Rapid detection of Burkholderia pseudomallei with a lateral flow recombinase polymerase amplification assay

• DOI: 10.1371/journal.pone.0213416
• Publication Date: 2019-07-08T17:25:00Z
• Authors: Yao Peng; Xiao Zheng; Biao Kan; Wei Li; Wen Zhang; Taozhen Jiang; Jinxing Lu; Aiping Qin
• Keywords: Burkholderia pseudomallei; Melioidosis; Recombinase Polymerase Amplification

Melioidosis is a severe infectious disease caused by gram-negative, facultative intracellular pathogen Burkholderia pseudomallei (B. pseudomallei). Although cases are increasing reported from other parts of the world, it an illness tropical and subtropical climates primarily found in southeast Asia northern Australia. Because 40% mortality rate, this life-threatening poses public health risk endemic area. Early detection B. infection vital for prognosis melioidosis patient. In study, novel isothermal recombinase polymerase amplification combined with lateral flow dipstick (LF-RPA) assay was established rapid pseudomallei. A set primer-probe targeting orf2 gene within putative type III secretion system (T3SS) cluster genes generated parameters LF-RPA were optimized. Result can be easy visualized 30 minutes limit (LOD) as low 20 femtogram (fg) (ca. 25.6 copies) genomic DNA without specific equipment. The highly no cross observed mallei, members cepacia-complex 35 non-B. bacteria species. Moreover, isolates patients Hainan (N = 19), Guangdong 1), Guangxi 3) province China well Australia Thailand 1) retrospectively confirmed newly developed method. LODs pseudomallei-spiked soil blood samples 2.1×103 CFU/g 4.2×103 CFU/ml respectively. sensitivity comparable to TaqMan Real-Time PCR (TaqMan PCR). addition, exhibited better tolerance inhibitors than PCR. Our results showed that alternative existing PCR-based methods potentiality early accurate diagnosis at point care or in-field use.