使用商业环介导等温扩增（LAMP）检测法直接从加标的血培养物中检测高毒力肺炎克雷伯菌（hvKp）菌株。
Detection of hypervirulent Klebsiella pneumoniae (hvKp) strains directly from spiked blood cultures using a commercial Loop-Mediated isothermal amplification (LAMP) assay.
作者信息
Fernández Vecilla Domingo, Rodríguez Grande Jorge, Fraile Valcárcel Nuria, Moure García Zaira, García Fernández Sergio, Siller Ruiz María, Roiz Mesones María Pía, Díaz de Tuesta Del Arco José Luis, Urrutikoetxea Gutiérrez Mikel Joseba, Lomoro María Catalina, Fariñas María Carmen, Ocampo-Sosa Alain
机构信息
Microbiology Service, Valdecilla Research Institute (IDIVAL), University Hospital Marqués de Valdecilla, Av. de Valdecilla, s/n, 39008, Santander, Cantabria, Spain.
CIBERINFEC, Health Institute Carlos III, Av. de Monforte de Lemos, 5, Fuencarral-El Pardo, 28029, Madrid, Spain.
出版信息
Ann Clin Microbiol Antimicrob. 2025 Aug 21;24(1):47. doi: 10.1186/s12941-025-00817-4.
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 Eazyplex 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.
背景
高毒力肺炎克雷伯菌（hvKp）菌株的特点是逃避免疫反应和全身播散的能力增强。快速鉴定hvKp菌株对于指导临床管理和实施有效的感染控制措施至关重要。环介导等温扩增（LAMP）检测为检测细菌病原体提供了一种快速且经济高效的方法。本研究评估了Eazyplex hvKp检测法直接从加样血培养物中检测hvKp菌株的性能。
方法
我们于2021年12月至2024年8月从西班牙北部的两家医院收集了20株肺炎克雷伯菌（Kp）分离株。使用PCR和全基因组测序（WGS）进行荚膜血清分型和毒力基因检测。在接种了hvKp分离株的加样血培养物上测试Eazyplex hvKp LAMP检测法。使用Kleborate评分系统评估毒力谱。
结果
17株Kp分离株的Kleborate评分为≥2，表明具有高毒力。LAMP检测法在95个毒力靶点中检测到87个，总体准确率为91.5%。虽然有8个靶基因未被直接检测到，但荧光信号表明在所有情况下均有扩增。该检测法鉴定出16株具有高毒力谱（评分≥3）的菌株，其中14株评分为4或5。基于LAMP的检测法能有效地直接从血培养物中检测hvKp，出结果时间为6分43秒至17分11秒。
结论
Eazyplex hvKp LAMP检测法是一种直接从血培养物中鉴定hvKp菌株的快速有效方法。本研究支持其在临床微生物学中对hvKp感染进行早期检测和流行病学监测的潜在用途。然而，Kleborate评分系统的局限性表明可能需要额外的毒力生物标志物来提高hvKp分类的准确性。
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