Objective To investigate the distribution of virulence-related genes in multidrug resistant Escherichia coli.Methods Seven virulence genes papA,cnf1,cnf2,cfaB,ipaB,hofQ and ompT were detected by PCR in 20 strains of multidrug resistant Escherichia coli clinically isolated,and the positive genes were further searched in 31 strains of Escherichia coli in BioCyc database whose genomies had been fully sequenced.Results Virulence genes hofQ and ompT were detected in 20 strains of Escherichia coli with a positive rate of 95.0％ (19/20) and 55.0％ ( 11/20),respectively.Among 31 strains of Escherichia coli in BioCyc,21 (67.7％) were positive for hofQ gene and 15 (48.4％) were positive for ompT gene.Conclusion hofQ and ompT genes are prevalent in multidrug resistant Escherichia coli.

Objective To investigate the expression of T cell immunoglobulin-and mucin-domaincontaining molecule-3 (Tim-3) in peripheral CD8 +T cells and its significance in patients with chronic hepatitis B (CHB).Methods Fifty-eight CHB patients and 16 healthy controls were enrolled.Tim-3 expression in CDs + T cells was detected by flow cytometry,and quantities of IFNγ-producing HBV-specific cytotoxic T lymphocytes (CTLs) in HLA-A2 positive subjects were detected by enzyme-linked immunosorbent spot (ELISPOT) test before and after the blockade of Tim-3/Tim-3L pathway.Paired t test was performed to compare the quantities of CTLs before and after the blockade,and nonparametric Spearman correlation analysis was performed to explore the correlation in quantitive data.Results Tim-3 expression in CHB patients was (14.2 ± 8.98 )％,which was higher than that of healthy controls (4.80 ± 2.92)％,and the difference was of statistical significance (x2 =92.48,P ＜ 0.05 ) Tim-3 expressions in 16 severe CHB patients and 42 mild CHB patients were ( 19.54 ± 10.95) ％ and (9.58 ± 7.30) ％,respectively,and the difference was statistically significant (x2 =77.24,P ＜ 0.05 ). Before the blockade of Tim-3/Tim-3L pathway,IFNγ-producing HBV-specific CTLs were 7.27 ± 3.14,and it increased to 19.62 ± 4.97 after the blockade ( t =2.95,P ＜ 0.05 ).Conclusion The upregulation of Tim-3 on peripheral CD8 + T cells may inhibit HBV-specific CTLs,and the blockade of Tim-3 pathway can enhance the proliferation of IFNγ-producing HBV-specific CTLs,thus can enhance antiviral effect.

Objective To determine the population structure of Streptococcus pncumoniae isolates collected from infants with eye infections,including drug resistance,resistance genes, serotypes and molecular types.Methods The susceptibility of 39 isolates to 10 antibacterial agents was tested by K-B disk diffusion and Etest.Latex agglutination test was performed to determine the serotype of the strains,and PCR was carried out to detect macrolides resistance genes mefE and ermB.Molecular types of the 20 strains were determined by multilocus sequence typing (MLST).Results A total of 39 Streptococcus pneumoniae isolates were obtained,in which 30 (76.9％) were resistant to 3 or more antibacterial agents,and no vancomycin,penicillin or cefotaxime resistant strain was found.ermB gene was found in 33 strains and mefE gene was found in 4 strains.Twelve serotypes were found,and the most frequent serotypes were 19 (8/39) and 14 (4/39). Seventeen strains (43.6％) were covered in PCV7 vaccine. The international clone Taiwan19F-14 and Spain23F-1 were found by MLST. Conclusions Streptococcus pneumoniae isolates from infants with eye infections include international resistance clones.The distribution of serotype and molecular type are dispersed, and the clones are sporadic. The isolates are highly resistant to commonly used antibacterial agents.

Objective To survey the incidence of hepatocellular carcinoma (HCC) in patients with HBV-related cirrhosis receiving nucleos(t)ide analogues treatment and to assess its risk factors.Methods A total of 141 patients with HBV-related liver cirrhosis receiving nucleos(t) ide therapy from April 2008 to June 2011 were enrolled.The clinical data including virological and biochemical tests were retrospectively analyzed.Univariate and multivariate Cox proportional hazards regression model was used to identify the risk factors of HCC occurrence.Results Patients were followed up for 6.4 to 87.6 months with a median followup time of 32.5 months.During the follow-up period,15 out of 141 patients developed HCC with an average annual incidence rate of 3.8％.HCC incidence was higher in HBeAg positive cirrhosis and in those with family history of liver cancer ( RR =4.524 and 3.858,P ＜ 0.05 ).Conclusions Patients with HBV-related cirrhosis have a high incidence rate of HCC even they recieve nucleos (t) ide analogues treatment.HBeAg positive cirrhosis and family history of liver cancer are independent risk factors for HCC.

Objective To investigate the infection and risk factors of multidrug-resistant Pseudomonas aeruginosa in diabetic foot.Methods Totally 85 strains of Pseudomonas aeruginosa were isolated from 428 samples of diabetic foot ulcers in Tianjin Metabolic Diseases Hospital from Jan 2008 to Dec 2010.Drug sensitivity tests were performed and multivariate logistic regression analysis was used to examine the risk factors of multidrug-resistant Pseudomonas aeruginosa infection. Results In 85 strains of Pseudomonas aeruginosa, 28 (32.94％ ) were multidrug resistant. Multidrug-resistant Pseudomonas aeruginosa showed high resistance to 3rd generation cephalosporins,quinolones and aminoglycosides with resistant rates of 42.86％-67.86％,32.14％ -57.57％,and 42.86％-67.86％,respectively. Logistic analysis indicated that previous antibiotic treatments, hypertension and anemia were associated with multidrug-resistant Pseudomonas aeruginosa infection ( OR =5.758,0.257 and 0.270,P =0.006,0.014 and 0.013 ). Conclusion Multidrug-resistant Pseudomonas aeruginosas isolated from diabetic foot are highly resistant to several commonly used antibiotics,and previous antibiotics use,hypertension and anemia are risk factors for multidrug-resistant Pseudomonas aeruginosa infections.

Objective To investigate the correlation between acquired drug resistance-related genes and mobile genetic elements from pandrug-resistant Acinetobacter baumannii. Methods Fifty-three horizontal transfer drug resistance-related genes ( β-lactamases,aminoglycoside and quinolones resistance related) and 12 mobile genetic elements (including zygosity plasmid,transposon,insertion sequence and integron) were detected by polymerase chain reaction (PCR) in 20 clinical isolates of pandrug-resistant Acinetobacter baumannii.Index cluster analysis was performed to explore the correlation.Results In 20 strains of pandrug-resistant Acinetobacter baumannii,there were 3 types of β-lactamases related genes (TEM-1,ADC-30,OXA-23 ),4 types of aminoglycoside modifying enzyme genes [ aac (3)-Ⅰ,aac(6′)-Ⅰ b,ant( 3″)-Ⅰ and aph( 3′)-Ⅰ ],and 5 kinds of mobile genetic elements ( int Ⅰ 1,tnpU,tnp513,IS26 and ISAba1 ). Index cluster analysis showed high correlations between resistance genes [TEM-1,ADC-30,aac( 6′)-Ⅰ b,ant( 3″)-Ⅰ,abeB,qacE Δ1] and mobile genetic elements ( int Ⅰ 1,tnpU,tnp513,IS26,ISAba1 ).Conclusion Clinical isolated pandrug-resistant Acinetobacter baumannii carries several acquired drug resistance-related genes and mobile genetic elements,and there may be a close association between them.

Objective To evaluate the killing efficacy of chlorine-releasing agents (CRAs) with different concentrations on multidrug-resistant Acinetobacter baumannii.Methods Totally 30 clinical strains of multidrug-resistant Acinetobacter baumannii were collected from November 2008 to December 2009.Killing efficacy of CRAs on these strains was evaluated by quantitative suspension test.The one-way analysis of variance was performed.Results The log values of killing to 30 strains of multidrug-resistant Acinetobacter baumannii were all ≥5.00,when the bacteria were exposed to available chlorine concentrations 400 mg/L of CRAs for 10 min,600 mg/L for 10 min,800 mg/L for 3 min and 1000 mg/L for 1 min,respectively.And effective rates were all 100％. Furthermore,there were significant differences among different available chlorine concentrations exposed for the same time ( except 10 min) ( F =72.72,64.79 and 32.33,P =0.00),and among different exposure time in the same available chlorine concentration ( except 1000 mg/L) ( F =110.42,20.41 and 3.20,P=0.00,0.00 and 0.03).Conclusion Satisfactory killing efficacy of CRAs to multidrug-resistant Acinetobacter baumannii can be achieved with available chlorine concentration 500 mg/L to 1000 mg/L and exposure time 10 min to 30 min.

Objective To establish a PCR method for rapid identification and sequence typing of VG Ⅱ allele of the intergenic spacer region (IGS) of Cryptococcus gattii.Methods Since IGS1 was of high sequence variation,multiple alignments were conducted by ClustalX 2 in IGS1 of Cryptococcus gattii and Cryptococcus neoformans,and then primer sets specific to genotype VG Ⅱ was designed for PCR analysis.The specificity of the primer pair was detected by amplification of the other genotypes in Cryptococcus gattii,Cryptococcus neoformans,and other pathogenic yeasts.The amplified fragments from VG Ⅱ genotype were sequenced and subtyped.Results Using the PCR analysis developed in this study,all VG Ⅱ genotype strains tested were amplified,whereas no amplification was obtained from other genotypes or yeast species involved herein.Three polymorphic nucleotide sites at 72,79 and 104 bp in the fragment amplified could be used to distinguish sub-genotypes within VG Ⅱ genotype.Conclusions The PCR analysis developed in this study can be used for rapid identification of genotype VG Ⅱ of Cryptococcus gattii.The sequence typing based on the amplified fragment from IGS1 may be performed for screening the highly virulent sub-genotype VGⅡ a.

Objective To investigate the distribution and drug resistance of major pathogens for urinary tract infections in the First Affiliated Hospital of Nanjing Medical University.Methods Strains from midstream urine culture of patients with urinary tract infections were collected during January 1 and December 31,2011.All strains were identified by API system,and disk diffusion method was used for drug sensitivity test.Results Totally 1129 strains were isolated,in which 667 (59.1％ ) were Gram-negative strains,266 (23.5％) were Gram-positive strains,and 196 (17.4) were Candida.Among Gram-negative strains,Escherichia coli,Klebsiella pneumoniae and Proteus mirabilis were highly sensitive to carbapenem antibiotics; while Acinetobacter baumannii and Pseudomonas aeruginosa were highly resistant to most antibiotics including cephalosporins and penicillinase inhibitor,and the resistance rates were over 50％.Among Gram-positive strains,the major strains Enterococcus avium and Enterococcusfaecalis were completely sensitive to vancomycin and teicoplanin,and highly sensitive to linezolid (resistance rate below 10％ ).Candida albicans and Candida glabrata were highly resistant to voriconazole and fluconazole (with the resistance rates of 47.2％ - 60.0％ ), but were completely sensitive to amphotericin and nystatin.Conclusion Gram-negative strains account for most urinary tract infections in the First Affiliated Hospital of Nanjing Medical University with high drug resistance rates.

Objective To investigate the distribution and antibiotic susceptibilities of quantitatively cultivated bacteria from bronchoalveolar lavage fluid (BALF) samples. Methods Totally 312 BALF samples were streak inoculated to chocolate,blood and MAC plates with 10 μL annulus,and the bacterial colony ＞ 104 CFU/mL was considered pathogenic bacteria. The identification of pathogenic bacteria was carried out with Vitek 2-Compact,and Kirby-Bauer disc agar diffusion method,Etest and dilution method were used for antibiotics sensitivity test.Results Totally 216 (69.2％) strains of pathogenic bacteria were isolated.The major gram-negative strains were Pseudomonas aeruginosa,Acinetobacter baumannii,Klebsiella pneumoniae and Escherichia coli, and the major gram-positive strains were Staphylococcus aureus,Streptococcus pneumoniae and Staphylococcus epidermidis.The resistance rate of Pseudomonas aeruginosa to aztreonam was high,but lower than 30％ to piperacillin/tazobactam,imipenem,cefepime,ofloxacin,ceftazidime and amikacin.Staphylococcus aureus was highly resistant to erythromycin,benzylpenicillin and clindamycin,but it was sensitive to furadantin,vancomycin,quinupristin/dalfoprisdn,tigecycline and linezolid.Conclusion The positive rate of BALF cultivation is high,and the main pathogenic bacteria Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella pneumoniae, Staphylococcus aureus and Escherichia coli are resistant to several commonly used antibiotics.