Domingo Fernández Vecilla, Jorge Rodríguez Grande, Nuria Fraile Valcárcel, Zaira Moure García, Sergio García Fernández, María Siller Ruiz, María Pía Roiz Mesones, José Luis Díaz de Tuesta Del Arco, Mikel Joseba Urrutikoetxea Gutiérrez, María Catalina Lomoro, María Carmen Fariñas, Alain Ocampo-Sosa
{"title":"利用商业环介导等温扩增（LAMP）法直接从加标血培养物中检测高致病性肺炎克雷伯菌（hvKp）菌株。","authors":"Domingo Fernández Vecilla, Jorge Rodríguez Grande, Nuria Fraile Valcárcel, Zaira Moure García, Sergio García Fernández, María Siller Ruiz, María Pía Roiz Mesones, José Luis Díaz de Tuesta Del Arco, Mikel Joseba Urrutikoetxea Gutiérrez, María Catalina Lomoro, María Carmen Fariñas, Alain Ocampo-Sosa","doi":"10.1186/s12941-025-00817-4","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>Hypervirulent K. pneumoniae (hvKp) strains are characterized by their enhanced ability to evade immune responses and disseminate systemically. Rapid identification of hvKp strains is critical for guiding clinical management and implementing effective infection control measures. Loop-Mediated Isothermal Amplification (LAMP) assays provide a rapid and cost-effective method for detecting bacterial pathogens. This study evaluates the performance of the Eazyplex<sup>®</sup> hvKp assay for the direct detection of hvKp strains from spiked blood cultures.</p><p><strong>Methods: </strong>We collected 20 K. pneumoniae (Kp) isolates between December 2021 and August 2024 from two hospitals in Northern Spain. Capsular serotyping and virulence gene detection were performed using PCR and whole-genome sequencing (WGS). The Eazyplex<sup>®</sup> hvKp LAMP assay was tested on spiked blood cultures inoculated with hvKp isolates. Virulence profiles were assessed using the Kleborate scoring system.</p><p><strong>Results: </strong>Seventeen Kp isolates had a Kleborate score of ≥ 2, suggesting high virulence. The LAMP assay detected 87 out of 95 virulence targets, demonstrating an overall accuracy of 91.5%. Although eight target genes were not directly detected, fluorescence signals indicated amplification in all cases. The assay identified 16 strains with high virulence profiles (score ≥ 3), with 14 strains scoring 4 or 5. The LAMP-based test effectively detected hvKp directly from blood cultures, with time-to-results ranging from 6:43 to 17:11 min.</p><p><strong>Conclussion: </strong>The Eazyplex<sup>R</sup> hvKp LAMP assay is a rapid and effective method for identifying hvKp strains directly from blood cultures. This study supports its potential utility in clinical microbiology for early detection and epidemiological surveillance of hvKp infections. However, limitations in the Kleborate scoring system indicate that additional virulence biomarkers may be needed to improve the accuracy of hvKp classification.</p>","PeriodicalId":8052,"journal":{"name":"Annals of Clinical Microbiology and Antimicrobials","volume":"24 1","pages":"47"},"PeriodicalIF":3.6000,"publicationDate":"2025-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12372381/pdf/","citationCount":"0","resultStr":"{\"title\":\"Detection of hypervirulent Klebsiella pneumoniae (hvKp) strains directly from spiked blood cultures using a commercial Loop-Mediated isothermal amplification (LAMP) assay.\",\"authors\":\"Domingo Fernández Vecilla, Jorge Rodríguez Grande, Nuria Fraile Valcárcel, Zaira Moure García, Sergio García Fernández, María Siller Ruiz, María Pía Roiz Mesones, José Luis Díaz de Tuesta Del Arco, Mikel Joseba Urrutikoetxea Gutiérrez, María Catalina Lomoro, María Carmen Fariñas, Alain Ocampo-Sosa\",\"doi\":\"10.1186/s12941-025-00817-4\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Background: </strong>Hypervirulent K. pneumoniae (hvKp) strains are characterized by their enhanced ability to evade immune responses and disseminate systemically. Rapid identification of hvKp strains is critical for guiding clinical management and implementing effective infection control measures. Loop-Mediated Isothermal Amplification (LAMP) assays provide a rapid and cost-effective method for detecting bacterial pathogens. This study evaluates the performance of the Eazyplex<sup>®</sup> hvKp assay for the direct detection of hvKp strains from spiked blood cultures.</p><p><strong>Methods: </strong>We collected 20 K. pneumoniae (Kp) isolates between December 2021 and August 2024 from two hospitals in Northern Spain. Capsular serotyping and virulence gene detection were performed using PCR and whole-genome sequencing (WGS). The Eazyplex<sup>®</sup> hvKp LAMP assay was tested on spiked blood cultures inoculated with hvKp isolates. Virulence profiles were assessed using the Kleborate scoring system.</p><p><strong>Results: </strong>Seventeen Kp isolates had a Kleborate score of ≥ 2, suggesting high virulence. The LAMP assay detected 87 out of 95 virulence targets, demonstrating an overall accuracy of 91.5%. Although eight target genes were not directly detected, fluorescence signals indicated amplification in all cases. The assay identified 16 strains with high virulence profiles (score ≥ 3), with 14 strains scoring 4 or 5. The LAMP-based test effectively detected hvKp directly from blood cultures, with time-to-results ranging from 6:43 to 17:11 min.</p><p><strong>Conclussion: </strong>The Eazyplex<sup>R</sup> hvKp LAMP assay is a rapid and effective method for identifying hvKp strains directly from blood cultures. This study supports its potential utility in clinical microbiology for early detection and epidemiological surveillance of hvKp infections. However, limitations in the Kleborate scoring system indicate that additional virulence biomarkers may be needed to improve the accuracy of hvKp classification.</p>\",\"PeriodicalId\":8052,\"journal\":{\"name\":\"Annals of Clinical Microbiology and Antimicrobials\",\"volume\":\"24 1\",\"pages\":\"47\"},\"PeriodicalIF\":3.6000,\"publicationDate\":\"2025-08-21\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12372381/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Annals of Clinical Microbiology and Antimicrobials\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.1186/s12941-025-00817-4\",\"RegionNum\":2,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"MICROBIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Annals of Clinical Microbiology and Antimicrobials","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1186/s12941-025-00817-4","RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"MICROBIOLOGY","Score":null,"Total":0}
Detection of hypervirulent Klebsiella pneumoniae (hvKp) strains directly from spiked blood cultures using a commercial Loop-Mediated isothermal amplification (LAMP) assay.
Background: Hypervirulent K. pneumoniae (hvKp) strains are characterized by their enhanced ability to evade immune responses and disseminate systemically. Rapid identification of hvKp strains is critical for guiding clinical management and implementing effective infection control measures. Loop-Mediated Isothermal Amplification (LAMP) assays provide a rapid and cost-effective method for detecting bacterial pathogens. This study evaluates the performance of the Eazyplex® hvKp assay for the direct detection of hvKp strains from spiked blood cultures.
Methods: We collected 20 K. pneumoniae (Kp) isolates between December 2021 and August 2024 from two hospitals in Northern Spain. Capsular serotyping and virulence gene detection were performed using PCR and whole-genome sequencing (WGS). The Eazyplex® hvKp LAMP assay was tested on spiked blood cultures inoculated with hvKp isolates. Virulence profiles were assessed using the Kleborate scoring system.
Results: Seventeen Kp isolates had a Kleborate score of ≥ 2, suggesting high virulence. The LAMP assay detected 87 out of 95 virulence targets, demonstrating an overall accuracy of 91.5%. Although eight target genes were not directly detected, fluorescence signals indicated amplification in all cases. The assay identified 16 strains with high virulence profiles (score ≥ 3), with 14 strains scoring 4 or 5. The LAMP-based test effectively detected hvKp directly from blood cultures, with time-to-results ranging from 6:43 to 17:11 min.
Conclussion: The EazyplexR hvKp LAMP assay is a rapid and effective method for identifying hvKp strains directly from blood cultures. This study supports its potential utility in clinical microbiology for early detection and epidemiological surveillance of hvKp infections. However, limitations in the Kleborate scoring system indicate that additional virulence biomarkers may be needed to improve the accuracy of hvKp classification.
期刊介绍：
Annals of Clinical Microbiology and Antimicrobials considers good quality, novel and international research of more than regional relevance. Research must include epidemiological and/or clinical information about isolates, and the journal covers the clinical microbiology of bacteria, viruses and fungi, as well as antimicrobial treatment of infectious diseases.
Annals of Clinical Microbiology and Antimicrobials is an open access, peer-reviewed journal focusing on information concerning clinical microbiology, infectious diseases and antimicrobials. The management of infectious disease is dependent on correct diagnosis and appropriate antimicrobial treatment, and with this in mind, the journal aims to improve the communication between laboratory and clinical science in the field of clinical microbiology and antimicrobial treatment. Furthermore, the journal has no restrictions on space or access; this ensures that the journal can reach the widest possible audience.
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