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IR@PKUHSC  > 北京大学第一临床医学院  > 感染疾病科  > 期刊论文
学科主题: 临床医学
题名:
Rapid, simple, and sensitive detection of the ompB gene of spotted fever group rickettsiae by loop-mediated isothermal amplification
作者: Pan, Lei1; Zhang, Lijuan1; Wang, Guiqiang2; Liu, Qinghui3
关键词: Rickettsia ; LAMP assay ; Molecular detection ; Spotted fever ; Clinical microbiology
刊名: BMC INFECTIOUS DISEASES
发表日期: 2012-10-11
DOI: 10.1186/1471-2334-12-254
卷: 12
收录类别: SCI
文章类型: Article
WOS标题词: Science & Technology
类目[WOS]: Infectious Diseases
研究领域[WOS]: Infectious Diseases
关键词[WOS]: BORNE BACTERIAL DISEASES ; ORIENTIA-TSUTSUGAMUSHI ; LAMP ; DNA ; IDENTIFICATION ; DIAGNOSIS ; EUROPE
英文摘要:

Background: Spotted fever caused spotted fever group rickettsiae (SFGR) is prevalent throughout China. In this study, we describe a rapid, simple, and sensitive loop-mediated isothermal amplification (LAMP) assay targeting the ompB gene of spotted fever group rickettsiae ideal for application in China. The LAMP assay has the potential to detect spotted fever group rickettsiae early in infection and could therefore serve as an alternative to existing methods.

Methods: A set of universal primers which are specific 7 common species of spotted fever group rickettsiae in China were designed using PrimerExplorer V4 software based on conserved sequences of ompB gene. The sensitivity, specificity and reproducibility of the LAMP were evaluated. The LAMP assay for detecting SFGR was compared with conventional PCR assays for sensitivity and specificity in early phase blood samples obtained from 11 infected human subjects.

Results: The sensitivity of the LAMP assay was five copies per reaction (25 mu L total volume), and the assay did not detect false-positive amplification across 42 strains of 27 members of the order Rickettsiales and 17 common clinical pathogens. The LAMP assay was negative to typhus group rickettsiae including R. prowazekii and R. typhi for no available conserved sequences of ompB was obtained for designing primers. To evaluate the clinical applicability of the LAMP assay, a total of 11 clinical samples, 10 samples confirmed serologically (3 cases), ecologically (1 case), by real-time polymerase chain reaction (PCR; 2 cases), ecologically and by real-time PCR (1 case), and serologically and by real-time PCR (3 cases) were analyzed by the ompB LAMP assay. Data were validated using a previously established nested PCR protocol and real-time PCR. A positive LAMP result was obtained for 8 of the 10 confirmed cases (sensitivity, 73%; specificity, 100%), while none of these samples were positive by nested PCR (sensitivity, 0%; specificity, 100%).

Conclusions: The LAMP assay described here is the most reliable among the three methods tested and would be an ideal choice for development as a rapid and cost-effective means of detecting SFGR in China.

语种: 英语
所属项目编号: 2010CB530200 /2010CB530206 ; 30771854 ; 1U2GGH000018-01
项目资助者: National Basic Research Program of China (973 Program) ; National Natural Science Foundation of China ; China-U.S. Collaborative Program on Emerging and Re-emerging Infectious Diseases
WOS记录号: WOS:000312776700001
Citation statistics:
内容类型: 期刊论文
URI标识: http://ir.bjmu.edu.cn/handle/400002259/55625
Appears in Collections:北京大学第一临床医学院_感染疾病科_期刊论文

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作者单位: 1.Laizhou Peoples Hosp, Laizhou 261400, Shandong, Peoples R China
2.China CDC, Natl Inst Communicable Dis Control & Prevent, Dept Rickettsiol, Beijing 102206, Peoples R China
3.Peking Univ, Hosp 1, Dept Infect Dis, Beijing 100034, Peoples R China

Recommended Citation:
Pan, Lei,Zhang, Lijuan,Wang, Guiqiang,et al. Rapid, simple, and sensitive detection of the ompB gene of spotted fever group rickettsiae by loop-mediated isothermal amplification[J]. BMC INFECTIOUS DISEASES,2012,12.
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