Journal of Clinical Microbiology最新文献

{"title":"Antibiotic resistance profiles of seven genomospecies of <i>Corynebacterium jeikeium</i> analyzed by whole genome sequencing.","authors":"Michaela Fischer-Wellenborn, Frank Imkamp, Valéria Pereira Pires, Reinhard Zbinden, Nicolas Personnic","doi":"10.1128/jcm.00418-25","DOIUrl":"10.1128/jcm.00418-25","url":null,"abstract":"<p><p><i>Corynebacterium jeikeium</i> is an opportunistic human pathogen with high genomic diversity. In the past, multi-drug resistance has been considered a hallmark of <i>C. jeikeium</i>. However, some strains of <i>C. jeikeium</i> are fully susceptible and thus, their reclassification would avoid the empirical therapy with glycopeptides. One hundred eighty-six clinical isolates and four reference isolates identified as <i>C. jeikeium</i> between 1994-1999 and 2012-2019, respectively, were analyzed by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS) and 16S ribosomal RNA (16S rRNA) gene sequence analysis. A total of 153 confirmed <i>C. jeikeium</i> isolates was further analyzed by whole genome sequencing (WGS) and by disk diffusion antimicrobial susceptibility testing (AST). WGS and subsequent core genome single nucleotide polymorphism-based phylogenetic analysis segregated the isolates into seven phylogenetic clusters. The average nucleotide identity between the clusters was <u><</u>95%, qualifying them as separate <i>C. jeikeium</i> genomospecies. Overall, AST profiles of all 153 isolates were heterogeneous for six of the 16 antibiotics tested. Notably, genomospecies 6 and 7 displayed mainly comparably small inhibition zones. For the other 10 antibiotics, all genomospecies displayed either small or large inhibition zones, respectively. Our study of <i>C. jeikeium</i> clinical isolates revealed a relatively complex phylogenetic structure. As soon as clear phenotypic traits can be attributed to the seven genomospecies by routine diagnostic tests, genomospecies encompassing mainly antibiotic-susceptible strains should be reclassified to fine-tune potential treatment strategies.IMPORTANCE<i>Corynebacterium jeikeium</i> isolates typically display multi-drug resistance (MDR), frequently triggering empirical therapies with glycopeptides. However, some strains are susceptible to a broad range of antibiotics. In our study, analysis of a large collection of <i>C. jeikeium</i> by whole genome sequencing (WGS) and antibiotic resistance testing revealed genomic diversity, and some correlations between MDR phenotypes and specific genomospecies. While WGS is not yet a routine method, identification of MDR genomospecies may confirm the need for glycopeptide therapy. Identification of a less-resistant genomospecies would have even a higher clinical impact, as unnecessary therapies with glycopeptides are avoided, thereby reducing the selection for vancomycin-resistant enterococci. In the future, databases of diagnostic tools, e.g., MALDI-TOF, may be expanded to allow for the differentiation of genomospecies. Furthermore, rational taxonomic reclassification of some <i>C. jeikeium</i> genomospecies would help clinicians to fine-tune potential treatment strategies.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0041825"},"PeriodicalIF":5.4,"publicationDate":"2025-10-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12505879/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144955902","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Performance and potential utility of the BioFire Joint Infection Panel with synovial fluid and joint tissue specimens.","authors":"Shivani Fox-Lewis, Marc Douglass, Sally Roberts","doi":"10.1128/jcm.00594-25","DOIUrl":"10.1128/jcm.00594-25","url":null,"abstract":"<p><p>The BioFire Joint Infection Panel (BJIP) (bioMérieux) is a rapid sample-to-answer multiplex polymerase chain reaction (PCR) platform for the diagnosis of joint infections. This retrospective study evaluated its performance with synovial fluid and joint tissue specimens from native and prosthetic joints. Joint fluid and tissue specimens received from November 2023 to September 2024 were included. Some specimens were pooled prior to BJIP testing. BJIP results were compared to composite standard laboratory testing (culture, 16S rRNA gene PCR and sequencing [where performed], and concurrent microbiological results for that infection episode). Clinical data were collected (diagnosis, surgical history, antibiotic treatment). Subgroup analysis of native and prosthetic joint specimens was conducted. There were 224 specimens from 134 patients: 107 synovial fluids and 117 joint tissues. The most common organisms, reliably detected by the BJIP, were <i>Staphylococcus aureus</i> and <i>Streptococcus</i> spp. The on-panel positive percent agreement (PPA) was 90.5% for synovial fluids and 86.8% for joint tissues. There was no significant difference in the PPA for synovial fluids vs joint tissues, nor for native vs prosthetic joints. The additional diagnostic yield was one case. The absence of important pathogens in prosthetic joint infection, e.g., <i>Cutibacterium acnes</i>, reduces the overall PPA (74.5% synovial fluid, 77.6% joint tissue). The BJIP performs well in synovial fluid and joint tissue specimens, from native and prosthetic joints for on-panel organisms. There was minimal additional diagnostic yield. We suggest an implementation pathway using the BJIP upfront to provide rapid results to optimize patient care.IMPORTANCEThis study compared the BioFire Joint Infection polymerase chain reaction panel to standard culture-based methods for detecting organisms causing joint infections. This is a relatively new test, and the data published so far show it performs well with synovial fluid. There are limited published data using it for joint tissue specimens. We found that the BioFire Joint Infection Panel reliably detects pathogens in synovial fluid and joint tissue specimens, though its performance is limited by the absence of important pathogens associated with prosthetic joint infection, e.g., <i>Cutibacterium acnes</i>. We suggest using the panel to provide rapid diagnosis to optimize patient care.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0059425"},"PeriodicalIF":5.4,"publicationDate":"2025-10-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12505999/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144956119","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The potential to improve Lyme disease diagnostics through quantification of immunoglobulin class switching patterns.","authors":"Anna M Schotthoefer","doi":"10.1128/jcm.01020-25","DOIUrl":"https://doi.org/10.1128/jcm.01020-25","url":null,"abstract":"<p><p>N. Nair, A. Marques , E. J. Horn, G. Brown et al., J Clin Microbiol 63:e0034725, 2025, https://doi.org/10.1128/jcm.00347-25 present data to demonstrate that infection by <i>Borrelia burgdorferi</i>, the primary causative agent of Lyme disease in the USA, leads to immunoglobulin class switching in antibodies specifically against the immunodominant antigen, VlsE (variable major protein-like sequence, expressed), in a predictable pattern between the early acute (<1 month illness duration) to late-stage Lyme arthritis stages. Detection of anti-VlsE isotypes in sera was highly specific to Lyme disease patients, and the frequencies and abundances of IgM, IgG, and IgA isotypes progressed in a pattern consistent with the development of an anti-<i>B</i>. <i>burgdorferi</i> antibody response. Applying multivariate and machine learning modeling methods, they found the profile of isotypes quantified performed particularly well at correctly classifying early acute sera samples. They also identified IgG4 as a potential biomarker unique to Lyme arthritis patients. Overall, the findings suggest that improvements in Lyme disease diagnostics may be attained by quantifying specific antibody isotypes against <i>B. burgdorferi</i> antigens.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0102025"},"PeriodicalIF":5.4,"publicationDate":"2025-09-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145149148","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Off-label evaluation of the BD MAX MDR-TB assay for rapid diagnosis of rifampicin and isoniazid resistance of <i>Mycobacterium tuberculosis</i> clinical isolates in a high-volume reference laboratory.","authors":"Angela Pires Brandao, Fabiane Maria de Almeida Ferreira, Fernanda Cristina Dos Santos Simeao, Lucilaine Ferrazoli, Erica Chimara, Rosângela Siqueira de Oliveira, Juliana Maira Watanabe Pinhata","doi":"10.1128/jcm.00912-25","DOIUrl":"https://doi.org/10.1128/jcm.00912-25","url":null,"abstract":"<p><p>Drug-resistant tuberculosis (TB) remains a primary global health concern. Multidrug-resistant TB is defined by resistance to at least rifampicin (RIF) and isoniazid (INH), the two key drugs used in TB treatment. The BD MAX Multi-Drug Resistant Tuberculosis (BD MAX) assay is a fully automated real-time PCR platform recommended by the World Health Organization for the initial diagnosis of TB and RIF and INH resistance (RIF-R and INH-R) directly from pulmonary clinical samples. This study aimed to assess the off-label performance of BD MAX in clinical <i>M. tuberculosis</i> complex (MTBC) isolates under routine laboratory conditions. The assay was first validated using non-tuberculous mycobacteria (NTM) and MTBC isolates with known mutations. For real-world validation, it was compared to the GenoType MTBDR<i>plus</i> by testing 1,440 clinical isolates prospectively. The BD MAX assay correctly excluded MTBC from all NTM cultures. Among MTBC isolates with known mutations, it identified 19 of 20 RIF-R isolates and 14 of 15 INH-R isolates. In prospective testing, BD MAX achieved 99.6% sensitivity (1,403/1,409), 96.8% specificity (30/31), and 99.5% overall accuracy (1,433/1,440) for MTBC detection. For drug resistance detection, it showed 95.2% (40/42) concordance for RIF, 96.8% (30/31) for INH, and 81.3% (13/16) for MDR when compared to MTBDR<i>plus</i>. Discrepancies between MTBDR<i>plus</i> and BD MAX included heteroresistant cases and unreportable resistance results by BD MAX due to infrequent mutations or low bacterial load. Overall, this study confirms BD MAX as an accurate and reliable tool for MTBC detection and drug resistance profiling in clinical isolates in high-volume TB laboratories.IMPORTANCEThis study highlights the importance of the BD MAX Multi-Drug Resistant Tuberculosis assay (BD MAX) applied in clinical isolates for the detection of multidrug-resistant tuberculosis (MDR-TB), i.e., <i>Mycobacterium tuberculosis</i> resistance to rifampicin and isoniazid. TB is a global health issue, and drug-resistant TB makes treatment more difficult, favoring transmission and disease amplification. The BD MAX platform offers a faster and more automated way to detect TB and drug resistance. The study showed that BD MAX, applied off-label in clinical isolates, accurately identified TB and resistance to rifampicin and isoniazid, with results comparable to those of the widely used line probe assay. This is significant in a high-volume laboratory because it is more straightforward and more rapid than the line probe assay. BD MAX showed some limitations, especially in detecting rare mutations and in cases of low bacterial levels. Overall, this tool could improve TB care, especially in high-volume laboratories.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0091225"},"PeriodicalIF":5.4,"publicationDate":"2025-09-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145124017","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"An extraction-free, lyophilized one-pot RAA-CRISPR assay for point-of-care testing of <i>Haemophilus influenzae</i>.","authors":"Yaling Cao, Junwen Wang, Zihao Fan, Ling Xu, Zhenzhen Pan, Yinkang Mo, Qiwen Yang, Jing Huang, Feng Ren","doi":"10.1128/jcm.00535-25","DOIUrl":"https://doi.org/10.1128/jcm.00535-25","url":null,"abstract":"<p><p>Timely and accurate diagnosis is essential for the effective treatment of <i>Haemophilus influenzae</i> (HI) infection. Notably, a rapid point-of-care testing (POCT) method for the diagnosis of HI DNA is still lacking. The aim of this study was to establish a one-pot detection device for HI DNA detection using CRISPR/Cas13a that would be applicable for POCT. We established a two-step polymerase chain reaction (PCR)-CRISPR assay, a two-step recombinase-aided amplification (RAA)-CRISPR assay, and a one-pot RAA-CRISPR assay. Additionally, reagent lyophilization, rapid lysis technology, and a one-pot detection device were integrated to construct an extraction-free one-pot detection device, named EFORCA (extraction-free one-pot RAA-CRISPR/Cas13a assay). Validation was performed on 90 simulated samples and 77 clinical samples. The two-step PCR/RAA-CRISPR assay for HI DNA detection was established and optimized, with a detection limit of 1 copy/μL and 100% specificity. The sensitivity and specificity of the two-step PCR-CRISPR assay for the 90 simulated clinical samples were 96.7% and 95%, respectively, and for the 77 clinical samples, 97.5% and 100%, respectively. The one-pot RAA-CRISPR assay and the EFORCA detected 50 and 25 CFU/mL HI in bacterial suspensions within 30 min, respectively. The valuation results for 77 clinical samples demonstrated that the sensitivity of both methods was 95%, which was greater than the sensitivity of quantitative real-time PCR (qPCR) (92.5%) with the same specificity of 100%. We developed an extraction-free one-pot RAA-CRISPR/Cas13a assay for HI detection, which effectively performs a POCT test and is a valuable tool for the early detection and monitoring of HI infection.IMPORTANCETimely and accurate diagnosis is essential for the effective treatment of <i>Haemophilus influenzae</i> (HI) infection. Notably, a rapid point-of-care testing (POCT) method for the diagnosis of HI DNA is still lacking. In this study, sensitive two-step polymerase chain reaction (PCR)-CRISPR and recombinase-aided amplification (RAA)-CRISPR assays were developed, followed by the establishment of the one-pot RAA-CRISPR assay through integration of RAA, CRISPR, and rapid lysis for HI detection. Additionally, reagent lyophilization, rapid lysis technology, and a one-pot detection device were integrated to construct the extraction-free one-pot RAA-CRISPR/Cas13a assay (EFORCA). With its high sensitivity, specificity, rapid turnaround time, and operational simplicity, this assay shows great potential as a practical diagnostic tool for HI infection in small laboratory settings.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0053525"},"PeriodicalIF":5.4,"publicationDate":"2025-09-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145124057","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Transforming tuberculosis diagnosis with clinical metagenomics: progress and roadblocks.","authors":"Yuqing Chen, Hui Tang, Jieyuan Zheng, Qing Yang, Dongsheng Han","doi":"10.1128/jcm.00537-25","DOIUrl":"https://doi.org/10.1128/jcm.00537-25","url":null,"abstract":"<p><p>Tuberculosis (TB) remains a leading global infectious killer, yet traditional diagnostic methods are inadequate. Acid-fast staining suffers from low sensitivity, and mycobacterial culture requires prolonged incubation because of the slow growth of <i>Mycobacterium tuberculosis</i>. PCR-based molecular assays allow rapid detection, but their capacity for resistance profiling is limited to a narrow set of mutations. Metagenomic next-generation sequencing (mNGS) has emerged as a promising culture-independent tool for TB detection, enabling broad-spectrum pathogen identification and offering added value in complex scenarios including extra-pulmonary disease, mixed infections, and infections in immunocompromised or pediatric populations. Clinical studies indicate that mNGS achieves moderate to high sensitivity and excellent specificity in the diagnosis of tuberculosis. However, its diagnostic performance is often constrained by low mycobacterial read counts, interference from abundant host nucleic acids, and the inability to distinguish active from latent infection. In addition, the accuracy of drug resistance prediction using mNGS remains limited, and the World Health Organization currently endorses targeted NGS as the preferred sequencing-based approach for resistance profiling. Despite these challenges, mNGS has facilitated novel diagnostic strategies that combine pathogen detection with host-response data, thereby broadening its potential clinical utility. Nevertheless, practical barriers such as high cost, complex laboratory workflows, and difficulties in data interpretation continue to restrict widespread adoption in routine practice. Future efforts should prioritize technical optimization, standardized protocols, and integration with conventional diagnostics to establish cost-effective and clinically meaningful roles for mNGS in TB diagnosis and management.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0053725"},"PeriodicalIF":5.4,"publicationDate":"2025-09-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145086369","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Toward diagnostic preparedness: detection of highly pathogenic avian influenza A(H5N1) in contrived nasal swab specimens using rapid antigen and point-of-care molecular tests.","authors":"Leda Bassit, Gregory L Damhorst, Heather B Bowers, Courtney Sabino, Julie Sullivan, Emily B Kennedy, Jacob Khouri, Pamela Miller, Eric Lai, Raymond F Schinazi, Wilbur A Lam, Nira R Pollock, Anuradha Rao","doi":"10.1128/jcm.00548-25","DOIUrl":"10.1128/jcm.00548-25","url":null,"abstract":"&lt;p&gt;&lt;p&gt;Highly pathogenic avian influenza (HPAI) A(H5N1) clade 2.3.4.4b was first detected in birds in 2021 in the United States (U.S.). An ongoing outbreak in dairy cattle began in early 2024 involving genotype B3.13. At least 70 U.S. cases have been identified in humans with exposure to infected cattle, poultry, and wild birds. No human-to-human transmission has yet been documented. However, for diagnostic preparedness, we evaluated the ability of currently available influenza tests to detect 2024 U.S. H5N1 genotypes, B3.13 and D1.1, predominant in wild birds and poultry. Contrived nasal swab samples were prepared using live or inactivated virus and used to test 12 rapid antigen tests (lateral flow assays, [LFA]), including 10 commercially available influenza A and two H5-specific LFAs. Five point-of-care (POC) molecular assays were also tested. An inclusivity testing protocol utilizing a predetermined dilution series was used to evaluate each assay, enabling a head-to-head comparison of performances. All LFAs and POC molecular tests were able to detect bovine 2024 H5N1 genotype B3.13. Sensitivity for the POC molecular tests (heat-inactivated B3.13) ranged from 1.55 to 7.75 TCID&lt;sub&gt;50&lt;/sub&gt;/swab. For 11/12 LFAs, 10 commercial influenza A, and an RUO H5 assay, the sensitivity (live B3.13) ranged from 78 to 1550 TCID&lt;sub&gt;50&lt;/sub&gt;/swab. The 12th LFA detected 77,500 TCID&lt;sub&gt;50&lt;/sub&gt;/mL transport media. Testing of four LFAs confirmed inclusivity for genotype D1.1. Available influenza tests can detect 2024 U.S. H5N1 strains in contrived samples with a wide range of analytical sensitivity. In the event of human-to-human transmission, clinical performance and optimal sample types would need to be established.IMPORTANCETo date, at least 70 human cases of highly pathogenic avian influenza A (HPAI) H5N1 have been reported. Human-to-human transmission has not yet been reported, but there is ongoing HPAI H5N1 spread within animal and bird populations. As part of pandemic preparedness, it is critical to know whether available tests capable of detecting seasonal influenza A can also detect HPAI H5N1. Our paper describes results from the first study to assess and compare the ability of 10 commercially available influenza A lateral flow assays (LFAs), two LFAs specific for H5 influenza, and five POC molecular tests from three manufacturers to detect the H5N1 virus circulating in U.S. cattle (genotype B3.13). We found that all tests detect HPAI H5N1 in contrived nasal swab specimens, with varying degrees of analytical sensitivity. We also tested a subset of LFAs (three 510k-cleared COVID/flu multiplex tests and an RUO H5-specific LFA) to confirm that they were able to detect a D1.1 genotype (the predominant genotype in wild birds and poultry) strain isolated from a human case of H5N1 in Washington State. Given the persistent circulation of HPAI H5N1 cases, it is critical to know that available tests can analytically detect H5N1. Clinical performance and optimal ","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0054825"},"PeriodicalIF":5.4,"publicationDate":"2025-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12421893/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144873440","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Detection, quantitation, and genotyping of human papillomavirus circulating tumor DNA by droplet digital PCR.","authors":"Emily C Fernholz, David M Routman, Kathryn M Van Abel, Eric J Moore, Daniel J Ma, Danielle E Hunter, Kathleen R Bartemes, James S Lewis, Erik B Wendlandt, Matthew J Binnicker","doi":"10.1128/jcm.00585-25","DOIUrl":"10.1128/jcm.00585-25","url":null,"abstract":"<p><p>Human papillomavirus (HPV) is comprised of >200 genotypes and has an ~8 kb, circular, double-stranded DNA genome. Transmission of HPV occurs through skin-to-skin contact and infection of squamous epithelial cells of cutaneous and mucosal surfaces. HPV genotypes are categorized as low- or high-risk (hrHPV) based on oncogenic potential. There are approximately 14 types of hrHPV that can cause several types of cancer, including HPV-associated oropharyngeal squamous cell carcinoma (HPV(+)OPSCC). Detection of HPV(+)OPSCC is traditionally accomplished using p16 immunohistochemistry (IHC) and HPV-specific testing, either DNA or RNA <i>in situ</i> hybridization (ISH) staining or DNA-based PCR of suspected tumor biopsy tissue. More recently, platelet-poor plasma (PPP) samples from patients with HPV(+)OPSCC have proven useful for detection and quantitation of fragments of HPV circulating tumor DNA (ctDNA). ctDNA has been shown to be useful in determining treatment response and monitoring for disease recurrence. In this study, a novel droplet digital PCR assay (ddPCR) was developed and validated for the detection and quantitation of ctDNA from 5 hrHPV genotypes in PPP. Analytical sensitivity ranged from 7.71 to 19.45 fragments of HPV ctDNA per milliliter of PPP across five hrHPV genotypes. In patients with confirmed primary or recurrent HPV(+)OPSCC or HPV(-)OPSCC, testing of corresponding PPP samples (<i>n</i> = 32) by ddPCR demonstrated 90.63% (29/32) overall agreement with p16/HPV-ISH biopsy results. Compared with reference ddPCR assays performed at outside laboratories, our ddPCR assay yielded 90% (9/10) overall agreement. This assay may provide clinicians with a tool for monitoring HPV ctDNA prior to, during, and after treatment of an HPV-associated cancer.</p><p><strong>Importance: </strong>At least 14 genotypes of human papillomavirus (HPV) have been identified to have high oncogenic potential. While molecular diagnostic testing for HPV is widely available for liquid cytologic cervical samples, testing is limited for other sample types, including liquid biopsy samples, such as platelet-poor plasma (PPP). With the rising incidence of HPV-associated oropharyngeal squamous cell carcinoma (HPV(+)OPSCC), laboratory testing is an essential part of patient diagnosis, management, and surveillance. Here, we summarize the development and analytical performance validation of a multiplexed, droplet digital PCR (ddPCR) assay for the detection and quantitation of HPV circulating tumor DNA (ctDNA) in PPP. This assay may provide clinicians with a tool to address minimal residual disease for patients with an HPV-associated cancer.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0058525"},"PeriodicalIF":5.4,"publicationDate":"2025-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12421868/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144873437","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The impact of commercially available media on cefiderocol susceptibility testing by broth microdilution method.","authors":"Boudewijn L M DeJonge, Christine Slover, Sean T Nguyen, Christopher Longshaw, Miki Takemura, Naomi Anan, Hidenori Yamashiro, Yoshinori Yamano","doi":"10.1128/jcm.00471-25","DOIUrl":"10.1128/jcm.00471-25","url":null,"abstract":"<p><p>The reference method for determining cefiderocol MIC is broth microdilution (BMD) using iron-depleted cation-adjusted Mueller-Hinton broth (ID-CAMHB). However, laboratories have reported variable MIC findings and difficulties in determining MIC endpoints. To investigate this, the BMD method was re-evaluated by examining the chelation time for iron depletion of the media and by testing reproducibility using four MHB sources (BD-BBL, BD-Difco, Oxoid, and Merck) and 85 Gram-negative isolates (<i>Escherichia coli</i>, <i>Klebsiella pneumoniae</i>, <i>Pseudomonas aeruginosa</i>, and <i>Acinetobacter baumannii</i>), of which 63 were previously evaluated <i>in vivo</i>. Extending chelation time to 6 h provided optimal iron reduction to ≤0.03 µg/mL for all media. MIC reproducibility was high for each medium source and varied by species (93.3%-99.0% of MIC values within one dilution of modal MIC values); however, different sources of ID-CAMHB produced (reproducible) different modal MIC values for isolates. ID-CAMHB from BD-BBL and BD-Difco showed the best correlation with 96.5% of isolates showing modal MIC values within one dilution between media. MIC values generated with these two media were the most predictive of efficacy <i>in vivo</i>. When trailing was observed, it was seen across all media, and refined reading guidelines were applied to enhance reproducibility of determining the MIC endpoint. The 6-h chelation and the refined reading guidelines for trailing are now incorporated into the Clinical and Laboratory Standards Institute M100 document. When using the BMD method to determine cefiderocol susceptibility, the impact of media should be considered, and based on these results, ID-CAMHB prepared with BD-BBL or BD-Difco is recommended.IMPORTANCEThe reference antibiotic susceptibility testing method for cefiderocol is the broth microdilution method approved by the Clinical and Laboratory Standards Institute in 2016, using iron-depleted cation-adjusted Mueller-Hinton broth. A few manual devices have come to the market replicating this method, but inconsistent results with these devices have been reported. The current study identified the source of Mueller-Hinton broth as a variable in cefiderocol MIC determinations and provides detailed description of the preparation of iron-depleted cation-adjusted Mueller-Hinton broth, and revised reading guidance to improve reproducibility of cefiderocol MIC determinations for <i>Escherichia coli</i>, <i>Klebsiella pneumoniae</i>, <i>Pseudomonas aeruginosa</i>, and <i>Acinetobacter baumannii</i>. The recommendations made in this study should enhance the reproducibility of cefiderocol broth microdilution susceptibility testing.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0047125"},"PeriodicalIF":5.4,"publicationDate":"2025-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12421808/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144956126","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Clinical development and performance of the First to Know Syphilis Self-Test for over-the-counter usage: a <i>de novo</i> rapid test for treponemal antibody.","authors":"Kevin Clark, Antony George Joyee, Diane Biddison, Sharon Rabine, Jianghong Qian, Sydney Spradlin, Vicki Thompson, Qinwei Shi, Jing Xu, Lu Zhang, Alicia Brown, Lisa Nibauer, Raji Pillai, Jody D Berry","doi":"10.1128/jcm.00244-25","DOIUrl":"10.1128/jcm.00244-25","url":null,"abstract":"<p><p>The resurgence of syphilis in the USA and globally has hastened the need for widely available syphilis testing. Early detection of syphilis is crucial for avoiding serious clinical complications and preventing the spread of infection. Self-tests enhance access to testing and promote timely treatment for positive cases. Herein, we describe the performance of First to Know Syphilis Test (FTK), the first over-the-counter (OTC) treponemal test. A prospective multi-site clinical study with 1,270 subjects was conducted, where subjects self-collected fingerstick capillary blood and self-tested without assistance. FTK test results were compared with a composite reference standard to evaluate the diagnostic performance. The FTK test exhibited an overall sensitivity of 93.4% (95% CI, 87.0% to 96.8%) and specificity of 99.5% (95% CI, 98.9% to 99.8%) and accuracy of 99%. The overall agreement was 98.9%, with Cohen's Kappa (ⱪ) value 0.93 (95% CI, 0.90 to 0.97) showing excellent agreement between the reference standard and FTK results. In addition, the test was validated in a panel of 125 clinically staged syphilis patient samples, and the results showed 100% agreement in detecting anti-treponemal antibodies in all the samples. These data show excellent performance of the FTK Test, demonstrating its utility in the screening and diagnosis of syphilis. The availability of this OTC test and its excellent performance will have a profound impact on syphilis detection and prevention strategies and could reduce comorbidities such as HIV and transmission of other STIs.</p><p><strong>Importance: </strong>The resurgence of syphilis in the USA and globally underscores the urgent need for rapid, accessible testing. The First To Know Syphilis Test, as the first at-home, over-the-counter (OTC) test for syphilis, represents a significant breakthrough in rapid diagnostics for increased access to testing and support early diagnosis and treatment. The FTK test demonstrated excellent overall clinical performance, with 93.4% sensitivity and 99.5% specificity, confirming its reliability for syphilis detection. This novel, easy-to-use OTC syphilis test allows individuals to privately test at home and can promote testing among those hesitant or unable to seek traditional healthcare, reducing barriers like stigma and limited access. The availability of this OTC test will have a significant impact on syphilis detection and prevention strategies and reduction of comorbidities such as HIV.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0024425"},"PeriodicalIF":5.4,"publicationDate":"2025-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12421861/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144784395","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}