AUTHOR=Feng Yanling , Xue Guanhua , Feng Junxia , Yan Chao , Cui Jinghua , Gan Lin , Zhang Rui , Zhao Hanqin , Xu Wenjian , Li Nannan , Liu Shiyu , Du Shuheng , Zhang Weiwei , Yao Hailan , Tai Jun , Ma Lijuan , Zhang Ting , Qu Dong , Wei Yongxiang , Yuan Jing 

TITLE=Rapid Detection of New Delhi Metallo-β-Lactamase Gene Using Recombinase-Aided Amplification Directly on Clinical Samples From Children

JOURNAL=Frontiers in Microbiology

VOLUME=12

YEAR=2021

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2021.691289

DOI=10.3389/fmicb.2021.691289

ISSN=1664-302X

ABSTRACT=<p>New Delhi metallo-β-lactamase, a metallo-β-lactamase carbapenemase type, mediates resistance to most β-lactam antibiotics including penicillins, cephalosporins, and carbapenems. Therefore, it is important to detect <italic>bla</italic><sub>NDM</sub> genes in children’s clinical samples as quickly as possible and analyze their characteristics. Here, a recombinase-aided amplification (RAA) assay, which operates in a single one-step reaction tube at 39°C in 5−15 min, was established to target <italic>bla</italic><sub>NDM</sub> genes in children’s clinical samples. The analytical sensitivity of the RAA assay was 20 copies, and the various bacterial types without <italic>bla</italic><sub>NDM</sub> genes did not amplify. This method was used to detect <italic>bla</italic><sub>NDM</sub> genes in 112 children’s stool samples, 10 of which were tested positive by both RAA and standard PCR. To further investigate the characteristics of carbapenem-resistant bacteria carrying <italic>bla</italic><sub>NDM</sub> in children, 15 carbapenem-resistant bacteria (<italic>Escherichia coli</italic>, <italic>Klebsiella pneumoniae</italic>, <italic>Acinetobacter baumannii</italic>, <italic>Citrobacter freundii</italic>, <italic>Klebsiella oxytoca</italic>, <italic>Acinetobacter junii</italic>, and <italic>Proteus mirabilis</italic>) were isolated from the 10 samples. Notably, more than one bacterial type was isolated from three samples. Most of these isolates were resistant to cephalosporins, cefoperazone-sulbactam, piperacillin-tazobactam, ticarcillin-clavulanic acid, aztreonam, co-trimoxazole, and carbapenems. <italic>bla</italic><sub>NDM</sub><sub>–</sub><sub>1</sub> and <italic>bla</italic><sub>NDM</sub><sub>–</sub><sub>5</sub> were the two main types in these samples. These data show that the RAA assay has potential to be a sensitive and rapid <italic>bla</italic><sub>NDM</sub> gene screening test for clinical samples. The common existence of <italic>bla</italic><sub>NDM</sub> and multi-drug resistance genes presents major challenges for pediatric treatment.</p>