Microbiology spectrum最新文献

The value of Fourier transform infrared (FTIR) spectroscopy as an investigation tool for outbreaks caused by Acinetobacter baumannii remains inconclusive. Here, we used FTIR as a real-time tool to investigate a carbapenem-resistant A. baumannii (CRAB) outbreak and later compared the FTIR results to results of whole genome sequencing (WGS). We used single-nucleotide polymorphism (SNP) analysis to confirm the correct FTIR cutoff and validate FTIR clustering. The outbreak occurred mainly in a general intensive care unit of an Israeli tertiary care hospital and involved 22 patients detected between December 2022 and August 2023. Isolates from 21 patients and two environmental sources were analyzed by FTIR. Eleven representative isolates underwent WGS. We performed SNP analysis using the earliest isolate as the reference. FTIR revealed that the outbreak was comprised of three clusters and four singletons. FTIR results were concordant with WGS for the 11 sequenced isolates. FTIR had higher discriminatory power than multilocus sequence typing. Within-cluster SNP differences were up to 21 SNPs. This finding confirmed that the cutoff we chose to define FTIR clusters (0.252) was appropriate. FTIR uncovered that what appeared to be one CRAB outbreak was actually three parallel outbreaks. FTIR analysis of CRAB outbreaks yields results similar to WGS, but faster and at a lower cost. A SNP threshold of less than 21 SNPs is appropriate when analyzing an A. baumannii outbreak.IMPORTANCEThis study demonstrated that Fourier transform infrared (FTIR) spectroscopy could effectively identify Acinetobacter baumannii outbreaks. Serving as a practical and rapid alternative to whole genome sequencing, FTIR can significantly accelerate A. baumannii outbreak confirmation, resulting in effective infection control interventions.

Klebsiella quasipneumoniae is an emerging opportunistic pathogen increasingly associated with multidrug-resistant infections. In this study, we characterized four clinical isolates of K. quasipneumoniae subsp. quasipneumoniae assigned to the ST978-KL103 clonotype, all linked to poor clinical outcomes. Antimicrobial susceptibility testing revealed an extensively drug-resistant phenotype, including resistance to β-lactams, carbapenems, aminoglycosides, fluoroquinolones, tigecycline, and trimethoprim/sulfamethoxazole, with polymyxin B as the only effective agent. Whole-genome sequencing and comparative analysis showed that all ST978-KL103 isolates harbored multiple plasmids, including a conserved IncU-type plasmid carrying bla_IMP-4 and the tmexCD2-toprJ2 tigecycline resistance cluster. Two isolates also carried IncC-type plasmids encoding bla_NDM-1. Conjugation assays confirmed efficient horizontal transfer of both plasmid types to E. coli recipients. Although classical virulence loci such as yersiniabactin, aerobactin, and rmpA/rmpA2 were absent, the isolates exhibited strong biofilm formation and serum resistance, indicating potential for persistence and immune evasion. Cytotoxicity assays and Galleria mellonella infection models demonstrated low acute virulence compared to the hypervirulent K. pneumoniae strain NUTH-K2044. Phylogenetic analysis placed the ST978 isolates within a distinct and globally distributed clonal lineage. The close genetic relatedness among isolates, particularly between KP57 and KP58, suggests recent nosocomial transmission. Resistome profiling of nine global ST978 genomes revealed over 40 resistance genes, with our isolates uniquely harboring the tmexCD2-toprJ2 cluster. These findings highlight ST978 K. quasipneumoniae as a globally disseminated and increasingly drug-resistant lineage with the potential to emerge as a high-risk clone in clinical settings.

Importance: The emergence of extensively drug-resistant Klebsiella quasipneumoniae strains poses a significant threat to global health yet remains underrecognized due to diagnostic limitations. Our study identifies ST978-KL103 as a cryptic high-risk lineage co-harboring bla_IMP-4, bla_NDM-1, and tmexCD2-toprJ2 on transferable plasmids. These findings highlight the species' underappreciated role as a reservoir and disseminator of last-line resistance. The strong biofilm formation and serum resistance further suggest persistence potential in clinical settings. Early recognition and surveillance of such emerging clones are critical to prevent silent dissemination and therapeutic failure in healthcare environments.

The increasing frequency of infections and drug resistance of Candida albicans has emerged as significant public health challenges, highlighting the urgent need for new antifungal agents. In this study, a decapeptide AntiCADP was designed, which could effectively inhibit the growth of C. albicans. AntiCADP killed C. albicans cells by disrupting the cell membrane, inducing ROS accumulation, damaging mitochondria, and ultimately leading to cellular necrosis. Additionally, AntiCADP inhibited the hyphal morphogenesis and biofilm formation of C. albicans. AntiCADP could also kill C. albicans cells in the mature biofilm. In a mouse subcutaneous infection model, AntiCADP significantly inhibited the development of abscesses, reduced C. albicans cell counts within abscesses, and suppressed inflammatory cell infiltration at the infected area. Taken together, AntiCADP has the potential to be an antifungal agent against skin infections caused by C. albicans.IMPORTANCETo effectively cope with the increasing frequency of infections and drug resistance of Candida albicans, various types of new antimicrobial molecules have been studied. Among these molecules, antimicrobial peptides have attracted great attention. In the present study, we designed a decapeptide AntiCADP, which showed good anti-C. albicans activity in vitro and in vivo. AntiCADP killed C. albicans cells via multiple modes, including disrupting the cell membrane, inducing ROS accumulation, damaging mitochondria, and inducing cellular necrosis. AntiCADP could also inhibit hyphal morphogenesis and biofilm formation of C. albicans and kill C. albicans cells in the mature biofilm. Thus, AntiCADP had the potential against skin infections caused by C. albicans.

Chikungunya virus (CHIKV), a major cause of acute febrile illness and joint pain, remains a significant public health threat in tropical regions. Rapid and accurate detection is essential for timely clinical management and outbreak control, particularly in resource-limited settings where real-time PCR (RT-qPCR) is often impractical. We developed and validated a SHERLOCK assay coupled with recombinase polymerase amplification for CHIKV RNA detection. Analytical performance was assessed by determining the limit of detection (LOD), cross-reactivity, clinical sensitivity and specificity, and predictive values. The assay achieved an LOD of 215 copies/reaction with no cross-reactivity against other alphaviruses or flaviviruses. Clinical testing of 146 plasma samples showed a sensitivity and specificity of 94.52% and 100% with lateral-flow readout and 97.26% and 100% with fluorescence readout, respectively. This study establishes a promising CRISPR-Cas13a-based SHERLOCK platform for CHIKV detection, demonstrating high analytical performance, rapid turnaround time, and potential for future adaptation to resource-limited settings.IMPORTANCEEarly and accurate detection of chikungunya virus (CHIKV) is critical for outbreak control, especially in resource-limited settings, where real-time PCR is not feasible. This study demonstrates that the CRISPR-Cas13a-based SHERLOCK platform, combined with RPA, achieves high diagnostic accuracy and a low detection limit, comparable to RT-qPCR. The assay's rapid turnaround time and simple lateral-flow readout make it a promising tool for point-of-care diagnostics during CHIKV outbreaks, potentially improving disease surveillance and clinical decision-making.

Staphylococcus aureus clones emerge and recede in roughly 10-year intervals unless virulence factors impose clonal continuation. This study investigates a newly emergent clone. Prior to 2020, all of 322 clinical isolates tested grew, as expected, yellow colonies on mannitol salt agar. After 2020, 10% of clinical S. aureus isolates grew as intensely pink colonies (hyper-pink, not pale red as seen in coagulase-negative staphylococci). The hyper-pink S. aureus lacked the gene mannitol-1-phosphate dehydrogenase within the mannitol fermentation operon. The organisms were methicillin-sensitive. S. aureus usually expresses cell-surface virulence factors during exponential growth and secretes toxins for a brief time in the post-exponential phase, but not in the stationary phase. The hyper-pink organisms expressed both cell-surface and secreted virulence factors well into the stationary phase. S. aureus, which is usually yellow on mannitol salt, showed the virulence factor production profile of hyper-pink organisms when the media pH was maintained at 8.0. Comparison of hyper-pink organism culture fluids with those of USA300 and USA400 S. aureus strains for interaction with epithelial cells revealed similar harmful pro-inflammatory properties. Known global regulatory systems in the hyper-pink organisms were intact, including the two-component system SrrA/B, which senses both oxygen and pH. This newly emergent clone appears to have been selected after 2020 by its inability to ferment mannitol. The clone was cultured from the skin of patients with moderate to severe atopic dermatitis.IMPORTANCEA novel clone of Staphylococcus aureus emerged after 2020, unusually, with production of cell-surface and secreted virulence factors into late stationary growth phase. This newly emerging clone may be confused with coagulase-negative staphylococci because it has a strong pink phenotype on mannitol salt agar, whereas S. aureus usually forms bright yellow colonies on this medium.

The QuantiFERON-TB Gold Plus (QFT-Plus; Qiagen, Hilden, Germany) assay is widely used for latent TB infection (LTBI) screening; however, borderline results often show variability during follow-up. We investigated the longitudinal variability of borderline TB1 and TB2 results throughout the follow-up period. A total of 770 individuals with initial borderline results (0.2-0.7 IU/mL) in either the TB1 or TB2 tube were retrospectively collected over a 5-year period. Agreement and correlation between the initial TB1 and TB2 results were analyzed. Trends in reversion and conversion were evaluated separately for each tube and for combined results among individuals with available follow-up tests. Initial TB1 and TB2 borderline results showed weak agreement (Cohen's κ = 0.441, 95% CI 0.401-0.480) and moderate correlation (Spearman's ρ = 0.640, 95% CI 0.596-0.680; P < 0.001). TB2 exhibited the highest variability during the first follow-up (33.8%, 47/139), which decreased with subsequent testing. Among those with ≥2 follow-ups, 30.2% (13/43) showed result changes. Cases with borderline values in both TB1 and TB2 showed higher rates of reversion and conversion, whereas having a low-negative result (<0.2 IU/mL) in at least one tube was associated with minimal variability. The QFT-Plus assay provides separate TB1 and TB2 values, and integrating these results may improve interpretation. Follow-up testing is particularly warranted when borderline results occur in both tubes. Further studies are needed to define appropriate criteria and optimal retesting intervals.

Importance: This is the first study to longitudinally assess borderline QuantiFERON-TB Gold Plus (QFT-Plus) results in an intermediate TB burden setting, highlighting the need for defined criteria and retesting intervals.

Chronic hepatitis B virus (HBV) infection is a major cause of liver-related morbidity and mortality worldwide. Serum HBV pregenomic RNA (pgRNA) is a surrogate marker for the transcriptional activity of covalently closed circular DNA. Here, we successfully designed a novel point-of-care (POC) diagnostic platform that allows specific, sensitive, rapid, and visual identification of HBV pgRNA. The platform integrates probe-based reverse transcription loop-mediated isothermal amplification (RT-LAMP) with either restriction endonuclease-mediated real-time fluorescence (REF) or a gold nanoparticle-based lateral flow biosensor (AuNPs-LFB), termed HBV-RT-LAMP. A unique set of probe-based LAMP primers targeting HBV-pgRNA was successfully designed. The optimal conditions for HBV-RT-LAMP were determined to be 64°C and 30 min. AuNPs-LFB and a pocket fluorescence detector (REF assay) were used for readout of the products. Our assay detected the target gene at concentrations as low as 50 copies/mL of HBV RNA standard and did not produce cross-reactions with HBV DNA (treated with DNase I) or other pathogens. The entire detection process, including HBV RNA extraction (45 min), LAMP (30 min), and the interpretation of results (AuNPs-LFB, less than 2 min), could be performed within 80 min, with no need for expensive devices. Therefore, the HBV-RT-LAMP diagnostic system developed in this study can potentially serve as a useful POC diagnostic tool for the evaluation of chronic HBV infection status and antiviral drug efficacy.IMPORTANCEChronic hepatitis B (CHB) is still a serious global concern that can result in severe liver-related diseases, including liver cirrhosis and hepatocellular carcinoma. Serum hepatitis B virus (HBV)-pregenomic RNA (pgRNA) has been proposed as a surrogate intrahepatic covalently closed circular DNA marker in CHB patients. Here, for the first time, a novel point-of-care diagnostic platform, termed HBV-reverse transcription loop-mediated isothermal amplification (RT-LAMP), which integrates probe-based RT-LAMP with either restriction endonuclease-mediated real-time fluorescence (REF) detection or a gold nanoparticle-based lateral flow biosensor (AuNPs-LFB), was developed and applied for accurate, sensitive, specific, rapid, and visual identification of HBV-pgRNA.

Enterovirus (EV) exhibits high evolutionary activity and is associated with a wide spectrum of human diseases. Environmental surveillance employing next-generation sequencing (NGS) is an effective approach for elucidating EV type diversity and genetic variation; however, similar studies in China are limited. Sewage samples were collected monthly in Jinan, China, from January to December 2024 and subsequently concentrated. EV isolation was conducted using cell culture, followed by VP1 amplification and Sanger sequencing. Concurrently, VP1 amplicons directly obtained from sewage concentrates were subjected to NGS. Similarity and phylogenetic analysis were performed. A total of 104 EV strains representing nine serotypes were isolated via cell culture, with echovirus 11 (E11) and E3 being the most prevalent, accounting for 34.62% (36/104) and 27.88% (29/104) of the total isolates, respectively. NGS technology identified 31 EV types, with CVB4 and CVA16 exhibiting the highest read counts. Additionally, uncommon types such as D68, A71, A76, B88, A90, and C99 and 11 rhinovirus types were detected in the sewage samples. Phylogenetic analysis revealed that CVA16 and D68 sequences from sewage had close genetic relationship with recent Chinese strains, and multiple branches of B88 and C99 were observed. This study represents the first detection of D68 and B88 in sewage in China, demonstrating co-circulation of diverse enteroviruses with significant genetic variability in the region. These findings underscore the utility of environmental surveillance for tracking EV epidemiology and evolution.IMPORTANCEEnteroviruses are significant human pathogens associated with frequent epidemics and outbreaks. Although numerous types exist, some exhibit low prevalence and are rarely detected. By implementing next-generation sequencing (NGS) in wastewater surveillance, this study comprehensively characterized enterovirus diversity, revealing multiple uncommon types of interest. Notably, we detected EV-D68-an emerging agent linked to acute flaccid myelitis-in sewage, underscoring its public health relevance. Additionally, we identified multiple rhinovirus types. These data demonstrate the local circulation of diverse viruses with substantial public health implications and provide a critical foundation for disease monitoring and early warning systems.