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Detection and differentiation of Burkholderia species with pathogenic potential in environmental soil samples
PLOS ONE ( IF 3.7 ) Pub Date : 2021-01-07 , DOI: 10.1371/journal.pone.0245175
Sujintana Janesomboon , Veerachat Muangsombut , Varintip Srinon , Chatruthai Meethai , Chayada S. Tharinjaroen , Premjit Amornchai , Patoo Withatanung , Narisara Chantratita , Mark Mayo , Vanaporn Wuthiekanun , Bart J. Currie , Joanne M. Stevens , Sunee Korbsrisate

The Burkholderia pseudomallei phylogenetic cluster includes B. pseudomallei, B. mallei, B. thailandensis, B. oklahomensis, B. humptydooensis and B. singularis. Regarded as the only pathogenic members of this group, B. pseudomallei and B. mallei cause the diseases melioidosis and glanders, respectively. Additionally, variant strains of B. pseudomallei and B. thailandensis exist that include the geographically restricted B. pseudomallei that express a B. mallei-like BimA protein (BPBM), and B. thailandensis that express a B. pseudomallei-like capsular polysaccharide (BTCV). To establish a PCR-based assay for the detection of pathogenic Burkholderia species or their variants, five PCR primers were designed to amplify species-specific sequences within the bimA (Burkholderia intracellular motility A) gene. Our multiplex PCR assay could distinguish pathogenic B. pseudomallei and BPBM from the non-pathogenic B. thailandensis and the BTCV strains. A second singleplex PCR successfully discriminated the BTCV from B. thailandensis. Apart from B. humptydooensis, specificity testing against other Burkholderia spp., as well as other Gram-negative and Gram-positive bacteria produced a negative result. The detection limit of the multiplex PCR in soil samples artificially spiked with known quantities of B. pseudomallei and B. thailandensis were 5 and 6 CFU/g soil, respectively. Furthermore, comparison between standard bacterial culture and the multiplex PCR to detect B. pseudomallei from 34 soil samples, collected from an endemic area of melioidosis, showed high sensitivity and specificity. This robust, sensitive, and specific PCR assay will be a useful tool for epidemiological study of B. pseudomallei and closely related members with pathogenic potential in soil.



中文翻译:

环境土壤样​​品中具有致病潜力的伯克霍尔德氏菌物种的检测和分化

类鼻疽伯克霍尔德菌系统发生簇包括假苹果花Bmalleithailandensisoklahomensishumptydooensis奇异的。视为该组中,唯一的成员致病假单胞菌Mallei分别引起类鼻疽和腺体疾病。另外,B的变异株。假苹果thailandensis存在包括地理上受限。表示B的假马来酸mallei样比马蛋白(BPBM),和。表示B的thailandensis假性马来样荚膜多糖(BTCV)。为了建立用于检测病原性的基于PCR的测定法伯克霍尔德物种或它们的变体,五个设计PCR引物以扩增种特异性序列的内比马urkholderiantracellularotility A)基因。我们的多重PCR方法可以区分致病。来自非致病性B的假芽孢杆菌和BPBM 。thailandensis和BTCV株。第二次单重PCR成功地将BTCV与B区别开来。thailandensis。除了humptydooensis,针对其他伯克霍尔德氏菌以及其他革兰氏阴性和革兰氏阳性细菌的特异性测试均产生阴性结果。用已知量的B人工掺加的土壤样品中多重PCR的检测限。假苹果粒Bthailandensis土壤分别为5和6 CFU / g土壤。此外,比较标准细菌培养和多重PCR检测B的能力。从类me虫病的流行地区收集的34个土壤样本中的假苹果花,具有很高的敏感性和特异性。这种健壮,灵敏,特异的PCR检测方法将成为B流行病学研究的有用工具。在土壤中具有潜在致病性的假苹果和与其密切相关的成员。

更新日期:2021-01-07
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