細田 卓也

With the widespread use of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS), the number of identifiable bacterial species has increased. However, anaerobic bacteremia remains challenging to accurately diagnose due to the diverse range of anaerobic bacteria and the frequent occurrence of polymicrobial infections. Consequently, MALDI-TOF MS often fails to achieve accurate species-level identification in such cases. To address this limitation, we evaluated whole-genome sequencing (WGS) as an alternative method for identifying anaerobic bacteria in blood cultures. Over a 4-year period (April 2020 to March 2024), 69 cases of anaerobic bacteremia were identified, involving 85 bacterial strains. WGS successfully identified 73 strains (89%) at the species level. MALDI-TOF MS accurately identified 43 strains (59%) at the species level and 6 strains (8.2%) at the genus level. Among the 24 discordant strains, 9 species were not included in the database, and 6 species had limited prior reports of bloodstream infections. Additionally, 21 of the 69 cases (30%) were polymicrobial, and WGS revealed 9 cases (13%) in which multiple species had not been identified by MALDI-TOF MS. These results highlight the limitations of MALDI-TOF MS in anaerobic bacterial identification, particularly in polymicrobial infections, and suggest that alternative molecular approaches are necessary to improve diagnostic accuracy.IMPORTANCEAccurate identification of anaerobic bacteria remains a significant challenge despite the widespread use of matrix-assisted laser desorption ionization time-of-flight mass spectrometry. While this technology has improved the detection of many bacterial species, some anaerobes remain unidentified due to their absence from reference databases and the difficulties associated with their isolation, particularly in polymicrobial infections. Whole-genome sequencing (WGS) has revealed previously unreported anaerobes and identified polymicrobial infections that were initially misclassified as monomicrobial. Our findings underscore the importance of implementing molecular approaches such as WGS- or PCR-based methods in clinical diagnostics to improve the detection of anaerobic pathogens.

Rapid detection and reporting of carbapenemase-producing Enterobacterales (CPE) is one of the top priorities of clinical microbiology laboratories. The Clinical and Laboratory Standards Institute recommends the modified carbapenem inactivation method (mCIM) as the preferred method for this purpose, but it requires a broth incubation process which can be cumbersome. Here, we compared the performance of mCIM with three alternative rapid CPE detection methods against a collection of genetically defined CPE, with most carrying blaIMP, and non-CPE clinical isolates. The sensitivities of mCIM, simplified carbapenem inactivation method (sCIM), Rapidec Carba NP, and NG-Test Carba 5 were 98.0%, 54.9%, 90.2%, and 72.5%, whereas the specificities were 89.5%, 84.2%, 89.5%, and 100%, respectively. Modification of the interpretive criteria of sCIM increased its sensitivity to 88.2% and specificity to 89.5%. The results suggest that mCIM is currently the optimal method for CPE detection in an epidemiological setting where CPE-producing IMP group carbapenemase is predominant. While sCIM is easier to perform, it requires further validation before it can be widely adopted as an alternative to mCIM in the clinical laboratory. IMPORTANCE Simple identification methods for carbapenemase-producing Enterobacterales are required for the clinical laboratory. The simplified carbapenem inactivation method (sCIM) is a carbapenemase detection method that can be performed with less hands-on time than mCIM, but its sensitivity and specificity were suboptimal compared with other phenotypic detection methods when tested against a collection of IMP-producing CPE. Insufficient inactivation of imipenem from inadequate inoculation was suspected as the cause. While sCIM is easier to perform, it requires optimization before it can be widely adopted as an alternative to mCIM in the clinical laboratory.

Carbapenemase-producing Escherichia coli sequence type (ST) 648 strains were isolated from two international visitors without previous medical exposure from Southeast Asian countries in a hospital in Japan. One isolate, FUJ80154, carried blaNDM-5 in a complex class 1 integron on an IncFIB/FII plasmid; the other isolate, FUJ80155, carried two copies of blaOXA-48 on the chromosome flanked by IS1R on both sides. The core-genome based-phylogenetic analysis with publicly available genome data of E. coli ST648 carrying blaNDM-5 or blaOXA-48-like demonstrated high genetic similarity between FUJ80154 and NDM-5-prooducing E. coli ST648 strains isolated in South and Southeast Asian countries. On the other hand, no closely related isolates of FUJ80155 were identified. In the absence of prior hospitalization overseas, neither patient had qualified for routine screening of multidrug-resistant organisms, and the isolates were incidentally identified in cultures ordered at the discretion of the treating physician. IMPORTANCE Although patients with history of international hospitalization are often subject to screening for multidrug-resistant organisms, it is unclear whether patients who reside in countries where carbapenemase-producing Enterobacterales (CPE) is endemic but have no history of local hospitalization contribute to the transmission of CPE. In this study, NDM-5-producing and OXA-48-producing Escherichia coli sequence type (ST) 648, a recently recognized high-risk, multidrug-resistant clone, were detected from two overseas visitors without previous medical exposure. The findings of this study suggest that active surveillance culture on admission to hospital may be considered for travelers from countries with endemicity of carbapenem-resistant organisms even without history of local hospitalization and underscore the need to monitor cross-border transmission of high-risk clones, such as carbapenemase-producing E. coli ST648.