Molecular Epidemic Characteristics And Preliminary Exploration On The Mechanism Of Evolutionary And Transmission In CR-hvKP

Object:To study the clinical and antimicrobial resistance rates characteristics of carbapenem-resistant Klebsiella pneumoniae（CRKP）and carbapenem-resistant hypervirulent Klebsiella pneumoniae（CR-hvKP）and analyze their molecular and virulence features;To preliminarily investigate the evolutionary mechanism of CR-hvKP.Methods:Non-duplicative of clinical CRKP strains and their clinical data were collected from Sichuan province hospital between August 2018 and June 2022.CR-hvKP strains were screened using string test and PCR.Microbroth dilution method was performed to study the in vitro antibacterial activity of CRKP and CR-hvKP.We used PCR test to analyze the molecular characteristics of CRKP and CR-hvKP by amplify their resistance genes,wzi and housekeeping genes.The virulence featuress of CR-hvKP strains were studied using the serum killing test and Galleria mellonella infection model.The horizontal transferability of plasmids was studied by plasmid conjugation test.We used electroshock conversion test to explore the possible evolutionary mechanisms.Results:A total of 114 CRKP and 40 CR-hvKP strains were collected.The resistance levels of CRKP and CR-hvKP to tigacycline（TGC）,colistin（COL）and polycolistin（POL）were less than 10.5%.The resistance rates of CRKP and CR-hvKP to eravacycline（ERV）were 23.7%and 62.5%,respectively,the resistance rates to ceftazidime-avibactam（CZA）were 36.0%and 22.5%,respectively and the tesistance rates to meropenem-vaborbactam（MEV）were at a high level（62.5%and 46.5%,respectively）.The resistance rates to imipenem-relebactam（IMR）were 7.5%and 28.1%,respectively and the resistance rates to aztreonam-avibactam（AZA）were 10.5%and 25.0%,respectively.There were significant differences in the neutrophils count（N）and percentage of neutrophils（N%）between the two groups.Multilocus sequence typing（MLST）showed that sequence type（ST）11 was the predominant type in two groups,accounting for 58.8%and 80.0%,respectively;K serotype was dominated by K64 serotype,accounting for 49.1%and 82.5%,respectively;Carbapenemase genes were dominated by bla _KPC-2 and bla _NDM-1.After conjugation and transfer between 36 CR-hvKP strains and E.coli J53,14（42.4%）E.coli J53 carried carbapenemase genes.Electric shock transformation test showed that ST23-K1 hvKP strain could obtain the drug resistance gene,on the contrary,ST11-K64CRKP strain could also obtain the virulence gene,but there were differences among different strains.Conclusion:CRKP and CR-hvKP were sensitive to only a few antibacterial agents.In our hospital,ST11-K64 type CR-hvKP was prevalent.They mainly carried bla _KPC-2and bla _NDM-1genes.CR-hvKP could be evolved from ST23-K1 hvKP and ST11-K64 CRKP.