Knappik Michael, Dance David A B, Rattanavong Sayaphet, Pierret Alain, Ribolzi Olivier, Davong Viengmon, Silisouk Joy, Vongsouvath Manivanh, Newton Paul N, Dittrich Sabine
Lao-Oxford-Mahosot Hospital-Wellcome Trust Research Unit, Microbiology Laboratory, Mahosot Hospital, Vientiane, Lao People's Democratic Republic Centre for Tropical Medicine and Global Health, Nuffield Department of Medicine, University of Oxford, Oxford, England, United Kingdom.
Lao-Oxford-Mahosot Hospital-Wellcome Trust Research Unit, Microbiology Laboratory, Mahosot Hospital, Vientiane, Lao People's Democratic Republic.
Appl Environ Microbiol. 2015 Jun;81(11):3722-7. doi: 10.1128/AEM.04204-14. Epub 2015 Mar 27.
Burkholderia pseudomallei is the cause of melioidosis, a severe and potentially fatal disease of humans and animals. It is endemic in northern Australia and Southeast Asia and is found in soil and surface water. The environmental distribution of B. pseudomallei worldwide and within countries where it is endemic, such as the Lao People's Democratic Republic (Laos), remains unclear. However, this knowledge is important to our understanding of the ecology and epidemiology of B. pseudomallei and to facilitate public health interventions. Sensitive and specific methods to detect B. pseudomallei in environmental samples are therefore needed. The aim of this study was to compare molecular and culture-based methods for the detection of B. pseudomallei in soil and surface water in order to identify the optimal approach for future environmental studies in Laos. Molecular detection by quantitative real-time PCR (qPCR) was attempted after DNA extraction directly from soil or water samples or after an overnight enrichment step. The positivity rates obtained by qPCR were compared to those obtained by different culture techniques. The rate of detection from soil samples by qPCR following culture enrichment was significantly higher (84/100) than that by individual culture methods and all culture methods combined (44/100; P < 0.001). Similarly, qPCR following enrichment was the most sensitive method for filtered river water compared with the sensitivity of the individual methods and all individual methods combined. In conclusion, molecular detection following an enrichment step has proven to be a sensitive and reliable approach for B. pseudomallei detection in Lao environmental samples and is recommended as the preferred method for future surveys.
类鼻疽伯克霍尔德菌是类鼻疽病的病原体，这是一种严重且可能致命的人畜共患病。它在澳大利亚北部和东南亚为地方性流行，存在于土壤和地表水中。类鼻疽伯克霍尔德菌在全球范围内以及在其地方性流行的国家（如老挝人民民主共和国）内的环境分布情况仍不清楚。然而，这些信息对于我们理解类鼻疽伯克霍尔德菌的生态学和流行病学以及推动公共卫生干预措施至关重要。因此，需要灵敏且特异的方法来检测环境样本中的类鼻疽伯克霍尔德菌。本研究的目的是比较分子方法和基于培养的方法在检测土壤和地表水中类鼻疽伯克霍尔德菌方面的效果，以便确定老挝未来环境研究的最佳方法。在直接从土壤或水样中提取DNA后或经过过夜富集步骤后，尝试通过定量实时PCR（qPCR）进行分子检测。将qPCR获得的阳性率与通过不同培养技术获得的阳性率进行比较。培养富集后通过qPCR从土壤样本中的检出率（84/100）显著高于单独培养方法以及所有培养方法合并后的检出率（44/100；P < 0.001）。同样，与单独方法以及所有单独方法合并后的灵敏度相比，富集后的qPCR是检测过滤后河水最灵敏的方法。总之，富集步骤后的分子检测已被证明是检测老挝环境样本中类鼻疽伯克霍尔德菌的一种灵敏且可靠的方法，建议将其作为未来调查的首选方法。
