Research on drug resistant genes and genotypes of carbapenem-resistant Klebsiella pneumoniae / 中华检验医学杂志

Objective To investigate the drug resistant genes and genotypes of carbapenem-resistant Klebsiella pneumoniae in Tianjin First Center Hospital. Methods A total of 52 strains of carbapenem-non-susceptible Klebsiella pneumoniae were collected from 2012 to 2015. The MICs of antimicrobial drugs were detected using agar dilution methods. Phenotypes of carbapenemases were screened using modified Hodge test. Drug resistant genes were detected by multiplex-PCR assay. Multilocus sequence typing ( MLST) was used to determine the genotypes and homology of these carbapenem-resistant Klebsiella pneumoniae strains. Results Susceptibility of antimicrobial agents indicated that all these strains with multiple drug resistance. The resistance rate to piperacillin/tazobactam, ceftriaxone, ceftazidime, cefepime, aztreonam imipenem,meropenem was 100%( 52/52 ) . The resistance rate of ST11 type to amikacin was 93. 5%( 43/46), ciprofloxacin was 97. 8%(45/46), levofloxacin was 97. 8%(45/46), compound sulfamethoxazole was 17. 4%(8/46), tigecycline was 0. The resistance rate of ST101 type to amikacin was 3/3, ciprofloxacin was 2/3, levofloxacin was 3/3, compound sulfamethoxazole was 3/3, tigecycline was 0. The resistance rate of ST709 type to amikacin was 1/1, ciprofloxacin was 1/1, levofloxacin was 1/1, compound sulfamethoxazole was 1/1, tigecycline was 0. The resistance rate of ST1393 type to amikacin was 1/1, ciprofloxacin was 1/1, levofloxacin was 1/1, compound sulfamethoxazole was 1/1, tigecycline was 0. The resistance rate of ST2068 type to amikacin was 1/1, ciprofloxacin was 1/1, levofloxacin was 1/1, compound sulfamethoxazole was 1/1, tigecycline was 0. PCR results showed that 43 isolates were blaKPC-2 positive and 5 isolates were blaOXA-48 positive, 1 isolate was blaDNM-1 positive. There were 46 strains of ST11 type. The 43 strains of Klebsiella pneumoniae producing KPC-2 type carbapenemase were all ST11. While among 5 strains of Klebsiella pneumoniae carrying OXA-48 carbapenem resistant gene, 3 strains were ST101, 1 was ST709, 1 was ST1393. One strain of Klebsiella pneumoniae harboring DNM-1 type carbapenemase was ST2068. Conclusions Drug resistant genes of carbapenem-resistant Klebsiella pneumoniae were KPC-2 dominant, OXA-48 and DNM-1 were sporadical;the genotype was mainly ST11 by MLST in the hospital. The research provided effective basic and reference for the hospital infection t control.

Identification of m/z characteristics of Staphylococcus aureus in agr and sigB regulatory system using MALDI-TOF-MS / 中华检验医学杂志

Objective The expression of agr and sigB regulation system in Staphylococcus aureus with different infection types were assessed by analyzing the characteristics of m /z value generated by MALDI-TOF-MS.Methods A total of 50 isolates with specific genotypes were collected from Tianjin First Center Hospital during Jun 2013 to Feb 2014 for retrospective study .The pattern profiles of these isolates were obtained by MALDI-TOF-MS with RUO model, and the m/z value was also analysed to evaluate the expression the agr and sigB regulation system .The phylogenetic tree based on mass spectrum peak feature was constructed using SARAMIS software .Results A total of 50 strains of Staphylococcus aureus were divided into two groups: acute infection and chronic persistent and recurrent infection .The expression of delta toxin in acute infection and in chronic infection was 99.2 ±4.1 and 60.5 ±10.1 ( t =16.83, P<0.05), respectively.The regulation of stress proteins of sigB system was enhanced in chronic persistent and recurrent infections , and the expression intensities of SAS 030, SAS049 and SA0772 were 27.1 ±14.7, 54.8 ±21.5 and 51.6 ±19.2, respectively; while in acute infections , those were 4.9 ±1.9, 12.4 ±2.8 and 15.7 ±6.9, respectively.The t values between the two groups were -6.88 (P<0.05),-8.98 (P<0.05) and -1.87 (P<0.05), respecitively.The expression of phenol-soluble modulins (PSMs) was inconsistent , and the relative strength of PSMα3 was 100%in the colony variants small strains .Conclusions Different types of the Staphylococcus aureus infections could be evaluated through the assessment of the agr and sigB regulation system .The m/z value obtained by MALDI-TOF mass spectrometry is a marker for the expression of agr and sigB regulation system .The application of this technology needs further development .

Infuening factors and measures of evidence-based nursing on the success rate of venipuncture for children / 现代临床护理

Objective To explore the effect of evidence-based nursing of the success rate of venipuncture for children and summarize the strategies . Method Three hundred and sixty children receiving intravenous therapy were investigated by self-designed questionnaire to explore the influencing factors of success veinpuncture. Results The unsuccessful rate of veinpuncture was 22.2%(88/360). And the main influencing factors included children′factors which accounted for 42.1%, nurses′factors which accounted for 29.4%, parents′factors which accounted for 18.1%and environment factors 10.4%. Conclusion The following strategies can be effective in increasing the success rate of veinpuncture, such as choosing the appropriate veins, improving the veinpuncture skills, creating favorable treatment environment and doing psychological nursing well.

Detection of PSM-mec by Vitek mass spectrometry for rapid identification of nosocomial-acquired methicillin-resistant Staphylococcus aureus / 中华检验医学杂志

Objective To evaluate the method of PSM-mec detection by Vitek MS for nosocomialacquired methicillin-resistant Staphylococcus aureus (MRSA) identification.Methods Totally 167 isolates of MRSA and 100 isolates of methicillin-sensitive Staphylococcus aureus (MSSA) used in this research were non-repetitively and prospectively collected between June 2012 and December 2013,two different SCCmec genotyping methods were applied for the MRSA strains,Vitek MS was used for identification of the isolates,the acquisition mass-spectrogram and the result mass-spectrogram at Myla system were analyzed among the different SCCmec type of MRSA.Results The 167 isolates of MRSA were classified into 5 major SCCmec types,among which SCCmec Ⅰ accounting for 3.6％ (6 isolates);SCCmec Ⅱ 6.0％ (10 isolates);SCCmec Ⅲ and Ⅲa 84.4％ (141 isolates);SCCmec Ⅳand Ⅳ a 4.8％ (8 isolates);SCCmec Ⅴ 1.2％ (2 isolates),respectively.The peak adjacent to the horizontal axis of a m/z 2 500 could be visually identified between the SCCmec Ⅱ and Ⅲ MRSA,of which the delta toxin peak were presented at m/z 3 005-3 009 or m/z 3 037-3 056,while the strains without delta toxin peak and the other types of MRSA or MSSA had no characteristic peak at the same position.Conclusions Nosocomial-acquired MRSA of the drug-resistant condition could be rapidly differentiated and forecasted by Vitek MS.Vitek MS could serve as a routine clinical assistance for epidemiological investigations of nosocomial-acquired MRSA in local area.

The Clumping factor B gene typing in discriminating the strains of nosocomial-acquired methicillin-resistant Staphylococcus aureus / 中华流行病学杂志

Objective To evaluate the clfB typing method in discriminating the ST239 methicillin-resistant Staphylococcus aureus (MRSA) isolated from patients under nosocomial infection in Tianjin first central hospital so as to access the clinical risk factors and outcomes of the MRSA nosocomial infection from ICU and non-ICU departments.Methods Forty-two stains of MRSA with known SCCmec type were chosen in both ICU (n=35) and non-ICU (n=7) wards,from 2006 to 2012,of which MLST genotype was ST239.Clinical risk factors and rates on drug resistant to MRSA were counted,respectively.Results All the isolates of MRSA belonged to the same lineage 3 and 6 heplotypes,based on clfB variable-number random repeats typing.Thirty-five isolates from ICU belonged to 6 heplotypes,among which clfB3-52,3-52E,3-50,3-52C,3-50A and 3-50E were accouted for 42.9％,37.1％,8.6％,5.7％,2.9％ and 2.9％,respectively.Seven isolates from non-ICU belonged to 3 heplotypes,in which 3-52,3-52E and 3-50 were accouted for 42.8％,28.6％,28.6％,respectively.When clfB typing was combined with SCCmec typing in use,results showed that the index of discrimination as 0.767,better than clfB (ID=0.688) or SCCmec (ID=0.303) used alone.SCCmec Ⅲ-clfB3-52E seemed as the major clone among the 10 heplotypes of clfB/SCCmec typing,which was accounted for 40.4％.There were significant differences on the length of hospitalization (P＜0.005) and the duration of antibiotics use (P＜0.05) between ICU and non-ICU.Conclusion The clfB typing method which was based on variable-numbers of tandom repeats showed powerful ability of resolution.It could also be combined with MLST and SCCmec typing to be used in local epidemiological investigations.

The cDNA cloning of human granulysin Mr 15000 and Mr 9000 active segments from CTL activated by allogenic antigen / 重庆医科大学学报

Objective; To clone, sequence and analyze the coding sequences of the Mr 15000 and Mr 9000 active segments of natural granulysin derived from human CTLs activated by allogenic antigen ,in order to establish the basis for further purifying and investigating the immune - impairing mechanism of granulysin. Methods; The coding sequences of the Mr 15000 and Mr 9000 granulysin gene were amplified from the total RNA of activated CTLs of healthy person after reverse transcription by Nested - PCR, inserted into pET32a ( + ) vectors and then transformed into E. Coli TOP10, respectively. The recons were identified by PCR, endonuclease digestion and sequencing. Results: The whole coding sequences of the Mr 15000 and Mr 9000 active segments were successfully cloned as expected. The accurate pET32a - Mr 15000 and pET32a - Mr 9000 recons were obtained through Colony - PCR, endonuclease digestion and sequencing. There lied polymorphism on the 119 th amino acid of the product encoded by GLS gene. Conclusion; The coding sequences of the Mr 15000 and Mr 9000 active segments of human granulysin can be obtained and cloned by the methods mentioned above and can be used for subsequent research.