneumoniae from all age-group patients at a teaching hospital in Japan. The capsular swelling reaction of all isolates yielded 24 different serotypes. Intensive visual observation (VO) of DiversiLab band pattern difference divided all isolates into 73 clusters. Multilocus sequence typing (MLST) of representative 73 isolates from each VO cluster yielded 51 different STs. Among them, PMEN-related lineages accounted for 63% (46/73). Although the serotype of PMEN-related isolates was identical to that of the original PMEN clone in 70% (32/46), CC156-related PMEN lineages, namely Greece(6B)-22 and Colombia(23F)-26, harbored various capsular types discordant to the original PMEN clones. Regarding automated analysis, genotyping by extended Jaccard (XJ) with a 75% similarity index cutoff (SIC) showed the highest correlation with serotyping (adjusted Rand's coefficient, 0.528). Elevating the SIC for XJ to 85% increased the discriminatory power sufficient for distinguishing two major PMEN-related isolates of Taiwan(19F)-14 and Netherlands(3)-31. These results demonstrated a potential utility of DiversiLab for identifying worldwide lineage of pneumococcus. An optimized parameters of automated analysis should be useful especially for comparison for reference strains by "identification" function of DiversiLab.

Incorrect diagnosis of Clostridium difficile infection in a university hospital in Japan.

Physicians often fail to suspect Clostridium difficile infection (CDI) and many microbiology laboratories use suboptimal diagnostic techniques. To estimate the extent of and reasons for incorrect diagnosis of CDI in Japan, we investigated toxigenic C. difficile isolated from all stool culture samples and clinical course. Over a 12-month period in 2010, all stool culture samples (n = 975) submitted from inpatients in a university hospital in Japan were cultured for C. difficile and routine microbiological testing was conducted. In total, 177 C. difficile isolates were recovered, and 127 isolates were toxigenic. Among the toxin-A-positive/toxin-B-positive isolates, 12 were also positive for the binary toxin gene. However, clinically important ribotypes, such as 027 and 078, were not identified. A total of 58 (45.7%) cases with toxigenic C. difficile had unformed stool, and the incidence CDI was 1.6 cases per 10,000 patient-days. Of these 58 cases, 40 were not diagnosed in routine testing due to a lack of clinical suspicion (24.1%, 14/58) or a negative C. difficile toxin assay result (44.8%, 26/58). A stool toxin assay was performed in 54 patients (78.2%, 54/69) who did not have unformed stool. The present study demonstrated that a significant number of CDI cases in Japan might be overlooked or misdiagnosed in clinical practice due to a lack of clinical suspicion and limitations of microbiological testing for CDI in Japan. Providing education to promote awareness of CDI among physicians is important to improve the accuracy of diagnosis in Japan.

[Usefulness of Immunochromatography Method for Malaria and Dengue Fever Diagnosis].

There are three major differential diagnosis of febrile patients with history of travels to the tropical countries i.e., malaria, typhoid fever and dengue fever. Diagnosis of malaria patients undergoes sometimes arduous process due to the variable skills of laboratory technician, and more convenient method is warranted. Immunochromatography (IC) method is simple method and recently used for diagnosis of several infectious diseases. Here, we reported usefulness of IC method for malaria and dengue fever diagnosis. Forty-seven samples from 46 patients were retrospectively analyzed by both malaria IC method and microscopic examination. Furthermore, three patients were undergone dengue IC method followed by PCR and antibody examination (ELISA) if the results were positive. Several factors such as rheumatoid factor (RF) are known to affect the results of IC method. We also checked malaria and dengue IC method using serum known to be high RF results without malaria infection. Totally six patients were diagnosed as malaria i.e., 1 vivax malaria and 5 falciparum malaria. Sensitivity and specificity of the malaria IC method were excellent, 100% and 97.6%, respectively. Among three patients, one patient revealed false-negative results of dengue IC method, however, results of the other two patients revealed good correlation between IC method and PCR/ELISA results. Among four RF positive serums, 2 malaria IC method and 4 dengue IC method revealed false-positive results. In summary, IC method for malaria and dengue fever might be quick and convenient method and considered to be used as an adjunctive diagnostic tool.

Clinical Efficacy of Therapeutic Agents for Clostridioides difficile Infection Based on Four Severity Classifications.

Japanese guidelines recommend metronidazole (MNZ) and vancomycin (VCM) for non-severe and severe cases of Clostridioides difficile infection (CDI), respectively. In the present study, we investigated the use of CDI antimicrobials and evaluated their clinical efficacy and validity using four severity classifications. A retrospective chart review was conducted using the data of 137 inpatients with initially positive C. difficile toxin test results and the initiation of CDI antimicrobials between April 2015 and March 2019. Patients treated with VCM or oral MNZ were included for clinical efficacy analysis of CDI antimicrobials and validation of severity classifications. The endpoints were CDI recurrence, 30-day mortality, and diarrhea cure rates. No significant differences were found between the VCM and oral MNZ groups in the CDI recurrence rate (10.4% vs. 12.7%, P = 0.707), 30-day mortality rate (12.5% vs. 5.6%, P = 0.162), and diarrhea cure rate (61.9% vs. 72.7%, P = 0.238), regardless of severity. Treatment with oral MNZ for non-severe cases was promising, confirming its usefulness according to Japanese guidelines. Further investigation of the clinical efficacy of oral MNZ in patients with first-episode CDI and evaluation of the preferred severity classification are warranted.

The outbreak of multispecies carbapenemase-producing Enterobacterales associated with pediatric ward sinks: IncM1 plasmids act as vehicles for cross-species transmission.

BACKGROUND: This study describes an outbreak caused by multispecies carbapenemase-producing Enterobacterales (CPE) occurring in a pediatric ward at an academic medical center in Tokyo. METHODS: The index case involved a 1-year-old boy with Klebsiella variicola (CPE) detected in anal swabs in June 2016. The second case was Klebsiella quasipneumoniae (CPE) occurred in March 2017 followed by further spread, leading to the declaration of an outbreak in April 2017. Extensive environmental and patient microbiological sampling was performed. The relatedness of the isolates was determined using draft-whole-genome sequencing. RESULTS: CPE surveillance cultures of patients and environments were positive in 19 patients and 9 sinks in the ward. The sinks in hospital rooms uninhabited by CPE patients exhibited no positive CPE-positive specimen during the outbreak. All CPE strains analyzed using draft-whole-genome sequencing harbored blaIMP-1, except for one harboring blaIMP-11; these strains harbored identical blaIMP-1-carrying IncM1 plasmids. CPE was detected even after sink replacement; infection-control measures focused on sinks were implemented and the CPE outbreak ended after 7 months. CONCLUSIONS: Multiple bacterial species can become CPE via blaIMP-1-carrying IncM1 plasmids of the same origin and spread through sinks in a hospital ward. Thorough infection-control measures implemented as a bundle might be crucial.

Molecular characterization of extraintestinal Escherichia coli isolates in Japan: relationship between sequence types and mutation patterns of quinolone resistance-determining regions analyzed by pyrosequencing.

Infection from fluoroquinolone-resistant Enterobacteriaceae is an increasing health problem worldwide. In the present study, we developed a pyrosequencing-based high-throughput method for analyzing the nucleotide sequence of the quinolone resistance-determining regions (QRDRs) of gyrA and parC. By using this method, we successfully determined the QRDR sequences of 139 out of 140 clinical Escherichia coli isolates, 28% of which were nonsusceptible to ciprofloxacin. Sequence results obtained by the pyrosequencing method were in complete agreement with those obtained by the Sanger method. All fluoroquinolone-resistant isolates (n = 35; 25%) contained mutations leading to three or four amino acid substitutions in the QRDRs. In contrast, all isolates lacking a mutation in the QRDR (n = 81; 57%) were susceptible to ciprofloxacin, levofloxacin, and nalidixic acid. The qnr determinants, namely, the qnrA, qnrB, and qnrS genes, were not detected in the isolates, and the aac(6')-Ib-cr gene was detected in 2 (1.4%) of the isolates. Multilocus sequence typing of 34 randomly selected isolates revealed that sequence type 131 (ST131) (n = 7; 20%) is the most prevalent lineage and is significantly resistant to quinolones (P < 0.01). The genetic background of quinolone-susceptible isolates seemed more diverse, and interestingly, neighboring STs of ST131 in the phylogenetic tree were all susceptible to ciprofloxacin. In conclusion, our investigation reveals the relationship between fluoroquinolone resistance caused by mutations of QRDRs and the population structure of clinical extraintestinal E. coli isolates. This high-throughput method for analyzing QRDR mutations by pyrosequencing is a powerful tool for epidemiological studies of fluoroquinolone resistance in bacteria.

Diagnostic usefulness of ribosomal protein l7/l12 for pneumococcal pneumonia in a mouse model.

The capsular antigen detection (CAD) kit is widely used in clinics to detect Streptococcus pneumoniae infection from urine, because it is rapid, convenient, and effective. However, there are several disadvantages, including false-positive results in children colonized with S. pneumoniae and prolonged positive readings even after the bacteria have been cleared. RP-L7/L12 is a component of the 50S ribosome that is abundant in all bacteria and is specific for each bacterial species. We investigated whether RP-L7/L12 could be used to accurately diagnose pneumococcal pneumonia infection in mouse models of pneumonia and colonization generated by infecting CBA/JN or CBA/N mice, respectively, with S. pneumoniae strain 741. RP-L7/L12 detection by enzyme-linked immunosorbent assay accurately assessed active lung infection, as RP-L7/L12 levels decreased simultaneously with the bacterial lung burden after imipenem administration in the pneumonia mouse model. Based on the data, antibodies detecting RP-L7/L12 were applied to rapid immunochromatographic strips (ICS) for urine sample testing. When we compared the ICS test with the CAD kit in the pneumonia model, the results correlated well. Interestingly, however, when the lung bacterial burden became undetectable after antibiotic treatment, the ICS test was correspondingly negative, even though the same samples tested by the CAD kit remained positive. Similarly, while the ICS test exhibited negative results in the nasal colonization model, the CAD kit demonstrated positive results. Bacterial RP-L7/L12 may be a promising target for the development of new methods to diagnose infectious disease. Further studies are warranted to determine whether such a test could be useful in children.

Protective effect of procysteine on Acinetobacter pneumonia in hyperoxic conditions.

OBJECTIVES: Ventilator-associated pneumonia (VAP) is an important cause of morbidity and mortality in critical care settings. Acinetobacter has become a leading cause of VAP. In particular, the appearance and spread of multidrug-resistant Acinetobacter is of great concern. In this study, we examined the effect of the antioxidant procysteine on Acinetobacter murine pneumonia in hyperoxic conditions in order to simulate VAP. METHODS: Acinetobacter was administered intranasally to BALB/c mice kept in hyperoxic conditions. At designated timepoints, bacterial number, cytokine production and histopathological findings in the lungs were examined. The effects of procysteine on survival rates, lung bacterial burdens and the phagocytic activities of alveolar macrophages were evaluated. RESULTS: Drastic decreases in survival were observed when the infected mice were kept in hyperoxic conditions (P < 0.001). Significant differences in pulmonary bacterial number and neutrophil accumulation were observed between mice kept in hyperoxic or normoxic conditions on day 3. Although all mice infected with Acinetobacter spp. and kept in hyperoxic conditions died by day 3, procysteine treatment significantly improved survival (60% survival on day 7, P < 0.01). Procysteine treatment decreased the lung bacterial burden on days 2 and 3. Finally, improved uptake of FITC-labelled beads by alveolar macrophages from mice treated with procysteine and kept in hyperoxic conditions was noted. CONCLUSIONS: These results suggest that hyperoxia increases mortality in mice with Acinetobacter pneumonia and that procysteine improves survival by increasing the phagocytic activity of alveolar macrophages in mice kept in hyperoxic conditions.

Virulence-suppressing effects of linezolid on methicillin-resistant Staphylococcus aureus: possible contribution to early defervescence.

In the present study, immunomodulatory effects of linezolid (LZD) on methicillin-resistance Staphylococcus aureus (MRSA) infections were evaluated. We have retrospectively reviewed treatment effects of LZD on 52 patients with severe MRSA infections. Sixty-four percent of the febrile patients demonstrated significant defervescence within 3 days, despite the presence of positive culture results. We speculated that this finding might be due to early anti-inflammatory effects of LZD, and to investigate this further we initiated in vivo experiments using mice MRSA pneumonia models. Mice were treated with either LZD or vancomycin (VCM) immediately after intranasal administration of MRSA. Bacterial numbers and levels of inflammatory cytokines in the lungs were determined. Although the bacterial burden in the lungs was not apparently different between the two groups, LZD but not VCM treatment significantly reduced induction of inflammatory cytokines in the lungs (P < 0.05). To evaluate whether this anti-inflammatory response was due to suppression of virulence factor expression, filter-sterilized supernatants of MRSA incubated in broth overnight with sub-MICs of LZD were subcutaneously administered to mice. To clarify whether LZD possesses direct host-modulating activity, cytokine responses to the supernatants were examined in mice pretreated with LZD. Interestingly, MRSA solutions prepared in the presence of sub-MICs of LZD revealed significant suppression of interleukin 6 (IL-6) in a dose-dependent manner (P < 0.05), but pretreatment of mice with LZD revealed no changes in cytokines. These findings suggest that sub-MICs of LZD might suppress virulence factors of MRSA, which may be associated with a reduction in endogenous pyrogens. These data may explain at least in part early defervescence observed in LZD-treated individuals.

Genotyping of skin and soft tissue infection (SSTI)-associated methicillin-resistant Staphylococcus aureus (MRSA) strains among outpatients in a teaching hospital in Japan: application of a phage-open reading frame typing (POT) kit.

We aimed to elucidate the current epidemiological features of outpatient skin and soft tissue infection (SSTI)-associated methicillin-resistant Staphylococcus aureus (MRSA) in Japan. Altogether, we evaluated the performance of a phage-open reading frame typing (POT) kit for genotyping these MRSA strains. We collected 57 MRSA strains from all outpatients with SSTIs attending a teaching hospital in Japan. Drug susceptibility measurement and genotyping including SCCmec typing, spa typing, multilocus sequence typing, pulsed-field gel electrophoresis, and commercial POT-kit were performed. The majority of strains (39 strains, 68 %) had the SCCmec-II element. Seventeen strains (30 %) with SCCmec-IV accounted for the second largest population. Strains with SCCmec-IV and SCCmec-V appeared multiclonal, and a predominance of Panton-Valentine leukocidin (PVL) gene-negative CC8/spa-CC008 strains, as well as the first isolate of an ST93 strain in Japan, was observed among them. Only one USA300 strain was identified. Strains with SCCmec-IV and SCCmec-V were significantly susceptible to antimicrobials. The PVL gene was found in 5 SCCmec-IV strains and 1 SCCmec-V strain. The POT-kit successfully predicted the SCCmec type in 54 strains (95 %), and typing by POT1 scores was highly concordant with SCCmec typing and spa typing. Moreover, three PVL-positive strains fell into a particular POT type (POT scores, 106-77-113). Simpson's index of the POT-kit was 0.977. In conclusion, the present study clarified the multiclonal nature of outpatient SSTI-associated MRSA in a teaching hospital in Japan. These data also underscore the utility of the POT-kit for non-outbreak surveillance through its simple platform consisting of two multiplex PCRs without sequencing.

[Isolation rate and susceptibilities of candida species from blood, vascular catheter, urine and stool].

We evaluated species distribution and antifungal susceptibility of Candida isolates during 2002-2008. Of 177 Candida isolates from blood, species distribution was 90 (51%) Candida albicans, 30 (17%) C. parapsilosis, 22 (12%) C. glabrata, 6 (3%) C. tropicalis and 29 (16%) other Candida spp.. Of 162 Candida isolates from vascular catheter, species distribution was 87 (54%) C. albicans, 14 (9%) C. parapsilosis, 36 (22%) C. glabrata, 5 (3%), C. tropicalis, 2 (1%) C. krusei and 18 (11%) other Candida spp.. Of 1889 Candida isolates from urine, species distribution was 1165 (62%) C. albicans, 22 (1%) C. parapsilosis, 484 (26%) C. glabrata, 83 (4%) C. tropicalis, 26 (1%) C. krusei and 109 (6%) other Candida spp.. Of 782 Candida isolates from stool, species distribution was 425 (54%) C. albicans, 3 (1%) C. parapsilosis, 103 (13%) C. glabrata, 28 (4%) C. tropicalis, 5 (1%), C. krusei and 218 (28%) other Candida spp. Both C. albicans and non-Candida spp. isolated from urine increased slightly over the past 7 years. Flucytosine, fluconazole, itraconazole and micafungin still have strong activity against Candida isolates.