Journal of microbiological methods最新文献

{"title":"Identification of novel Listeria monocytogenes serotype 4h with specific antisera against wall teichoic acid","authors":"Zhenhua Wu ,&nbsp;Hao Yao ,&nbsp;Jing Li ,&nbsp;Fanzeng Meng ,&nbsp;Chao Chen ,&nbsp;Ye Wang ,&nbsp;Yuelan Yin ,&nbsp;Xin'an Jiao","doi":"10.1016/j.mimet.2025.107210","DOIUrl":"10.1016/j.mimet.2025.107210","url":null,"abstract":"<div><div><em>Listeria monocytogenes</em> (<em>Lm</em>) strains are serotyped based on their somatic (O) and flagellar (H) antigens. The newly designated serotype (ST) 4h exhibits a greater ability to colonize organs than well-characterized hypervirulent strains. Therefore, simple and rapid identification methods for <em>Lm</em> ST 4h are urgently needed for effective monitoring and prevention. Here, we report the development of rabbit polyclonal antibodies against the O-antigen of <em>Lm</em> ST 4h, targeting its unique galactosylated wall teichoic acid (WTA) structure. These antisera exhibit high titers and enable the rapid and accurate identification of ST 4h isolates via slide agglutination and ELISA assays, demonstrating their specificity in labeling <em>Lm</em> ST 4h. The WTA-antisera provide a valuable tool for investigating the pathogenic mechanisms of hypervirulent strains and enhancing existing methods for <em>Lm</em> ST identification.</div></div>","PeriodicalId":16409,"journal":{"name":"Journal of microbiological methods","volume":"236 ","pages":"Article 107210"},"PeriodicalIF":1.9,"publicationDate":"2025-08-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144763796","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Validation of a Relative Centrifugal Force method for the enumeration and detection of Campylobacter from chicken carcass rinsates","authors":"Joanne M. Kingsbury ,&nbsp;Anne Midwinter ,&nbsp;John Mills ,&nbsp;Mark Englefield ,&nbsp;Roy Biggs ,&nbsp;Anne-Marie Perchec Merien ,&nbsp;Nicola Dermer ,&nbsp;Aswathi Soni ,&nbsp;Milana Blakemore","doi":"10.1016/j.mimet.2025.107207","DOIUrl":"10.1016/j.mimet.2025.107207","url":null,"abstract":"<div><div>Campylobacteriosis is the most frequently notified foodborne disease in New Zealand and poultry is the predominant infection source. New Zealand monitors <em>Campylobacter</em> present in poultry carcass rinsates under the National Microbiological Database (NMD) programme. To better monitor <em>Campylobacter</em> control improvements, a more sensitive method is required that can enumerate rinsates with lower <em>Campylobacter</em> numbers. This study developed a modification of the current NMD method involving adding a relative centrifugal force (RCF) step for concentrating <em>Campylobacter</em> from poultry carcass rinsates. Centrifugation for 30 min significantly improved <em>Campylobacter</em> recovery compared with 15 min (<em>p</em> &lt; 0.001), but there were no differences between RCFs of 3500, 4000 and 4430 x g (<em>p</em> = 0.992). RCF and NMD method performances were compared in a single laboratory validation study that used different inoculation levels of twelve <em>Campylobacter</em> strains, including poultry isolates. <em>Campylobacter</em> was detected from more samples (<em>p</em> &lt; 0.001) using the RCF method (93 of 126; 73.8 %) than the NMD method (65 of 126; 51.6 %). The RCF method had a seven-fold lower detection limit (28 colony forming units (CFU)/400 ml) than the NMD method (200 CFU/400 ml). The detection limit accounted for an observed 70.3 % of the inoculated CFU captured within the centrifuged pellet. <em>Campylobacter</em> was also detected from significantly more (<em>p</em> &lt; 0.001) commercial chicken rinsate samples tested by poultry industry laboratories using the RCF method (257 of 863; 29.8 %) than the NMD method (114 of 863; 13.2 %). Taken together, results support the RCF method as a modification of the NMD method to enumerate lower numbers of <em>Campylobacter</em> in rinsates.</div></div>","PeriodicalId":16409,"journal":{"name":"Journal of microbiological methods","volume":"236 ","pages":"Article 107207"},"PeriodicalIF":1.9,"publicationDate":"2025-07-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144768688","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Synthesis and evaluation of lipophilic fluorescent probes for the labelling of Listeria and their impact on biofilm formation","authors":"Nika Janež ,&nbsp;Márta Ladányi ,&nbsp;Nika Zaveršek ,&nbsp;Petra Čotar ,&nbsp;Aleksandar Sebastijanović ,&nbsp;Janez Štrancar ,&nbsp;Jerica Sabotič ,&nbsp;Stane Pajk","doi":"10.1016/j.mimet.2025.107206","DOIUrl":"10.1016/j.mimet.2025.107206","url":null,"abstract":"<div><div>Imaging bacterial biofilms using confocal fluorescence microscopy is used to study their structures, but its wider application is constrained by the limited availability of effective labelling tools. Small chemical fluorescent probes offer a versatile alternative to heterologous expression of fusion or reporter proteins, but data on their effects on biofilm formation are lacking. In this study, we synthesized a series of new lipophilic fluorescent probes based on Nile blue, Nile red and coumarin scaffold. We investigated them for the labelling of <em>Listeria</em> biofilms and determined their effects on the growth and biofilm biomass formation. The Nile red probe <strong>SP-AM 7</strong> and the coumarin probe <strong>PAG 31</strong> inhibited biofilm development and showed a strong bactericidal effect. The Nile blue probe <strong>PAG 19</strong> had the least effect on the tested parameters, but labelled slowly, while the fast-labelling Nile red probe <strong>SP-AM 8</strong> promoted biofilm formation. Both are suitable for use during biofilm growth, resulting in less variation in biomass-related measurements than probes added prior to imaging. In the 3D imaging-based measurements for selected probes, we found no difference in the total biomass formed compared to the control dye, but a redistribution of biomass in the 3D layers was observed. Other probes were found to be slow to label, leave traces of unused probes or interfere with attachment to the surface. Our results show that fluorescent probe labelling should be evaluated from chemical, physical and biological points of view to understand their reliability and credibility.</div></div>","PeriodicalId":16409,"journal":{"name":"Journal of microbiological methods","volume":"236 ","pages":"Article 107206"},"PeriodicalIF":1.9,"publicationDate":"2025-07-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144758094","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Diagnostic potential of recombinant Mycobacterium tuberculosis PcaA antigen and its enhancement of protective efficacy as a subunit vaccine booster following BCG priming","authors":"Yun Xu ,&nbsp;Xiaochun Wang ,&nbsp;Qiangsen Zhong","doi":"10.1016/j.mimet.2025.107202","DOIUrl":"10.1016/j.mimet.2025.107202","url":null,"abstract":"<div><div>Bacillus Calmette-Guérin (BCG) is the only vaccine currently used in clinical practice to prevent tuberculosis (TB), however, it remains limited in preventing latent infection, TB reactivation, and providing comprehensive protection. In this study, the <em>pcaA</em> gene, an antigen associated with persistent infection, was selected, and the recombinant plasmid pET28a-PcaA was successfully constructed for protein expression and purification. The specificity of the antigen was further verified using chemiluminescence, enzyme-linked immunosorbent assay (ELISA), flow cytometry, and mycobacterial growth inhibition assay (MGIA). Significant differences were observed in the expression levels of IFN-γ, IL-2, IL-8, and IgG in the peripheral blood of patients with <em>Mycobacterium tuberculosis</em> (<em>M. tb</em>) following stimulation with the PcaA antigen in the Active Tuberculosis (ATB) and Latent Tuberculosis Infection (LTBI) groups. An IL-8 combined diagnostic model could effectively distinguish between ATB and LTBI, while anti-PcaA IgG demonstrated strong performance in ruling out <em>M. tb</em> infection. Recombinant PcaA protein (rPcaA) was formulated with liposome dimethyl dioctadecylammonium bromide (DDA) / colloidal manganese salt (MnJ)/DM to immunize mice. Serum-specific antibody levels, cytokines secreted by splenocytes, and the number of multifunctional T cells in splenocytes were assessed. The results indicated that the BCG + rPcaA-DM vaccine group exhibited significantly elevated levels of Th1-type cytokines, antibody titers, and the frequencies of IFN-γ⁺/TNF-α⁺ single and double-positive CD4⁺ and CD8⁺ T cells compared to the BCG group. Furthermore, splenocytes and lung cells from immunized mice significantly inhibited mycobacterial growth. These findings suggest that the rPcaA-DM vaccine, as a BCG booster, significantly enhances Th1 polarization and provides robust protective efficacy, with potential to prevent LTBI progression to ATB.</div></div>","PeriodicalId":16409,"journal":{"name":"Journal of microbiological methods","volume":"236 ","pages":"Article 107202"},"PeriodicalIF":1.9,"publicationDate":"2025-07-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144722982","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Improved pre-treatment for lipid staining of microalgae with a rigid cell wall","authors":"Samira Reuscher ,&nbsp;Gerd Klock ,&nbsp;Antje Kersten ,&nbsp;Samuel Schabel ,&nbsp;Rüdiger Graf","doi":"10.1016/j.mimet.2025.107205","DOIUrl":"10.1016/j.mimet.2025.107205","url":null,"abstract":"<div><div>The worldwide depletion of energy and feedstock resources is an important topic of the current decade. Microalgae are an interesting option, offering rapid biomass production in combination with CO₂-fixation. For ongoing research in microalgae applications, well-functioning laboratory methods are important. One major aspect in microalgae-related studies is the measurement of the biomass' lipid content. In order to avoid elaborate extraction protocols, lipid staining with fluorescent dyes such as Nile Red or BODIPY is currently preferred in small-scale screening trials. Although practical, staining was hindered for microalgae with a rigid cell wall. Additional pre-treatment using solvents has already been described, but this was not sufficient for all strains. Besides sonication, we show here that fluorescent staining can be successfully implemented following the disruption of microalgal cell walls by bead beating. The protocol functions with conventional laboratory equipment (ball mill, shaker and vortex mixer), all of them allowing for small-volume samples and processing of multiple samples in parallel. The bead beating method extends the fluorescence lipid staining approach to microalgae with an extremely rigid cell wall and can be applied for screening trials in biofuel or biomaterial related research.</div></div>","PeriodicalId":16409,"journal":{"name":"Journal of microbiological methods","volume":"236 ","pages":"Article 107205"},"PeriodicalIF":1.9,"publicationDate":"2025-07-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144721285","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Site-2 protease-like protein drives organic solvent tolerance in Rhodococcus ruber","authors":"Jinfang Wu,&nbsp;Zhonghao Wu,&nbsp;Ren Peng","doi":"10.1016/j.mimet.2025.107204","DOIUrl":"10.1016/j.mimet.2025.107204","url":null,"abstract":"<div><div>Engineered overexpression of a Site-2 protease-like protein (S2plp) in <em>Rhodococcus ruber</em> SD3 significantly enhanced organic solvent tolerance. Through electro-transformation with the recombinant plasmid pNV18-<em>s2plp</em>-<em>s2plp</em>, we generated a strain exhibiting superior growth under multiple organic solvent stresses. Transcriptomic analysis identified 13 significantly upregulated genes, including those encoding aminoglycoside O-phosphotransferase, replication initiation protein, two M50 family metallopeptidases, transposase, SMC family ATPase, ATP-dependent helicase, three hypothetical proteins, and three small RNAs (sRNAs). KEGG analysis linked S2plp overexpression to enhanced DNA repair mechanisms, directly contributing to organic solvent tolerance. These results establish S2plp as a novel molecular target for boosting solvent resilience in this important bacterium, advancing its potential in biocatalysis and bioremediation.</div></div>","PeriodicalId":16409,"journal":{"name":"Journal of microbiological methods","volume":"236 ","pages":"Article 107204"},"PeriodicalIF":1.7,"publicationDate":"2025-07-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144707790","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Advances in molecular diagnostics of Neisseria gonorrhoeae","authors":"Choo Yee Yu ,&nbsp;Xin Rui Lim ,&nbsp;Thanh Van Phan ,&nbsp;Nurulfiza Mat Isa ,&nbsp;Chan Yean Yean ,&nbsp;Kok-Gan Chan ,&nbsp;Geik Yong Ang","doi":"10.1016/j.mimet.2025.107197","DOIUrl":"10.1016/j.mimet.2025.107197","url":null,"abstract":"<div><div><em>Neisseria gonorrhoeae</em> is a human obligate pathogen that causes the sexually transmitted infection (STI) gonorrhea. As the second most commonly reported STI of bacterial origin, gonorrhea is a growing global public health concern given that the causative pathogen has developed resistance to antibiotics that are currently used for treatment. A holistic approach is thus essential to reduce the incidence of gonorrhea and to control the spread of antimicrobial resistance <em>N. gonorrhoeae</em>. Improvement in diagnostics is one of the avenues to combat rising STI rates and this review aims to provide an overview of the strategies used for <em>N. gonorrhoeae</em> nucleic acid detection by examining nucleic acid amplification tests (NAATs) that are cleared by the United States Food and Drug Administration (FDA). The updated and comprehensive information of FDA-cleared NAATs presented in this review includes the recent clearance of NAATs for point-of-care testing and home-based specimen collection kit for gonorrhea testing that can potentially revolutionize STI services. Further progress in the molecular diagnostics for gonorrhea is anticipated along with vaccine development as the challenge remains to meet the World Health Organization goal of reducing the incidence of <em>N. gonorrhoeae</em> infection by 90 % by 2030.</div></div>","PeriodicalId":16409,"journal":{"name":"Journal of microbiological methods","volume":"236 ","pages":"Article 107197"},"PeriodicalIF":1.7,"publicationDate":"2025-07-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144655788","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Evaluation of nucleic acid extraction methods for recovery of Cyclospora cayetanensis, Salmonella enterica, and murine norovirus from water and sludge","authors":"Amy Kahler ,&nbsp;Jessica Hofstetter ,&nbsp;Camila Rodrigues ,&nbsp;Mia Mattioli","doi":"10.1016/j.mimet.2025.107195","DOIUrl":"10.1016/j.mimet.2025.107195","url":null,"abstract":"<div><div>The coccidian parasite <em>Cyclospora cayetanensis</em> is the causative agent for foodborne outbreaks of cyclosporiasis and multiple fresh produce recalls annually. In recent years, this organism has been reported in the water near produce growing operations during outbreak investigations, prompting a call for more research on its environmental prevalence in the United States. Currently, there is a lack of performance data available on methods for conducting this research, including the performance of DNA extraction methods for molecular testing. Extraction methods for environmental samples must be efficient due to the often-limited amount of target nucleic acid and the potential for molecular inhibitors present in an environmental sample. This study assessed the performance of <em>C. cayetanensis</em> nucleic acid extraction seeded into surface water, produce wash water, and tap water by two methods designed for use with environmental samples: the PowerViral and UNEX methods. The PowerSoil extraction method (2 g) was assessed for <em>C. cayetanensis</em> extraction from seeded sewage sludge – an environmental sample type used to evaluate parasite carriage within communities. Extraction performance of the PowerViral and UNEX methods were also assessed for the detection of the foodborne bacterial pathogen <em>Salmonella</em> and a surrogate for foodborne viruses, murine norovirus (MNV) seeded into surface water, produce wash water, and tap water. The PowerViral method resulted in consistent detection (83–100 %) of <em>C. cayetanensis</em>, <em>S. enterica</em>, and MNV across all water types. Detection rates for the UNEX method ranged from 56 to 100 % prevalence for tap water and wash water, but there were no detections for any microbe from surface water. The PowerSoil method resulted in poor recovery of <em>C. cayetanensis</em> from sludge (≤1 % recovery), while both the PowerViral and UNEX methods effectively recovered <em>C. cayetanensis</em> from sludge (4–36 % recovery).</div></div>","PeriodicalId":16409,"journal":{"name":"Journal of microbiological methods","volume":"236 ","pages":"Article 107195"},"PeriodicalIF":1.7,"publicationDate":"2025-07-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144649631","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Evaluation of a high-throughput method for processing sponge-stick samples to detect viable, non-spore-forming biothreat agents","authors":"Vanessa L. Brisson ,&nbsp;Staci R. Kane ,&nbsp;M. Worth Calfee ,&nbsp;Stephen Cendrowski ,&nbsp;Sanjiv R. Shah","doi":"10.1016/j.mimet.2025.107194","DOIUrl":"10.1016/j.mimet.2025.107194","url":null,"abstract":"<div><div>After a bioterrorism incident, surface sampling is often used to determine the extent of contamination and exposure, guiding decontamination efforts and decisions for re-occupancy of affected sites. The sponge-stick (SS) is a preferred and commonly used device for sample collection to detect both spore-forming and non-spore-forming biothreat agents from non-porous surfaces. Here, a recently developed high-throughput method (HTM) for processing SS samples to detect viable <em>Bacillus anthracis</em> spores was adapted for detection of non-spore-forming biothreat agents, <em>Yersinia pestis</em> and <em>Francisella tularensis</em>. The scalable HTM was used to process up to 20 SS samples simultaneously, compared to the current stomacher-based method which processes one SS at a time. Comparisons of the HTM and the stomacher-based method were statistically indistinguishable for most experiments (<em>P</em> &gt; 0.05) with HTM recoveries of 37–60 % for <em>Y. pestis</em> inoculated at 10²–10³ cells/SS and held 48 h at 4 °C to mimic sample transport/storage. The HTM was integrated with Rapid Viability-Polymerase Chain Reaction (RV-PCR) analysis to detect viable <em>Y. pestis</em> in the presence of particulate contamination (Arizona Test Dust, ATD). This approach detected <em>Y. pestis</em> inoculated at 20 cells/SS and ATD did not impact detection (<em>P</em> &gt; 0.05). <em>F. tularensis</em> showed significantly lower recoveries between no-hold time and 48-h hold time (4 °C, <em>P</em> &lt; 0.05) using the HTM, which further testing showed could be due to toxicity of the neutralizing buffer used for SS pre-wetting. With modifications, this method could enhance throughput capacity while maintaining similar recovery efficiencies to current methods for other non-spore-forming bacterial pathogens.</div></div>","PeriodicalId":16409,"journal":{"name":"Journal of microbiological methods","volume":"236 ","pages":"Article 107194"},"PeriodicalIF":1.7,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144626574","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Validation of pleural fluid group A Streptococcus and Staphylococcus aureus PCR assays and their potential clinical impact in children with complicated pneumonia","authors":"Erin C. Ho ,&nbsp;Molly Butler ,&nbsp;Kaitlin E. Olson ,&nbsp;Dennis Simmons ,&nbsp;Colsen Beveridge ,&nbsp;Kristen Miller ,&nbsp;Meghan Birkholz ,&nbsp;Sarah A. Jung ,&nbsp;Samuel R. Dominguez","doi":"10.1016/j.mimet.2025.107192","DOIUrl":"10.1016/j.mimet.2025.107192","url":null,"abstract":"<div><h3>Background</h3><div>Better diagnostic tools are needed to increase the yield and speed of pathogen identification in pediatric complicated community-acquired pneumonia (cCAP), and thereby, improve patient care through optimization of antibiotic therapy.</div></div><div><h3>Methods</h3><div>We performed analytical and clinical validations of a laboratory-developed group A <em>Streptococcus</em> (GAS) PCR and a commercially available <em>Staphylococcus aureus</em> (<em>S. aureus</em>), including methicillin-resistant <em>S. aureus</em> (MRSA), PCR for pleural fluid specimens and studied the potential clinical impact of pleural fluid GAS and <em>S. aureus</em> PCR testing in children with cCAP.</div></div><div><h3>Results</h3><div>Both assays demonstrated high analytical sensitivity, specificity, reproducibility, and accuracy in detection of their target pathogens in pleural fluid. In potential clinical impact analysis for 62 children with cCAP requiring pleural fluid drainage, the addition of GAS and <em>S. aureus</em> PCR testing to existing diagnostic testing increased pathogen yield from 71.0 % to 83.2 % (<em>p</em> = 0.023), decreased theoretical median time from hospitalization to optimal therapy from 5.1 days (95 % CI: 2.4–7.2) to 3.7 days (95 % CI: 1.9–6.9, <em>p</em> = 0.001) and theoretical median time from start to stop of unwarranted MRSA therapy from 1.5 days (95 % CI: 1.0–3.7) to 0.8 days (95 % CI: 0.5–1.2, <em>p</em> &lt; 0.001).</div></div><div><h3>Conclusion</h3><div>PCR testing is a sensitive and specific method for detecting GAS and <em>S. aureus</em> in pleural fluid. Clinical implementation of these targeted pleural fluid PCR assays has the potential to significantly increase pathogen yield, facilitate faster de-escalation of MRSA therapy and transition to narrow, targeted antibiotics, suggesting their utility for pediatric complicated pneumonia.</div></div>","PeriodicalId":16409,"journal":{"name":"Journal of microbiological methods","volume":"236 ","pages":"Article 107192"},"PeriodicalIF":1.7,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144626589","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}