Rapid Detection Of Carbapenemase-producing Klebsiella Pneumoniae Based On GCMS

Objective:To explore the application value of mass spectrometry in the detection of carbapenem-resistant Klebsiella pneumoniae(CRKP)which producing carbapenemases.Methods:1.CRKP isolated from blood culture in the Second Affiliated Hospital of Nanchang University from January 2016 to December 2019 were collected.Strain identification and drug sensitivity test were carried out by VITEK-2 compact.Modified carbapenem inactivation method(m CIM)and EDTA modified carbapenem inactivation method(e CIM)were used to detect whether the strain produced carbapenemase and its phenotype.The carbapenemases genes of the strain was amplified by polymerase chain reaction(PCR)and its genotype was determined by bidirectional sequencing.The clinical information of patients was collected by LIS and HIS system,and the clinical characteristics of patients were statistically analyzed.2.Continue to collect and select clinical isolates of CRKP that produce different types of carbapenemases,CRKP that do not produce carbapenemases and carbapenemsensitive Klebsiella pneumoniae(CSKP).Using broth as culture medium,and imipenem was added during the enrichment culture.The gas components in the upper layer of bacterial liquid were detected by headspace-solid phase microextraction-gas chromatography-mass spectrometry(HS-SPME-GCMS),and the data were processed and analyzed by Mass Hunter GC/MS Acquisition software.Results:1.A total of 41 strains of CRKP isolated from blood culture were collected.Among them,40 isolates produced KPC-2 carbapenemase(97.6%,40/41)and one produced OXA-48(2.4%,1/ 41).Clinical data showed that,among the 41 patients,56.1%(23/41)were from ICU;90.2%(37/41)were hospital acquired infection;and 51.2%(21/41)of patients had bloodstream infection which from pulmonary infection;the proportion of patients with Pitt bacteremia score ≥ 4 was 73.2%(30/41);a total of 23 patients died and the mortality was 56.1%(23/41).2.CRKP producing KPC carbapenemase,CRKP producing NDM carbapenemase,CRKP producing IMP carbapenemase,CRKP without producing carbapenemase and CSKP were analyzed by GC-MS.Results showed that in the group of producing KPC and NDM carbapenemase CRKP had characteristic peaks when the retention time(RT)was 6.1min and the mass-to-charge ratio was 41,42,43,55 and 70.After searching and comparing the NIST standard spectrum database,it was found that the biomarker was3-methyl-1-butanol.Conclusions:1.Carbapenem-resistant Klebsiella pneumoniae,which causes bloodstream infection,mainly produces KPC-2 carbapenemase.The patients with bloodstream infection caused by CRKP are mainly from ICU,and whose bacteremia is serious,and the mortality is high.2.Microbial volatile organic compounds(m VOCs)can be used for the rapid identification of carbapenemase-producing CRKP.3-methyl-1-butanol is a biomarker of the CRKP which producing KPC and NDM carbapenemases.