Journal of Clinical Microbiology最新文献

Matrix-assisted laser desorption-ionization-time of flight (MALDI-TOF) mass spectra can be used to predict antimicrobial resistance (AMR) using machine learning (ML). This study aimed to validate the performance of ML models for AMR prediction using own and publicly available MALDI-TOF data and to test how these models perform over time. Mass spectra of Escherichia coli (n = 7,897), Klebsiella pneumoniae (n = 2,444), and Staphylococcus aureus (n = 4,664) from routine diagnostics (Germany) and the DRIAMS-A database (Switzerland) were used. Six classification models were benchmarked for AMR prediction using cross-validation (regularized logistic regressions [LR], multilayer perceptrons [MLP], support vector machines [SVM], random forests [RF], gradient boosting machines [LGBM, XGB]). Performance was prospectively observed for 18 months after training. The performance of AMR prediction evaluated by the mean area under the receiver operating characteristic curve (AUROC) was comparable between the DRIAMS-A data set and own data. The best predictive performance (classifier, AUROC) on own data was achieved for oxacillin resistance in S. aureus (RF, 0.85), ciprofloxacin resistance in E. coli (XGB, 0.83), and piperacillin-tazobactam resistance in K. pneumoniae (XGB, 0.81). ML performance was poor if training and test data were unrelated in terms of location and time. Performance (change in AUROC) decreased within 18 months after training for S. aureus (oxacillin resistance, RF: -0.10), E. coli (ciprofloxacin, XGB: -0.19), and K. pneumoniae (piperacillin-tazobactam, XGB: -0.25). The performance of ML for the prediction of AMR based on MALDI-TOF data is good (AUROC ≥ 0.8) but classifiers need to be trained on local data and retrained regularly to maintain the performance level.

Importance: MALDI-TOF mass spectrometry can be used not only for bacterial species identification but also for the prediction of antimicrobial resistance (AMR) using machine learning (ML). Such an approach would provide antimicrobial susceptibility test results one day earlier than conventional routine diagnostics. This is essential for an early targeted treatment to reduce mortality of severe infections. We show that the performance of ML for the prediction of AMR based on MALDI-TOF data is good (AUROC ≥ 0.8). However, the ML models need to be trained on local data and retrained regularly to maintain a good performance.

The resurgence of whooping cough in regions utilizing acellular pertussis vaccines underscores emerging public health challenges. Here, we characterized 178 Bordetella pertussis isolates collected from patients across all age groups in Shanghai (2018-2024) to assess genomic evolution and antibiotic susceptibility. Macrolide resistance to erythromycin, azithromycin, and clarithromycin escalated from ≤50% (pre-2020) to nearly 100% (post-2020), mechanistically linked to the 23S rRNA A2047G mutation. Genome-based analysis identified a genotype MT28-ptxP3-MRBP rapidly dominated post-2020, exhibiting significantly higher prevalence in adults than in other age groups. Phylogenetic analysis of 178 Shanghai and 1,596 global genomes revealed two major lineages corresponding to ptxP1 and ptxP3 alleles. MT28-ptxP3-MRBP cluster was identified in France, Japan, and the United States in 2024, indicating potential cross-border transmission. These findings advocate for integrated surveillance spanning all ages and international borders to contain the global spread of macrolide-resistant B. pertussis.

Importance: In recent years, despite high coverage of acellular pertussis vaccines in China, pertussis cases have increased substantially. Drawing on 178 Bordetella pertussis isolates obtained through age-inclusive active surveillance in Shanghai (2018-2024), we found that macrolide resistance rose from ≤50% before 2020 to nearly 100% thereafter, with all resistant isolates harboring the 23S rRNA A2047G mutation. A resistant MT28-ptxP3 lineage became dominant after 2020 (61.7%) and was disproportionately represented among older age groups; the primary affected population shifted from children ≤36 months toward those aged 37 months to 18 years. Incorporating NCBI public genome data, we further observed that this resistant lineage is not confined locally, suggesting a risk of cross-border spread. These findings provide an early warning of the expansion of macrolide-resistant pertussis and underscore the need for age-inclusive, cross-regional genomic surveillance and re-evaluation of diagnostic workflows, antimicrobial stewardship, and immunization strategies.

Diagnosing pulmonary tuberculosis (TB) and monitoring treatment remain challenging. New tools such as the PATHFAST TB LAM assay (PHC Corporation, formerly LSI Medience Corporation, Tokyo, Japan; distributed by BioSynex, France; PF-LAM) may complement microscopy, culture, and NAAT. PF-LAM is an immunoassay quantifying lipoarabinomannan (LAM), a mycobacterial cell wall component, in sputum within 1 hour using an automated chemiluminescent reader. Diagnostic performance was first assessed on 100 sputum samples: 40 culture-positive for Mycobacterium tuberculosis complex (MTBC), 40 culture-positive for nontuberculous mycobacteria (NTM), and 20 culture-negative for both. We then tested 61 longitudinal sputum samples from 19 pulmonary TB patients under treatment. Four additional samples from one patient with NTM pulmonary disease (NTM-PD) were also tested. PF-LAM showed 75% sensitivity for MTBC detection, with a strong correlation between LAM concentration and culture time-to-positivity (Spearman ρ = 0.915, P < 0.0001). Specificity was 95% on culture-negative specimens. The test was also positive for 26/40 (65%) NTM-culture-positive samples. Notably, 19 of these 26 samples were obtained from NTM-PD patients, yielding 73% sensitivity for NTM-PD detection. The assay's potential for treatment monitoring was demonstrated by significant negative correlations between LAM concentration and (a) treatment duration (ρ=-0.434, P = 0.0012) and (b) time to positive culture (ρ=-0.665, P < 0.0001). PF-LAM is a rapid and easy-to-use test for diagnosing pulmonary TB. While NTM cross-reactivity reduces specificity, it may provide diagnostic value for NTM-PD. Results on monitoring TB treatment are promising as no test is currently available for this indication.

Importance: Effective monitoring of the treatment response is essential for successful tuberculosis (TB) management as prolonged therapies require accurate evaluation to prevent relapse, treatment failure, or drug resistance. This study highlights the diagnostic and treatment-monitoring potential of the PATHFAST TB LAM Ag assay, which quantifies lipoarabinomannan (LAM) concentrations in sputum samples and correlates with bacterial load. For the first time, we also demonstrate the assay's applicability for detecting and monitoring nontuberculous mycobacterial (NTM) pulmonary diseases, which are increasingly prevalent in industrialized countries. The semi-automated, rapid format (<17 minutes) of the PATHFAST TB LAM Ag assay provides a simple and reliable approach for assessing bacillary load during treatment, representing a promising tool for improving patient management and diagnostic efficiency in both TB and NTM-PD.

Sexually transmitted infections, e.g., Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), and Mycoplasma genitalium (MG), present with similar signs/symptoms and are often treated empirically due to delays in receiving diagnostic test results. This study evaluated the clinical performance of the Cobas Liat CT/NG/MG nucleic acid amplification test (NAAT) in detecting these pathogens in a point-of-care (POC) setting. This non-interventional, multicenter study recruited symptomatic/asymptomatic participants from 13 geographically diverse POC sites across the USA. Prospective clinician- and self-collected specimens (urine/vaginal swabs) were used to determine the sensitivity and specificity of the Cobas Liat CT/NG/MG test relative to the composite reference standard (CRS), determined using a combination of results from three FDA-approved NAATs and one laboratory-developed test. Among 4,800 evaluable participants, 40.4% (n = 1,941) were symptomatic, and 51.9% (n = 2,489) were female at birth. The Cobas Liat CT/NG/MG test demonstrated good clinical performance across all specimen types regardless of symptom status, with specificity >97% for each analyte, and sensitivity ≥92%, except female urine (≥CT 87%, NG ≥ 83%, MG ≥ 77%). The test was considered easy to use, and no statistically significant difference in performance results was observed between trained/untrained users. The Cobas Liat CT/NG/MG test demonstrated good clinical performance, with high sensitivity and specificity for CT/NG/MG detection, regardless of symptom status. The assay provides a short turn-around time (approximately 20 min) with centralized testing laboratory accuracy at the POC for self- and clinician-collected samples. POC testing can facilitate rapid pathogen identification and accurate treatment, thereby mitigating the need for empiric treatment.IMPORTANCENucleic acid amplification tests are the preferred method for diagnosing CT and NG infections and are the only reliable tests for MG; however, delays in receiving results can lead to empiric treatment, potentially causing misdiagnosis and overtreatment of STIs. Point-of-care testing could mitigate these issues by enabling rapid pathogen identification and treatment, but no rapid POC tests are currently available for the detection of CT, NG, and MG despite the similarity of symptoms, and these three pathogens being responsible for the majority of symptomatic STIs in some settings. Our findings suggest that the Cobas Liat CT/NG/MG assay may help to reduce reliance on empiric treatment of symptoms and minimize resulting return visits for unresolved infections. Use of the Cobas Liat CT/NG/MG assay may also result in improved patient outcomes, help realize population benefits by reducing the duration of infection and potentially transmission, and reduce healthcare costs.

Trypanosoma brucei infections cause African trypanosomiasis, also known as sleeping sickness in humans and nagana in animals, presenting a significant global health and economic burden, especially in sub-Saharan Africa. Animal trypanosomiasis also affects the economic development in Asia and South America. Accurate diagnosis of diseases caused by infection with the parasite of the T. brucei group remains a major challenge due to the persistence of infection-induced antibodies long after parasite clearance, complicating serological discrimination between active and past infections. To address this limitation, we developed a sensitive and specific antigen detection assay targeting Trypanosoma brucei enolase (TbrENO) using a panel of camelid single-domain antibodies (sdAbs also known as nanobodies). Among the candidates, the sdAbsR3-77/sdAbR2-103 sandwich enzyme-linked immunosorbent assay (ELISA) exhibited robust performance in detecting circulating TbrENO in plasma from experimentally infected mice. Additionally, this assay showed strong potential as a "test-of-cure" tool by monitoring real-time antigenemia throughout a chronic T. brucei infection course. We further validated the assay's diagnostic utility in human clinical samples, detecting Trypanosoma brucei rhodesiense infections at both early and advanced stages and Trypanosoma brucei gambiense infections in advanced stage. The sdAbsR3-77H/sdAbR2-103HA ELISA targeting TbrENO shows potential for point-of-care pan-diagnosis of active T. brucei infections (including Trypanosoma brucei brucei, T. b. gambiense, T. b. rhodesiense, Trypanosoma brucei evansi, and Trypanosoma brucei equiperdum) in both animals and humans. Therefore, this assay addresses gaps in current diagnostic capabilities by overcoming the key limitations of antibody-based tests, offering a promising tool for improved disease control.IMPORTANCEAfrican trypanosomiasis, commonly known as sleeping sickness in humans and nagana in animals, is a life-threatening disease that remains a major health and economic concern in many parts of the world. One of the key difficulties in managing this disease is detecting ongoing infections, as existing antibody-based tests cannot reliably distinguish between current and past infections. In this study, we developed a novel laboratory test that detects a specific protein released by the parasite during infection. This test uses special antibodies derived from camels, known for their exceptional stability and precision, to accurately identify infections caused by multiple Trypanosoma brucei subspecies. Our approach not only enables accurate diagnosis but also offers a way to monitor treatment success. This work provides a valuable tool for disease control efforts and could help improve the health of both humans and animals in regions where trypanosomiasis is endemic.

Leptospirosis, caused by Leptospira spp. infection, is a globally significant zoonotic disease that affects a wide range of animals. Although renal colonization is well-documented, genital infection by leptospires remained less explored for decades, despite its impact on reproduction. Evidence suggests that genital infection occurs as a primary condition rather than secondary to renal colonization, particularly in cattle suffering from bovine genital leptospirosis (BGL), linked to chronic infections by strains of the Sejroe serogroup. In horses, a similar condition is suggested to be associated with strains of serogroup Australis. Molecular studies confirmed the presence of Leptospira DNA in uterine, follicular, and vaginal samples, strengthening the hypothesis of an independent genital physiopathology. Despite significant advances in molecular diagnostics, the detection of genital carriers remains challenging, requiring refined methodologies beyond standard serology. This review critically examines the historical detection of Leptospira spp. in genital samples of cattle, small ruminants, swine, and equines, emphasizing its relevance to reproductive health. Moreover, we highlight the limitations of current diagnostic approaches, advocating for increased use of genital samples in leptospirosis research of large animals and shedding light on future directions regarding genital leptospirosis in livestock research. Enhanced understanding and diagnosis of genital leptospirosis will contribute to better livestock reproductive management and disease prevention.

The diagnosis of congenital toxoplasmosis or disseminated toxoplasmosis in immunocompromised patients nowadays relies on molecular tools, in particular real-time PCR. There are many reagents on the market, and their evaluation by independent experts provides valuable information to medical biologists who are looking for a high-performance kit among the different references. Under the aegis of the French National Reference Center for Toxoplasmosis, we report here a multicenter evaluation of the analytical and clinical performances of the "quanty TOXO (RH region)" PCR assay manufactured by Clonit. The kit showed good analytical performance, as indicated by the results of serial dilution tests and external quality control samples. PCR efficiencies varied from 95% to 105%; linearity zone extended over four log units (R² >0.99), and limit of detection varied from <1 parasite/mL to between 1 and 5 parasites/mL, i.e., from <0.08 parasite/PCR to between 0.2 and 1 parasite/PCR, depending on the center. Based on 141 cryopreserved DNAs from a large range of clinical specimens, we determined a clinical sensitivity of 94.7% (71/75; 95% confidence interval [CI]: 87.1%-97.9%) and a clinical specificity of 100% (66/66; 95% CI: 94.5%-100%). Four false negative results were detected despite amplification carried out in duplicate. Overall, the "quanty TOXO (RH region)" PCR assay demonstrated satisfactory analytical and clinical performances for the diagnosis of toxoplasmosis, even using extraction and amplification techniques or biological matrices not validated by the manufacturer.IMPORTANCEDue to its speed and accuracy, PCR is now the gold standard for diagnosing congenital and disseminated toxoplasmosis. High-performance molecular testing is essential, especially for immunocompromised patients and congenital infections, to initiate early treatment. This diagnostic approach increasingly relies on commercial assays. However, commercially available kits do not guarantee performance. In this study, conducted by the French National Reference Center for Toxoplasmosis, we performed an independent multicenter evaluation of the "quanty TOXO (RH region)" PCR assay manufactured by Clonit. Our results showed that this kit delivered satisfactory results for routine diagnostic use. However, among the 141 clinical samples tested, four false negative results were noted, corresponding to specimens with low parasitic load.