AUTHOR=Dong Derong , Liu Wei , Li Huan , Wang Yufei , Li Xinran , Zou Dayang , Yang Zhan , Huang Simo , Zhou Dongsheng , Huang Liuyu , Yuan Jing 

TITLE=Survey and rapid detection of Klebsiella pneumoniae in clinical samples targeting the rcsA gene in Beijing, China

JOURNAL=Frontiers in Microbiology

VOLUME=6

YEAR=2015

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2015.00519

DOI=10.3389/fmicb.2015.00519

ISSN=1664-302X

ABSTRACT=<p><italic>Klebsiella pneumoniae</italic> is a wide-spread nosocomial pathogen. A rapid and sensitive molecular method for the detection of <italic>K. pneumoniae</italic> in clinical samples is needed to guide therapeutic treatment. In this study, we first described a loop-mediated isothermal amplification (LAMP) method for the rapid detection of capsular polysaccharide synthesis regulating gene <italic>rcsA</italic> from <italic>K. pneumoniae</italic>in clinical samples by using two methods including real-time turbidity monitoring and fluorescence detection to assess the reaction. Then dissemination of <italic>K. pneumoniae</italic> strains was investigated from ICU patients in three top hospitals in Beijing, China. The results showed that the detection limit of the LAMP method was 0.115 pg/μl DNA within 60 min under isothermal conditions (61°C), a 100-fold increase in sensitivity compared with conventional PCR. All 30 non- <italic>K. pneumoniae</italic> strains tested were negative for LAMP detection, indicating the high specificity of the LAMP reaction. To evaluate the application of the LAMP assay to clinical diagnosis, of 110 clinical sputum samples collected from ICU patients with clinically suspected multi-resistant infections in China, a total of 32 <italic>K. pneumoniae</italic> isolates were identified for LAMP-based surveillance of <italic>rcs</italic>A. All isolates belonged to nine different <italic>K. pneumoniae</italic> multilocus sequence typing (MLST) groups. Strikingly, of the 32 <italic>K. pneumoniae</italic> strains, 18 contained the <italic>Klebsiella pneumoniae</italic> Carbapenemase (KPC)-encoding gene <italic>bla</italic><sub>KPC-2</sub> and had high resistance to β-lactam antibiotics. Moreover, <italic>K. pneumoniae</italic> WJ-64 was discovered to contain <italic>bla</italic><sub>KPC-2</sub> and <italic>bla</italic><sub>NDM-1</sub>genes simultaneously in the isolate. Our data showed the high prevalence of <italic>bla</italic><sub>KPC-2</sub> among <italic>K. pneumoniae</italic> and co-occurrence of many resistant genes in the clinical strains signal a rapid and continuing evolution of <italic>K. pneumoniae</italic>. In conclusion, we have developed a rapid and sensitive visual <italic>K. pneumoniae</italic> detection LAMP assay, which could be a useful tool for clinical screening, on-site diagnosis and primary quarantine purposes.</p>