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Objective:To investigate the role of type Ⅵ secretion system (T6SS) in the pathogenicity and antibiotic resistance of Acinetobacter baumanii. Methods:From January 1 to December 31, 2016, a total of 45 Acinetobacter baumanii isolates were collected from patients with bloodstream infection in the First Affiliated Hospital of Wenzhou Medical University. The susceptibilities to commonly used antimicrobial agents were determined by VITEK 2 Compact automatic microbiology analyzer. Detection of T6SS characteristic gene hemolysin coregulated protein ( hcp) was achieved by polymerase chain reaction. Biofilm formations, serum resistances and competition tests of T6SS-positive/negative Acinetobacter baumanii were performed in vitro. The clinical data of patients with bloodstream infection were collected and analyzed. Chi-square test, t test and Kruskal-Wallis test were conducted for statistical analysis. Results:The positive rate of T6SS in 45 Acinetobacter baumanii isolates was 53.3% (24/45). The resistance rates of T6SS-positive Acinetobacter baumanii to ceftazidime, ciprofloxdcin, gentamicin, imipenem, levofloxacin, piperacillin/tazobactam, tobramycin and cefepime (95.8%, 95.8%, 66.7%, 95.8%, 79.2%, 95.8%, 79.2%, 91.7%)were all higher than that of T6SS-negative Acinetobacter baumanii (28.6%, 28.6%, 28.6%, 28.6%, 9.5%, 23.8%, 23.8%, 28.6%), and the differences were all statistically significant ( χ2=22.12, 22.12, 6.51, 22.12, 21.83, 24.72, 13.79, 18.97, respectively, all P<0.05). The biofilm formation ability, serum resistance and competitive ability of T6SS-positive Acinetobacter baumanii were stronger than those of T6SS-negative Acinetobacter baumanii, and the differences were all statistically significant ( t=4.99, Z=-2.61 and -2.27, respectively, all P<0.05). The positive rate of T6SS isolated from intensive care unit (ICU) ward (80.0%, 16/20) was significantly higher than that from non-ICU ward (32.0%, 8/25; χ2=10.29, P<0.05). But T6SS had no effect on the prognosis of patients ( χ2=1.74, P=0.188). Conclusions:T6SS of Acinetobacter baumanii is associated with high pathogenicity, and the high drug resistance rate makes treatment extremely difficult. Physicians need to pay much attention, especially to the patients from ICU wards.
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Objective@#To investigate the distribution and expression of T3SS virulence genes in clinically isolated Pseudomonas aeruginosa (P. aeruginosa) strains and its correlation with drug resistance. @*Methods@#A total of 68 P. aeruginosa isolates were collected from the First Affiliated Hospital of Wenzhou Medical University in 2015. The antimicrobial susceptibility was detected by the agar dilution method. The distribution of virulence genes such as exoU, exoS, exoT and exoY from different isolates was detected by PCR. The expression levels of transcriptional regulator genes (ptrA and exsA) and effector-related genes (exoT and exoS) in some isolates were determined by real-time fluorescence quantitative PCR, and the Pearson correlation analysis was used to analyze the results. @*Results@#The detection rates of exoT and exoY in 68 P. aeruginosa isolates were higher, accounting for 79.4% and 75.0%, respectively. The detection rate of exoT in wound-sourced isolates was significantly higher than that in sputum (97.0% vs 61.8%, P＜0.01). In addition, the genotype of exoU - /exoS + was the most common, accounting for 51.5% (35/68). The resistance rates of sputum-sourced isolates to imipenem and meropenem were significantly higher than that of wound-sourced isolates (47.1% vs 8.8%, 47.1% vs 14.7%, P＜0.01). The resistance rates of isolates carrying exoU gene to carbapenems, aminoglycosides and fluoroquinolones were higher than those of isolates carrying exoS, exoT or exoY genes. Pearson correlation analysis showed that the expression level of ptrA gene was negatively correlated with those of exoT, exoS and exsA genes (P＜0.05). @*Conclusion@#The P. aeruginosa isolates from our hospital carrying T3SS virulence genes exoT and exoY are common, and the virulence genes are related to the drug resistance of P. aeruginosa. In addition, ptrA may be a potential negative regulatory gene for the expression of T3SS virulence genes.
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Objective@#To investigate the molecular mechanism of colistin resistance in Klebsiella pneumoniae (K.pneumoniae).@*Methods@#Three clinical isolates of colistin-resistant K. pneumoniae (FK1149, FK1920 and FK1934) and three colistin-resistant mutants (FK660R, FK713R and FK729R) were investigated. Resistance genes of pmrAB, phoPQ, mgrB, crrAB, mcr-1 and mcr-2 were detected by PCR and then analyzed by sequencing. PROVEAN platform was used to predict changes in the biological functions of proteins related to drug resistance. Expression of pmrH, pmrC, mgrB and phoP genes was measured using quantitative real-time PCR. LPS silver staining and conjugation assay were performed to analyze the three clinical colistin-resistant isolates.@*Results@#Amino acid substitutions in PmrA (G53V), PmrB (T157P, R256G), MgrB (F44C) and CrrB (E189K) were detected. ISkpn14 and IS5-like insertion sequences were detected in FK713R and FK729R, respectively. FK1149, FK1920 and FK1934 were negative for mcr genes. Compared with the wild-type strain, expression of pmrH and pmrC genes at the transcriptional level was increased in all investigated isolates. Changes in the expression of phoP and mgrB genes were also observed. A partial deletion of LPS was identified in FK1149.@*Conclusion@#LPS modification induced by inactivation of PmrAB or MgrB is the main molecular mechanism of colistin resistance in K. pneumoniae isolates in this study. Mutations in PmrA (G53V), MgrB (F44C) and CrrB (E189K) that might be related to colistin resistance are detected for the first time in clinical isolates of K. pneumoniae.
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@#Tumor occurrence is usually recognized as the interplay between genetic variations within the tumor and the environment. During a long time, great effort has been made in killing cancer cells. However, the role of tumor microenvironment has been largely ignored, which plays an important role in tumor generation, growth, invasion and metastasis. Meanwhile, tumor microenvironment not only facilitates the tumor infiltration, but also promotes the exchange of enzymes and cytokines to aid tumor proliferation, differentiation and self-renewal. Thus, better understanding of tumor microenvironment shows great importance. Recent developments in nanotechnology have brought new approaches to cancer diagnosis and therapy. Nanoparticles were suggested to show enhanced efficacy, while simultaneously reducing side effects and promoting bioavailability, owing to properties such as tumor localization and active cellular uptake. Additionally, nanoparticle surface chemistry has evolved from conventional synthetic polymers to more biologically inspired strategies, including cell membrane and self-recognition peptides, to minimize nonspecific uptake of nanoparticles. In the current review, we highlight the targets in tumor microenvironment and the strategies of nano drug delivery system to target tumor microenvironment for the treatment of cancer. We also highlight design considerations to improve nano drug delivery.
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@#To evaluate pharmacokinetic and metabolic characteristics of clevidipine butyrate lipid microspheres(CDB-LM)injection in mice, a novel HPLC-FLD method was developed for simultaneous measurement of clevidipine butyrate(CDB)and its metabolites clevidipine acid(MI)in whole blood samples. The chromatographic column was Waters C18(4. 6 mm×150 mm, 5 μm)and the mobile phase is consisted of acetonitrile-methanol-phosphate(2 ∶1 ∶2). The detection wavelength of FLD included excitation wavelength at 358 nm and emission wavelength at 440 nm. The pharmacokinetic parameters of CDB and MI were calculated by using DAS 2. 0. Then obtained parameters were statisticaly analyzed using PASW Statistics 18. The results showed that the half-life of CDB and MI were about 4 min and 20 min, respectively. Pharmacokinetic parameters of the low- and high-dose groups were as follows: CL of CDB were 4. 21 and 2. 72 L ·min-1 ·kg-1; AUC0-t were 3. 86 and 6. 43 mg/L ·min; MRT0-t were 7. 09 and 6. 17 min. CL of MI were 0. 34 and 0. 22 L ·min-1 ·kg-1; AUC0-t were 52. 23 and 74. 90 mg/L ·min; MRT0-t were 201. 24 and 217. 33 min. A method of protein precipitation was established, and acetonitrile was used to deal with whole blood samples. This method was simple, fast, with no interference with endogenous impurities. The results showed that the established HPLC-FLD method was simple and sensitive. It can be used to determine CDB and MI simultaneously. Comparing the low-dose group with the high-dose group, it was found that the plasma concentration-time curve of the two groups revealed the same tendency, which confirms that CDB has a short half-life and that it metabolizes to MI quickly.
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Total paeony glycoside (TPG) is extracted and purified from a traditional Chinese herbal medicine. It has many biological and pharmacological activities. However, there are few dosage forms of TPG in the market because of its low bioavailability. Self-microemulsifying drug delivery system (SMEDDS) is a vital tool in solving low bioavailability of poor absorption drugs. So the objective of this study is to develop a new TPG-SMEDDS for the oral delivery of poorly soluble TPG. Through the construction of pseudo-ternary phase diagrams, the optimum prescription was obtained, which consisted of 18.70% TPG, 16.27% ethyl oleate as oil, 43.34% Cremophor RH40 as surfactant and 21.73% Transcutol P as cosurfactant. The characterizations of TPG-SMEDDS including morphological characterization, droplet size, zeta-potential, emulsification time, and dissolution study of TPG-SMEDDS were evaluated. The results showed that TPG-SMEDDS is stable and its release rate is high in four different media (0.1 mol x L(-1) HCl, pH 6.8 PBS, pH 7.4 PBS, and water). The relative bioavailability of SMEDDS was dramatically enhanced in an average of 1.52-fold that of TPG-suspension. It is concluded that the bioavailability of TPG is enhanced greatly by SMEDDS.
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OBJECTIVE To investigate the production of metallo-beta-lactamase in clinical isolates of multi-resistant Pseudomonas aeruginosa and evaluate the validity of the detection methods.METHODS The multi-resistant strains were selected by K-B method according to the standard Aloush et al recommended.The metallo-beta-lactamase phenotypes were detected by multi-disk-multi-inhibitors synergy test(MDMIST),and the genotypes of IMP and VIM gene were analyzed by PCR amplification.RESULTS A total of 192 strains of multi-resistant P.aeruginosa were selected from 1081 clinical strains.The antimicrobial agents test in these multi-resistance strains demonstrated that ciprofloxacin and piperacillin had the highest resistant rate(92.5%),and the next were aztreonam and trimethoprim-sulfamethoxazole(91.5%),the polymyxin showed sensitive in all of these strains.Sixty-seven strains of metallo-beta-lactamase phenotypes were positive and the amplification PCRs showed that 65 strains were IMP or VIM in these 192 multi-resistant strains.CONCLUSIONS The resistance mechanisms in multi-resistant P.aerugionsa present multiple and changeable.The clinical laboratory should enhance the detection of metallo-beta-lactamase in these multi-resistant strains.
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OBJECTIVE To investigate the clinical distribution and antibiotic resistance of Enterococcus faecium in our hospital and provide the reference for the clinical treatment. METHODS The statistical method was used to analyze the status of drug resistance of 519 E. faecium strains which were isolated form our hospital from Jun 2006 to May 2008. RESULTS Among 519 isolated E. faecium strains, most common sources of specimens were urine (34.9%), sputum(26.8%) and feces (18.3%); E. faecium infection mainly distributed at ICU, respiratiory wards and EICU; E. faecium was resistant to multiple antibiotics. The drug-resistant rates to vancomycin and teicoplanin were 6.2% and 3.2%, respectively. CONCLUSIONS E. faecium could cause various infection, and has a high antibiotic resistance which is difficult to treat, we should be paid high clinical attention.
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OBJECTIVE To review and analyze the change in the MICs of vancomycin,teicoplanin and linezolid in meticillin-resistant Staphylococcus aureus(MRSA) strains isolated in our hospital from 2003 to 2007. METHODS The MICs of vancomycin,teicoplanin and linezolid were tested by Etest method on a sample of randomly selected MRSA strains. RESULTS The incidences of MRSA increased from 52.2% in 2003 to 74.5% in 2007.MIC of vancomycin increased from 1.85 ?g/ml in 2003 to 2.15 ?g/ml in 2007,and teicoplanin MIC geometric mean increased even more markedly from 1.28 ?g/ml in 2003 to 2.07 ?g/ml in 2007.The linezolid MIC remained almost unchanged. CONCLUSIONS The incidences of MRSA were increasing from 2003 to 2007.There is a upward trend in MIC of glycopeptide over the years,in which the increase for teicoplanin is higher than others two.