Objective To investigate the molecular types and related clinical features of methicillin-resistant Staphylococcus aureus (MRSA) in Jingzhou area, Hubei Province.Methods A total of 80 MRSA strains confirmed by mecA gene were isolated from inpatients in Jingzhou Central Hospital of Hubei province during January and December 2014. Vitek 2 Compact was used for antibiotic susceptibility test . Staphylococcus protein A (SPA) types and Staphylococcal cassette chromosome mec (SCCmec) genotypes were detected by multiplex polymerase chain reaction ( PCR ) and gene sequencing . Panton-valentine leucocidin ( pvl) gene of the strains was detected by PCR .Chi-square test and Wilcoxon test were used for data analysis .Results There were 16 spa types in 80 MRSA isolates , in which t030 and t437 were the most prevalent ones accounting for 50.0% ( 40 strains ) and 28.8% ( 23 strains ) of the total strains, respectively.There were 77 strains of SCCmec type Ⅰ-Ⅴ, in which SCCmecⅢ and SCCmecⅣ were the most prevalent ones accounting for 45.0% (36 strains) and 35.0% (28 strains), respectively.t030 was the main spa type in isolates of SCCmecⅢ(33/36, 91.7%), while t437 was the main spa type in isolates of SCCmecⅣ(20/28, 71.4%).Patients infected with t030/SCCmecⅢMRSAs were with higher ages than those infected with t437/SCCmecⅣMRSAs (T=446.500 and 607.500, P<0.01).Patients infected with t030/SCCmecⅢ MRSAs were mainly from surgical wards and intensive care unit ( ICU ) , while those infected with t437/SCCmecⅣ MRSAs were mainly from pediatrics wards , and there were significant differences in ward distribution between two groups (χ2 =33.724 and 29.768, P <0.01).Seventy percent and above strains of t030/SCCmec type Ⅲ were resistant to rifampin, erythromycin, clindamycin, tetracycline, levofloxacin, moxifloxacin, ciprofloxacin and gentamicin .Strains of t437/SCCmec type Ⅳwere resistant to erythromycin , clindamycin and tetracycline , but were sensitive to most non-β-lactam antimicrobial drugs (with resistance rates <20%).Virulence gene pvl was found in 11 strains (13.8%), in which 7 were strains of t437-SCCmec typeⅣ.Conclusions MRSAs in Jinzhou are of various genotypes , in which t030-SCCmecⅢand t437-SCCmecⅣare the most prevalent ones .Strains of t030-SCCmec typeⅢare usually multiple-drug resistant , mainly seen in elderly patients in surgical wards and ICU .Strains of t437-SCCmecⅣare sensitive to most non-β-lactam antimicrobial drugs , and its infection is mainly seen in children and young people .

Objective To investigate the epidemiology and antibiotic resistance of communityassociated and hospital-associated meticillin-resistant Staphylococcus aureus (CA-MRSA and HA-MRSA) in Jingzhou.Methods A total of 159 MRSA isolates were successively collected from patients in Jingzhou Central Hospital during January 2012 and December 2013.The minimum inhibitory concentrations of 16 antimicrobial agents against 159 MRSA isolates were detected.SCCmec types of the strains were detected by multiplex PCR,and the homology of the strains was analyzed using pulsed field gel electrophoresis (PFGE) and cluster analysis of antibiogram.WHONET 5.6 and SPSS 19.0 were used for data analysis.Results Among 159 MRSA strains,131 were hospital-associated,and 28 were community-associated,which accounted for 82.4％ and 17.6％,respectively.There were significant differences in the age of patients,ward distribution,specimen type,length of stay,length of anti-infection treatment,type of infection and underlying diseases between patients with CA-MRSA or HA-MRSA infections (x2 =19.103,31.372,59.756,71.703,54.153,59.756 and 54.232,all P ＜ 0.01).No vancomycin,linezolid,tigecyeline and nitrofurantoin resistant strains were found,but all strains were resistant to penicillin,cefoxitin and oxacillin.HA-MRSA had higher resistance rates to moxifloxacin,levofloxacin,rifampicin,ciprofloxacin and gentamicin than CA-MRSA (x2 =30.179,27.352,28.523,28.523 and 25.987,all P ＜ 0.01),but its resistance rates to erythromycin and clindamycin were lower (x2 =13.106 and 11.743,both P ＜ 0.01).Among 159 MRSA strains,12 (7.5％) were of SCCmec type Ⅱ,113 (71.1％) were of SCCmec type Ⅲ,26 (16.4％) were of SCCmec type Ⅳ,and 8 were of undifferentiated type.The predominant SCCmec types were type Ⅳ for CA-MRSA (26/28,92.9％) and type Ⅲ for HA-MRSA (113/131,86.3％),respectively.Six PFGE patters were found in 49 HA-MRSA isolates from ICU,and the predominant patters were A1 (24,49.0％),A2 (9,18.4％) and B (9,18.4％).Cluster analysis of antibiogram showed that three groups of HA-MRSA were of high correlations,and they were of PFGE patter A1,A2 and B,respectively.Conclusions HA-MRSA is the predominant MRSA in Jingzhou area,and it is different from CA-MRSA in the age of patients,ward distribution,type of infection and antibiotic resistance.Most HA-MRSA strains are of type SCCmec Ⅲ,and may cause epidemic outbreak in ICU.

Objective To establish loop-mediated isothermal amplification(LAMP)method for detecting Streptococcus pneumoniae in clinical samples.Methods Four Streptococcus pneumoniae-specific LAMP primers were designed according to the published sequence of strain R6(GenBank accession number AE008540).Genomie DNA in positive blood cultures was extracted by nanidine hydrochloride and benzene-methanol.Then lytA was amplified by LAMP at 63℃ for 45 minutes.We observed the turbidity in thereaction tube.For further confirmation.The amplified products were also detected using electrophoresis in 2% agarose gels,followed by ethidium bromide staining.Resuits 16 strains of Streptococcus pneumoniae were detected in 196 positive blood cuhure bottles by LAMP.Compared with traditional method.It8 sensitivity and specificity were both 100%and the detection could be finished in an hour.The assay had a minimum detection limit of 102 CFU/m1.Conclusions This IJAMP-based assay is simple,rapid.Sensitive and specific.It can be used to detect trept OCOCCUS pneumoniae in clinical samples.

Objective In order to choose a suitable method in detecting MRS,the detection rate,sensitivity and specificity of Vitek-32 auto microbacteria indentity system,oxacillin agar dilution test,cefoxitin disk diffusion test and PCR for mecA gene are evaluated.Methods MRSA and MRCNS are detected by the four methods metioned above in 175 staphylococci,then comparing the postive detection rate by Chi-square test and calculating sensitivity and specificity of the other three methods based on PCR for meeA gene as a gold standard.Results The detection rate of four methods have no difference in detecting MRSA,but the detection rate of Vitek-32 auto microbacteria indentity system and oxacillin agar dilution test is better than that of PCR for mecA gene in detecting MRCNS.Sensitivity and specificity of Vitek-32 auto microbacteria indentity system,oxacillin agar dilution test and cefoxitin disk diffusion test in detecting MRSA are 100%、96.3%、96.3% and 88.2%、100%、100% respectively,the sensitivity of detecting MRCNS are all 100%,the specificity of detecting MRCNS are 50%、46.1% and 65.4% respectively.Conclusions Both mecA gene and the determination for MIC of Oxacillin should be considered in final decision for MRS,Furthermore.the MRS mediated by mecA gene and the MRS mediated by non-mecA gene should be treated differentially.

OBJECTIVE To establish a method of polymerase chain reaction(PCR) for detecting staphylococci in positive blood culture bottles.METHODS Genomic DNA in 493 positive blood culture bottles was extracted by guanidine hydrochloride and benzenemethanol,then genes 16S rRNA,ssa and mecA were amplified by PCR to identify staphylococci.Finally,the results of PCR were compared with that of traditional method.RESULTS To compare with traditional method,as the golden standard the sensitivity and specificity of PCR method were 98.6% and 100.0%,respectively,the detection could be finished in four hours.Method of PCR was better than traditional method in detecting meticillin-resistant staphylococci.CONCLUSIONS The PCR-based assay is simple,rapid,sensitive and specific,it can be used to detect staphylococci in positive blood culture bottles.

OBJECTIVE To investigate the prevalence of erythromycin resistance genes ermB and mefA and the relationship of drug resistance and genes in Streptococcus pneumoniae.METHODS Forty three strains of S. pneumoniae were collected from respiratory system infected children from Dec 2004 to Oct 2005 at Yuying Pediatric Hospital of Wenzhou Medical College.Erythromycin sensitivity test was done by using MIC method.The erythromycin resistance genes ermB and mefA were detected by PCR.RESULTS In all forty three strains,forty were erythromycin resistant(93%),three were erythromycin sensitive.The total detection rate of erythromycin resistance genes ermB and mefA was 76.7% and 23.3%,respectively.There were neither gene ermB nor gene mefA in 3 erythromycin-sensitive S.pneumoniae.In 40 strains the detection rate of gene ermB was 82.5% and that of gene mefA was 25%.The erythromycin resistance gene ermB or mefA were detected in 35 of the 43 strains.The total detection rate of erythromycin resistance gene was 81.4%.In the 35 erythromycin resistance strains there were 25 strains in which gene ermB existed lonely and 2 strains in which gene mefA existed lonely.There were both genes ermB and gene mefA in 8 of the 35 erythromycin resistance strains.CONCLUSIONS The erythromycin resistance of S.pneumoniae can be caused mainly by gene expression of ermB or mefA,but the gene mefA seems to be less important than gene ermB.Obviously the erythromycin isn′t useful in treating S.pneumoniae infection.

Objective To investigate mutation and deletion of genes mecR1 and mecI in clinical methicillin-resistant staphylococci isolates and study the mutation and deletion have effect on gene mecA expression and drug resistance phenotype.Metheods PCR was used to detecte gene mecA and the regulatory genes mecR1 and mecI in staphylococci which were separated from clinical specimen in 2006,then the sequence of gene mecI was determined and compared with the sequence obtains from pre-MRSA strain N315(GI:BA000018).Results Gene mecA was detected in 60 strains of Staphylococcus aureus,58 strains of Staphylococcus epidermidis and 37 strains of Staphylococcus heamolyticus,but gene mecA in 6 strains of Staphylococcus epidermidis and 4 strains of Staphylococcus heamolyticus were only amplified by primer mecA2-F/R and not by primer mecA1-F/R.The percentage of gene mecR1 exist in Staphylococcus aureus was higher than Staphylococcus epidermidis and Staphylococcus heamolyticus,but the percentage of gene mecR1 exist in Staphylococcus epidermidis was not higher than Staphylococcus heamolyticus.The mutation and deletion of gene mecI were often seen,the wild type mecI was only detected in 14 strains,the point mutation of nucleotice 202 was detected in 36 strains.Conclusions Gene mecA expression in Staphylococcus aureus could be chiefly induced by mecR1,but which in coagulase-ngeative staphylococci could be other factors.The mutation and deletion of mecI were universal phenomenon in clinical strains,there could be a mechanism for overcoming the repressing of resistance caused by mecI in staphylococci.

OBJECTIVE To establish a method of loop-mediated isothermal amplification(LAMP) for detecting Staphylococcus aureus in positive blood culture bottles.METHODS Genomic DNA in 293 positive blood culture bottles was extracted by guanidine hydrochloride and benzenemethanol,then genes ssa and mecA were amplified by LAMP to identify S.aureus.Finally,the results of LAMP were compared with the results of traditional method.RESULTS Twenty-two strains of S.aureus were detected in 293 positive blood culture bottles by LAMP method.Compared with traditional method,the sensitivity and specificity of LAMP method were both 100%,respectively,the detection could be finished in an hour.CONCLUSIONS The LAMP-based assay is simple,rapid,sensitive and specific which can be used to detect S.aureus in positive blood culture bottles rapidly.

Objective To evaluate the prognostic value of China Classification System and TNM staging in patients with liver cancer undergoing resection. Methods From Jan 1986 to Dec 2000, 246 patients underwent resection of liver cancer. At least three years of follow-up was made in these 246 cases. Results The 1, 3, 5, 7, and 10-year disease-free survival rates were 55%, 30%, 25%, 20% and 18%, respectively. The 1,3,5,7,and 10-year disease-free survival rates predicted by China Classification System and TNM staging were statistically different and positively correlated with each other. Differences of survival rate between stageⅠa、Ⅰb、Ⅱa 、Ⅱb and Ⅲ by China Classification System were all statistically significant. Conclusions Based both on tumor extension and liver function, China Classification System was more accurate in than TNM stage predicting the prognosis of liver cancer patients undergoing resection.

Objective To investigate the clinicopathologic features and treatment of thyroid microcarcinoma (TMC). Methods From January 1997 to December 2006,311 patients who underwent surgery and defined as TMC(tumor size≤1 cm)were enrolled. Results TMC was identified incidentally by frozen pathologic examination on thyroidectomy specimens in tentative benign goiters in 181 patients; another 130 patients with clinically detectable primary tumors or suspected nodal metastases were grouped to as clinically overt TMC. The clinically overt TMC had a higher incidence of bilateral multifocal tumors (18.5%vs.9.4%,P=0.03),and cervical lymph node metastases(27.7%vs.10.5%,P=0.000)than that in clinically occult TMC group. Conclusion TMC may vary considerably in clinical and biologic behaviors between these two subtypes: clinically overt and occult. Lobectomy for single lesion, total or near total thyroidectomy for multifocal with central compartment nodal dissection should be performed, lateral nodal dissection was not carried out unless US or physical examination detected nodal metastases. Lobetomy, subtotal or more limited thyroidectomy for occult TMC, diagnosed incidentally following thyroid surgery for initially tentative benign thyroid disease, could all be treatment of choice depending on the preference of surgeons.