Journal of Clinical Microbiology最新文献

The aim was to develop an RT-qPCR targeting Aspergillus fumigatus and compare its performance to that of Aspergillus fumigatus qPCR for the diagnosis of invasive aspergillosis (IA). Samples from patients of the Lyon University hospitals for whom a suspicion of IA led to the realization of an Aspergillus fumigatus qPCR molecular diagnostic test over a 2-year period were included. The patients were classified according to the European Organization for Research and Treatment of Cancer/Mycoses Study Group (EORTC-MSGERC) criteria for suspected IA; RT-qPCR and qPCR assays were performed on all included samples. The sensitivities and specificities of RT-qPCR and qPCR were calculated and compared using the results of the EORTC-MSGERC classification as reference. The cycle threshold (Ct) results were compared according to IA classification and sample type. Among the 193 samples analyzed, 91 were classified as IA excluded, 46 as possible IA, 53 as probable IA, and 3 as proven IA. For all sample types, RT-qPCR was significantly more sensitive than qPCR for all IA classifications with an additional 17/102 samples detected (P-value < 0.01). For plasma samples, sensitivity was significantly higher and specificity significantly lower using RT-qPCR for all IA classifications (P-value < 0.001). The mean Ct obtained with RT-qPCR were significantly lower than those obtained with qPCR for all IA classifications and all sample types (P-value < 0.001 and P-value < 0.0001, respectively). RT-qPCR presents a higher sensitivity than qPCR for the diagnosis of IA due to Aspergillus fumigatus, particularly in samples with an intrinsically low fungal load.IMPORTANCEAspergillus fumigatus belongs to the critical priority group of the World Health Organization fungal priority pathogens list. Invasive aspergillosis (IA) is a life-threatening infection with poor prognosis and challenging diagnosis. PCR has been integrated into the 2020 European Organization for Research and Treatment of Cancer/Mycoses Study Group consensus definitions for IA diagnosis. However, due to frequent low fungal burdens, its sensitivity needs to be improved. This work presents an innovative method for detecting total nucleic acids, corresponding to both ribosomal RNA and DNA, that enables IA diagnosis with greater sensitivity than conventional techniques, especially in non-invasive samples such as blood, enhancing the monitoring of this infection in high-risk patients.

Non-sputum tests are needed to improve tuberculosis (TB) diagnosis and close the diagnostic gap. The World Health Organization's target product profile (TPP) for point-of-care (POC) screening tests requires a minimum sensitivity of 90% and a specificity of 70%. Our objective was to identify host blood protein biomarkers meeting TPP criteria. A systematic review was conducted and reported following PRISMA guidelines. Data extraction and quality assessment with Quality Assessment of Diagnostic Accuracy Studies-2 (QUADAS-2) were completed for the included studies. Heterogeneity was assessed. For biomarkers reporting sensitivity and specificity in at least four studies, a random-effects meta-analysis was performed for biomarkers with similar cut-offs. We screened 4,651 citations and included 65 studies that enrolled 16,010 participants and evaluated 156 host proteins. Most (47/65) studies enrolled adult pulmonary TB (PTB), with 15 studies in adult extra-pulmonary TB and 5 in children. Small early-stage discovery studies with case-control design were common (24/65) and had a high risk of bias. For adult PTB, CRP, IP-10, NCAM-1, and SAA met TPP criteria in high-quality studies. There was a high degree of heterogeneity in biomarker cut-offs and study design. CRP at 10 mg/L cut-off was meta-analyzed from 10 studies; pooled sensitivity 86% [95% confidence interval (CI): 80-95] and pooled specificity 67% (95% CI: 54-79). In people living with HIV (six studies), CRP pooled sensitivity was 93% (95% CI: 90-95), and pooled specificity was 59% (95% CI: 40-78). We identified promising biomarkers that performed well in high-quality studies. Data overall are limited and highly heterogenous. Further standardized validation across subgroups in prospective studies is needed before translating into POC assays.

Importance: To our knowledge, this is the first comprehensive systematic review of host blood protein biomarkers for tuberculosis (TB), and we identified promising biomarkers for a TB screening test.

The description of new taxa and nomenclature updates to currently known taxa from aquatic animal species continues. After a review of the literature from 2022 and 2023, multiple lists of bacteria, including members of Phylum Planctomycetota, were compiled. As with the previous review, most bacteria are oxidase-positive Gram-negative bacilli with familiar families including new taxa in Aeromonadaceae, Flavobacteriaceae, and Vibrionaceae. A number of Gram-positive bacilli are described including new taxa in the Nocardioides, Paenibacillus, and Streptomyces genera. Two anaerobic species are listed, and one new member of Family Planctomycetaceae is noted. Revised taxa are briefly mentioned. The majority of new and revised taxa are isolated from healthy aquatic animals, and therefore, the role of these new bacteria in health and disease is unknown. Bacteria with pathogenic association and potential production of bioactive substances are highlighted.

Pathogenic gram-negative bacteria frequently carry genes encoding extended-spectrum beta-lactamases (ESBL) and/or carbapenemases. Of great concern are carbapenem resistant Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter baumannii. Despite the need for rapid AMR diagnostics globally, current molecular detection methods often require expensive equipment and trained personnel. Here, we present a novel machine-learning-aided platform for the rapid detection of ESBLs and carbapenemases using Loop-mediated isothermal Amplification (LAMP). The platform consists of (i) an affordable device for sample lysis, LAMP amplification, and visual fluorometric detection; (ii) a LAMP screening panel to detect the most common ESBL and carbapenemase genes; and (iii) a smartphone application for automated interpretation of results. Validation studies on clinical isolates and urine samples demonstrated percent positive and negative agreements above 95% for all targets. Accuracy, precision, and recall values of the machine learning model deployed in the smartphone application were all above 92%. Providing a simplified workflow, minimal operation training, and results in less than an hour, this study demonstrated the platform's feasibility for near-patient testing in resource-limited settings.IMPORTANCEExtended-spectrum beta-lactamases (ESBL) and carbapenemases confer resistance to third-generation cephalosporins and carbapenems in pathogenic Gram-negative bacteria such as Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter baumannii. Conventional antimicrobial susceptibility testing is based on phenotypic methods, and results can take several days to be obtained. Current genotypic detection methods can be rapid but require expensive equipment and trained personnel. In this study, we present a novel machine learning-aided platform for the rapid detection of ESBLs and carbapenemases using Loop-mediated isothermal Amplification (LAMP). The validation of the platform demonstrated percent positive and negative agreements above 95% for all targets. The newly developed platform provided a simplified workflow, minimal technical training, and results in less than an hour. This study demonstrated the platform's feasibility for rapid testing of ESBL and carbapenemases in bacteria and urine specimens.

Nontyphoidal Salmonella (NTS) is a predominant cause of invasive disease in sub-Saharan Africa especially among children under 5 years. Asymptomatic fecal shedding of NTS is hypothesized to contribute to the human-to-human transmission of NTS especially in low-resource settings. However, the role of pathogen shedding in invasive disease is unknown. This study aimed to investigate the prevalence and duration of fecal shedding of NTS among children under 5 years convalescing from invasive NTS disease and among healthy individuals in the community. Children presenting with fever of ≥38°C with or without diarrhea were recruited at four health facilities in Nairobi, between June 2021 and August 2023. Blood and stool samples collected were subjected to culture for the isolation of NTS (S. Enteritidis and S. Typhimurium). Children with NTS culture-positive samples (index cases) were followed up post-acute disease where household contacts and controls provided stool samples for isolation of NTS. NTS prevalence among the 3,293 individuals recruited was 1.52%. Asymptomatic shedding post-treatment was observed in almost one-third (31%) of the 42 index cases followed up. Of the 13 with intestinal shedding, 7 were shedding NTS of the same sequence type (ST) as the one recovered during acute disease. The longest duration of intestinal shedding was 3 months post-treatment. Of the 241 healthy individuals recruited, 8 had asymptomatic shedding of NTS, and 2 of these were closely related to those recovered from index cases. These findings support the hypothesis of human-to-human transmission of NTS in sub-Saharan Africa highlighting the possible benefit of vaccine introduction.

Importance: Asymptomatic fecal shedding of nontyphoidal Salmonella (NTS) is hypothesized to contribute to the human-to-human transmission of NTS especially in low-resource settings which could lead to invasive disease among high-risk populations, especially children. Our findings reiterate the hypothesis that human reservoirs could be important in the transmission of nontyphoidal Salmonella in sub-Saharan Africa. This underscores the importance of developing infection prevention measures which could include vaccine deployment and improving water, sanitation and hygiene infrastructure.

The prevalence of invasive candidiasis caused by non-albicans Candida species is increasing. Candida guilliermondii is an infrequent cause of candidemia but has been associated with decreased susceptibility to triazoles. Clinical data related to the infection with C. guilliermondii are sparse. Our study evaluated the antifungal susceptibility testing (AST) for C. guilliermondii isolates submitted to a reference laboratory over a 12-year period (2012-2023). AST patterns were examined using Clinical and Laboratory Standards Institute (CLSI) and European Committee on Antimicrobial Susceptibility Testing (EUCAST) epidemiological cutoff values (ECVs) and breakpoints. Where isolates were identified from patients treated at our institution, retrospective chart review was performed to describe patient risk factors, treatment approaches, and outcomes associated with C. guilliermondii fungemia. One hundred twelve blood culture isolates of C. guilliermondii were identified, and clinical data were available for 21 fungemic patients. A significant number of isolates (9.8-20.5%) were observed to be non-wild type for various triazoles. All isolates were susceptible to micafungin. A majority (76.2%) of cases of C. guilliermondii fungemia treated at our tertiary care center were hospital-acquired, and two-thirds of patients were immunocompromised at the time of diagnosis. Ten of the 21 patients died within 60 days of fungemia, although mortality was directly or partially attributed to C. guilliermondii fungemia in only four cases (19.0%). Echinocandins may be used for empiric therapy for C. guilliermondii until the results of AST are available. Further research is required to determine appropriate clinical breakpoints for triazoles.

Importance: Our study addresses a significant knowledge gap in the clinical management of this non-Candida albicans species. Our retrospective review includes comprehensive AST data for 112 Candida guilliermondii isolates, which is the largest number of isolates reported from the United States to date. Susceptibility data are supplemented by clinical outcomes, where isolates were identified for patients treated at Mayo Clinic. Key findings from our study include the observation that a notable proportion of C. guilliermondii isolates exhibit non-wild-type profiles for various triazoles. Importantly, all isolates remained susceptible to echinocandins, suggesting their efficacy as first-line therapy in the absence of timely susceptibility results. Furthermore, our study highlights the high mortality associated with C. guilliermondii fungemia in immunocompromised patients, emphasizing the urgent need for optimized treatment strategies.

Antimicrobial resistance is a growing health threat, but standard methods for determining antibiotic susceptibility are slow and can delay optimal treatment, which is especially consequential in severe infections such as bacteremia. Novel approaches for rapid susceptibility profiling have emerged that characterize either bacterial response to antibiotics (phenotype) or detect specific resistance genes (genotype). Genotypic and Phenotypic AST through RNA detection (GoPhAST-R) is a novel assay, performed directly on positive blood cultures, that integrates rapid transcriptional response profiling with the detection of key resistance gene transcripts, thereby providing simultaneous data on both phenotype and genotype. Here, we performed the first clinical pilot of GoPhAST-R on 42 positive blood cultures: 26 growing Escherichia coli, 15 growing Klebsiella pneumoniae, and 1 with both. An aliquot of each positive blood culture was exposed to nine different antibiotics, lysed, and underwent rapid transcriptional profiling on the NanoString platform; results were analyzed using an in-house susceptibility classification algorithm. GoPhAST-R achieved 95% overall agreement with standard antimicrobial susceptibility testing methods, with the highest agreement for beta-lactams (98%) and the lowest for fluoroquinolones (88%). Epidemic resistance genes including the extended spectrum beta-lactamase bla_CTX-M-15 and the carbapenemase bla_KPC were also detected within the population. This study demonstrates the clinical feasibility of using transcriptional response profiling for rapid resistance determination, although further validation with larger and more diverse bacterial populations will be essential in future work. GoPhAST-R represents a promising new approach for rapid and comprehensive antibiotic susceptibility testing in clinical settings.IMPORTANCEExposure to antibiotics causes differential transcriptional signatures in susceptible vs resistant bacteria. These differences can be leveraged to rapidly predict resistance profiles of Escherichia coli and Klebsiella pneumoniae in clinically positive blood cultures.

Diagnostic stewardship (DxS) for infectious disease testing requires a multi-disciplinary approach to optimize test selection, performance, interpretation and patient treatment. Nucleic acid amplification-based tests for the diagnosis of infectious diseases, or "molecular microbiology tests," have rapidly expanded over the past two decades. With the increased availability and complexity of these tests, there is also an increased need for collaborative approaches to optimize test use to promote positive impacts on patient care, while mitigating potential negative impact or resource waste. In this review, we provide recommendations on building collaborative DxS teams, including microbiologists and the diverse stakeholders that use and interpret molecular microbiology tests. We then detail approaches to identify high-priority molecular microbiology tests that may need utilization assessment, select appropriate diagnostic stewardship interventions, and monitor the impact of implemented interventions. This strategic process may be employed by laboratories to realize optimal testing for selected tests at their institution.