Study On Carbapenem Resistance Mechanism And Molecular Epidemiology In Klebsiella Pneumoniae

ObjectiveTo investigate carbapenem resistance mechanism, epidemiology, and clinical features of Klebsiella pneumoniae isolated from clinical samples in Xinhua Hospital, Affiliated to Shanghai Jiao Tong University. Provide theoretical foundation for prevention and control of nosocomial infection of CRKP, and offer advices to clinician for rational usage of antibiotic.Methods1. CRKP strains isolated from clinical samples in our hospital during 2009 to 2013 were collected. 2. Modified Hodge Test and Imipenem/Imipenem-EDTA(IP/IPI) E-test was performed for screening of carbapenemase and mental β-lactmases expression phonotype, respectively. 3. PCR and DNA sequencing was used to detect and identify carbapenemase, Amp C β-lactamases and ESBLs genes. 4. Expression of outer membrane porin was determined by SDS-PAGE. 5. Plasmid conjugation experiment was carried out for the carbapenemase producing strains. 6. Collect environment specimen from Surgery Intensive Care Unit(SICU) for CRKP sreening. 7. Relatedness of the strains was determined by MLST and PFGE, and the characteristic of molecular epidemiology of CRKP in our hospital was investigated by analysis of the typing results with clinical information of patients.Results1. A total of 84 CRKP clinical strains were collected during 2009 to 2013. 2. 72 isolates were positive by Modified Hodge Test, and 1 was positive by IP/IPI E-test. 3. PCR and DNA sequencing revealed that the 69 isolates of 72 Modified Hodge Test positive strains carry KPC-2 gene, 2 isolates carry IMP-4 gene and 1 isolate carries NDM-1 gene, all strains above also carry 1~3 types of ESBLs genes. 12 Modified Hodge Test negitive strains carry DHA-1 gene and/or 1~2 types of ESBLs genes. 4. 5 strains express normal Outer Membrane Porin, 3 strains have Ompk36 deficiency, 1 strain has Ompk35 deficiency and the rests express outer membrane porin variant. 5. 4 carbapenemase-producing strains successfully transmited the plasmid to recipient strain. 6. 2 CRKP strains were isolated from environment samples of SICU. 7. MLST showed that the most common type of CRKP in our hospital was ST11(73/84), the next is ST323(3/84). ST29,ST37,ST571,ST1696,ST1721,ST1722,ST1723 and ST1725 were also detected, one strain for each type. The later 5 types were new sequence types discovered in this study. PFGE divided the 84 strains into the following 3 classes: A(A1, A2, A3, and A4), B(B1, B2, B3, and B4), C(C1, C2) and the most prevalent type were B3, and the typing result of PFGE was agreed with MLST. The two environment strains were belonging to type B3 and ST11 as well.ConclusionThe most common carbapenem resistance mechanism of CRKP in our hospital was KPC-2 producing combined with abnormal expression of outer membrane porin. Our hospital had an outbreak of CRKP, mainly caused by Klebsiella pneumoniae ST11(B3 type) clone. Patients who infected with CRKP and the materials him who had touched were the most important infection source. The prevention and control measures for nosocomic infection should be enforced, in order to limit the prevalent range of CRKP.