Development of New PCR Assay with SYBR Green I for Detection of Mycoplasma, Acholeplasma, and Ureaplasma sp. in Cell Cultures

被引：6

作者：

Krzyszton-Russjan, Jolanta ^{[1
]}

Chudziak, Jakub ^{[2
]}

Bednarek, Malgorzata ^{[2
]}

Anuszewska, Elzbieta Lidia ^{[1
]}

机构：

[1] Natl Med Inst NMI, Dept Biochem & Biopharmaceut, PL-00725 Warsaw, Poland

[2] NMI, Dept Biochem & Biopharmaceut, Chelmska 30-34 Str, PL-00725 Warsaw, Poland

来源：

DIAGNOSTICS | 2021年 / 11卷 / 05期

关键词：

Mycoplasma; Acholeplasma and Ureaplasma sp; detection; qPCR validation; cell cultures; quality control; GUIDELINES MINIMUM INFORMATION; 16S RIBOSOMAL-RNA; TIME; CONTAMINATION; LINES; IDENTIFICATION; COMPLEMENTARY; PUBLICATION; INFECTIONS; REGION;

D O I：

10.3390/diagnostics11050876

中图分类号：

R5 [内科学];

学科分类号：

1002 ; 100201 ;

摘要：

Mycoplasma, Acholeplasma, and Ureaplasma sp. are atypical bacteria responsible for in vitro cell culture contaminations that can warp the results. These bacteria also cause human and animal infections and may lead to chronic diseases. In developed polymerase chain reaction (PCR) in this study a quantitative PCR with SYBR Green I fluorochrome was applied to facilitate the Mycoplasma, Acholeplasma, and Ureaplasma sp. DNA detection and identification. Screening Test-1 v.1 (triplex qPCR) allowed for the detection of 11 species. Test-1 v.2 (three single qPCRs) pre-identified three subgroups, allowing for the reduction of using single qPCRs in Test-2 for species identification. The range of both tests was consistent with pharmacopeial requirements for microbial quality control of mammal cells and included detection of M. arginini, M. orale, M. hyorhinis, M. fermentans, M. genitalium, M. hominis, M. pneumoniae, M. salivarium, M. pirum, A. laidlawii, and U. urealyticum. Limit of detection values varied between 125-300 and 50-100 number of copies per milliliter in Test-1 and Test-2, respectively. Test-1 and Test-2 showed fully concordant results, allowed for time-saving detection and/or identification of selected species from Mycoplasma, Acholeplasma, and Ureaplasma in tested cell cultures.

引用

页数：22

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