OBJECTIVE:To construct the homologous recombination plasmid of sigB gene from staphylococcus aureus so as to provide the basis for further study of the formation mechanism of its biofilm.METHODS:The target fragment“upstream sequence of sigB,kana-resistance gene,and downstream sequence of sigB”were amplified by PCR from staphylococcus aureus DNA and pEGFP-N2plasmid,respectively,in which the terminals of3interlinked fragments were complementary,and re-stricted enzyme EcoRⅠand XhoⅠsite existed in the sequence terminal,respectively.The fragments were ligated with fusion PCR in turn.Then target fragment was recombinated into plasmid pBluescriptⅡSK(+)according to the instruction manual of commercial kits.After both recombination of pBluescriptⅡSK(+)and vector plasmid pGEM-7zf(+),restricted enzyme SacⅠand XhoⅠ,the homologous recombination plasmid pGEM-7zf(+)-sigB was finally made via T 4 ligation enzyme.RE-SULTS:The target fragments were amplified by PCR and were confirmed with the agarose gel electrophoresis and sequencing.CONCLUSION:The homologous recombination plasmid of sigB gene from staphylococcus aureus is constructed successfully.

Objective To understand the present situation of therapeutic drug monitoring(TDM)on cyclosporine A(CsA) application in renal transplant recipients.Methods The clinical data of the first postoperative month of 50 renal transplant cases admitted to our hospital during May 2000 to March 2005 were collected and retrospectively analyzed.Results Total 50 cases(male∶female= 3∶1) had accepted the trigeminy of CsA,prednisone and azamun.The average initial dose and average maintenance dose for the male were higher by 59.94 mg/d and 52.00 mg/d than that for the female(P

OBJECTIVE:To provide a basis for management of use of cephalosporins in hospital METHODS:The cephalosporins used in 18 hospitals of Chongqing in 2001～2003 were statistically analysed in respect to the kind,consumption sum and DDDs etc RESULTS:During the 3-year period,the sum of cephalosporins consumption increased with an annual rate of 23 34%,of which the third generation cephalosporins were the maximum consumption in kind accounting for 50% of total cephalosporins used Compared to those in 2001,DDDs of the third generation cephalosporins increased 540 49%,but DDDc decreased 75 22% As for fourth generation cephalosporins,the annual sum and consumption were under 2% and 0 3% respectively CONCLUSION:The consumption of the third generation cephalosporins has increased sharply In order to reduce and delay the bacteria resistance,it is very important to make measures as soon as possible to control the cephalosporins abuse strictly

OBJECTIVE:To probe into the tendency and influencing factors of the drug expenses of the inpatients in our hospital.METHODS:The total drug expense,the antibiotic expense and its proportion in the total of the inpatients in our hospital were analyzed statistically in this retrospective study.RESULTS:During 2003～2007,the average daily total drug expense per inpatient was 371.77 yuan with average annual growth rate of 11.94%;the average daily antibiotics cost per inpatient was 110.35 yuan with average annual growth rate of 11.92%.Both the drug expense and antibiotic expense were correlated positively with the consumer price index (CPI).And the proportion of antibiotic expense in the total drug cost remained stable at about 29.68%.CONCLUSION:The average daily drug cost and antibiotic expenses per inpatient between 2003 and 2007 fluctuated yet with upward trend on the whole,and the proportion of antibiotic expense in the total drug cost remained stable.

OBJECTIVE:To survey and evaluate the current situation and developing trend of antibiotic/beta-lactamase inhibitor(BLI)preparations consumed.METHODS:The antibiotic/BLI preparations used in18hospitals of Chongqing in2001～2003were collected in respect to the compositions,dosage forms,specifications,drug name,manufacturers and sum of money for consumption,and the clinical application information was analysed.RESULTS:The sum of money for these antibi?otics/BLI consumed in the years2001,2002and2003were2229,2937and4119ten-thousand yuan(RMB),respectively,with an average annual increase rate of26.41%.Among them the consumption costs of antibiotics plus sulbactam,clavulanate and tazobactam were6280,2276and729ten-thousand yuan(RMB),accounting for67.64%,24.51%and7.85%of the total consumption sum,with an annual increase rates of32.20%,35.93%and94.50%,respectively.CONCLUSION:The consump?tion of the antibiotic/BLI preparations was increasing fast in these years and the more requirement for the antibiotic/BLI preparations by market would be expected in the future.

Objective To investigate the correlation of biofilm forming ability with biofilm associated gene of the clinically isolated Staphylococcus aureus for providing basis for the further studies of the mechanisms of biofilm formation.Methods Safranine staining was conducted for the detection of the forming ability of biofilm in 96-well plates.icaAD,icaBC,sar,agr and sigB was amplified by PCR.Results Formation of biofilm could be found macroscopically in 17 out of 27 strains,especially to X387 and X409.icaAD and icaBC were amplified in 22 isolates and sar,agr and sigB in all 27 S.aureus strains.Conclusion ica operon is the key gene for biofilm formation but cooperative action of other genes is needed during the process of biofilm formation.

Objective To study the macrolide-resistance,ability of biofilm formation and icaA-genetypes of 68 stains of clinically isolated Staphylococcus epidermidis(S.epidermidis) so as to explore the efficacy of macrolide to prevent biofilm-associated infections caused by S.epidermidis.Methods Minimum inhibitory concentration(MIC) were detected by agar-plate dilution method,and microtiter-plate assay was used to investigate the ability of biofilm formation,and icaA genetype was identified by PCR.Results High macrolide-resistant rate(88.2%) was showed for clinical isolated S.epidermidis,and biofilms for macrolide-resistant strains were significantly stronger than that of macrolide-sensitive strains,but there was no dependability found between these two for icaA positive rate.Conclusion Frequently-used macrolide such as erythromycin,azithromycin,and clarithromycin may incompetence to prevent biofilm-associated infections caused by S.epidermidis.

Objective To study the current status of antibiotic usage in Southwest hospital, so as to improve the rationality in applying antibiotic. Methods A total of 1 668 patients in Southwest hospital during Jan 1 to Dec 30, 2003 were investigated in antibiotic usage and the data were sorted and analyzed. Results Antibiotics were administered in 1 247 cases (74.76%), including only 76 cases (6.09%) based on susceptibility test, 39 cases (3.12%) occurred hospital infection. 297 cases (23.81%) abused antibiotics, among which 131 cases (38.04%) were without definite symptoms or with weak signs, 93 cases (31.31%) used more than two kinds of antibiotics, 24 cases (8.08%) frequently changed antibiotic in the treatment, 27 cases (9.09%) inappropriately prolonged administration and 22 cases (7.40%) improperly combined the same antibiotics. Conclusion Irrational utilization of antibiotics exits in clinical work. Therefore, the rational use of antibiotic based on susceptibility tests should be emphasized.

Objective To study the effect of Bifidobacterium spent culture supernatant (SCS) on adhesion of Pseudomonas aeruginosa to intestinal epithelial cells. Methods The intestinal epithelial cells were cultured,then divided into three groups: control,Pseudomonas aeruginosa adhesion group,SCS and Pseudomonas aeruginosa adhesion group. The effects of SCS on cell viability,the number of adhering and invasive bacteria were evaluated with MTT assay and lysis-counting assay in 1,3,6 h. Results At 3 h after SCS treatment,the amount of Pseudomonas aeruginosa decreased significantly,and the number of adhering and invasive bacteria decreased by 71% and 87% respectively. Conclusion SCS could protect the intestinal epithelial cells by inhibiting the adhesion and invasion of Pseudomonas aeruginosa.

Objective To evaluate the endotoxin-neutralizing activity of modeling peptides from the limulus antilipopolysaccharide factor (MPLALFs, Ms) in vitro. Methods The endotoxin-binding activity of Ms was examined by biosensor technique and shown in values of Kon and Kd. The endotoxin-neutralizing effect was analyzed by limulus amebocyte lysate test. Results The biosensor technique results showed that the Kon values of M_0, M_1, M_2, M_3, M_4, M_6, M_8 and M_ 10 binding to LPS 055∶B5 were (840?5.716), (549?6.532), (842?6.530), (627?2.450), (996?5.716), (814?8.982), (556?1.633) and (635?2.449) arc second, of which M_4 and M_1 had the highest and lowest endotoxin-binding activity, respectively. The M_4 reacted to LPS with a Kd of 72.377 ?mol/L. The results obtained by the limulus amebocyte lysate test were the same with those from the biosensor technique. Conclusion M_4 has a potential good endotoxin-neutralizing effect in vitro.