Prevalence and characteristics of clinical Aeromonas isolates carrying mcr-3 gene / 中华微生物学和免疫学杂志

Objective:To investigate the prevalence and characteristics of mcr genes in clinical isolates of Aeromonas spp. in our hospital, and provide reference for clinical analysis of the prevalence and expression of colistin resistance genes. Methods:Polymerase chain reaction (PCR) was used to detect mcr genes in 183 Aeromonas spp. strains. The minimum inhibitory concentrations (MICs) of colistin and polymyxin against mcr-positive Aeromonas spp. were detected by micro broth dilution method. Broth conjugation and filter mating conjugation were performed. Whole genome sequencing was used to analyze the genetic environment of mcr-3 gene in Aeromonas spp.. A recombinant Escherichia coli ( E. coli) DH5α-pGEM-T: : p mcr-3 strain was constructed to verify the expression of mcr-3 gene. Results:The positive rate of mcr-3 gene in 183 strains of Aeromonas spp. was 2.19% (4/183). No mcr-1 or mcr-2 gene was detected among these isolates. Antimicrobial susceptibility test showed that four mcr-3-carrying Aeromonas hydrophilia ( A. hydrophilia) strains were sensitive to colistin and polymyxin (MIC<2 μg/ml). Conjugation experiments indicated that mcr-3 gene could not be transferred between strains. Whole-genome sequencing analysis suggested that the mcr-3 genes carried by the A. hydrophilia isolates belonged to mcr-3.2 and mcr-3-like variants, and no adjacent transfer element was detected upstream and downstream. The recombinant E. coli DH5α-pGEM-T: : p mcr-3 strain was sensitive to colistin (MIC=2 μg/ml). Conclusions:The clinical isolates of A. hydrophilia in our hospital carried mcr-3 gene, but does not exhibit colistin resistance, and no evidence supported the transfer of mcr-3 gene for the time being.

Molecular epidemiology and virulence characteristics of polymyxin-resistant Klebsiella pneumonia / 中华传染病杂志

Objective:To analyze the molecular epidemiology and virulence characteristics of polymyxin-resistant Klebsiella pneumoniae ( K. pneumoniae). Methods:From 2011 to 2016, 1 376 strains of K. pneumoniae were isolated from various clinical specimens of hospitalized patients in First Affiliated Hospital of Wenzhou Medical University. Agar dilution method was used to screen out the polymyxin-resistant strains.Polymerase chain reaction (PCR) was used to detect the genes related to polymyxin resistance, and real-time fluorescence quantitative PCR was used to detect the relative mRNA expression level of drug resistant genes. Pulsed-field gel electrophoresis, multilocus sequence typing and Galleria mellonella larvae infection model were performed to analyze the molecular epidemiological and virulent characteristics. Results:A total of 14 strains (1.02%) of polymyxin-resistant K. pneumoniae were detected among 1 376 K. pneumoniae isolates. Subsequent sequencing identified mutations leading to amino-acid changes (K2E, F28C) in MgrB of 10 isolates and D150G in PhoQ of nine isolates, and genes such as mcr and crrB were not detected in all drug-resistant strains. Compared with standard strains, the relative expression levels of pmrH and pmrD mRNA of these drug resistant strains were increased. Analysis of the molecular epidemiology indicated that the 14 drug-resistant strains were divided into nine clones. Galleria mellonella larvae infection model revealed polymyxin-resistant K. pneumoniae isolates had higher virulence. Conclusions:Polymyxin-resistant K. pneumoniae has mutations in mgrB and phoQ genes, and mgrB mutation may play a key role in the change of virulence profiles. The homology among the polymyxin-resistant K. pneumoniae stains in this study is low.

Role of type Ⅵ secretion system in the pathogenicity and antibiotic resistance of Acinetobacter baumanii / 中华传染病杂志

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.

Molecular mechanism of colistin resistance in Klebsiella pneumoniae / 中华微生物学和免疫学杂志

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.

In vitro antibiotic effects of drug combinations against colistin-heteroresistant Acinetobacter baumannii / 中华微生物学和免疫学杂志

Objective@#To evaluate the in vitro antibiotic effects of colistin combined with meropenem, levofloxacin or fosfomycin against colistin-heteroresistant Acinetobacter baumannii (A.baumannii).@*Methods@#A total of 576 A. baumannii clinical isolates were collected from the First Affiliated Hospital of Wenzhou Medical University from 2014 to 2015. Minimal inhibitory concentrations (MICs) of colistin against A. baumannii were detected by broth dilution method. Colistin-heteroresistant A. baumannii isolates were screened using population analysis profiles (PAPs). MICs of colistin combined with meropenem, levofloxacin or fosfomycin, and the four drugs used alone against colistin-heteroresistant A. baumannii were detected by checkerboard method and broth dilution method. Fractional inhibitory concentration index (FICI) was calculated to evaluate antibiotic effects.@*Results@#None of the 576 A. baumannii isolates was resistant to colistin as indicated by the broth dilution method. Nine colistin-heteroresistant A. baumannii isolates were identified using PAPs. Compared with the MICs of colistin used alone, the MICs of colistin used in combination with meropenem, levofloxacin or fosfomycin against colistin-heteroresistant isolates were all decreased. Colistin-meropenem combination showed synergistic (55.6%), additive (33.3%) and indifferent effects (11.1%), but no antagonistic effect. Colistin-levofloxacin combination showed synergistic (55.6%), additive (22.2%) and indifferent effects (22.2%), but no antagonistic effect. Colistin-fosfomycin combination showed synergistic (77.8%) and additive (22.2%) effects, but no indifferent or antagonistic effect.@*Conclusion@#In vitro use of colistin in combination with meropenem, levofloxacin or fosfomycin has synergistic and additive antibacterial effects against colistin-heteroresistant A. baumannii. Combinations of colistin-meropenem and colistin-levofloxacin have fewer indifferent effects and no antagonistic effect.

Study of mitochondrial energy metabolism in D-galactose-induced cell ageing model / 中华老年医学杂志

Objective To investigate the mitochondrial energy metabolism in D-galactoseinduced cell ageing model.Methods MRC-5 cells were cultivated for 72 hours in a medium containing 55 mmol/L D-galactose.The analysis of cell proliferation capacity by CCK8 method,β-galactosidase staining and detection of p21 protein expression level were performed for identifying cell senescence.The cell oxidation-reduction state was evaluated by an analysis of cellular ROS levels,SOD activity,MDA content and oxidative damage level of mitochondrial DNA(mtDNA).For purpose of detecting mitochondrial function and its impairment,mitochondrial morphology was observed by electron microscope,mitochondrial quantity was analyzed by flow cytometry,mitochondrial membrane potential(△Ψm) was measured by JC-1 staining,and ATP content was analyzed by HPLC,and mitochondrial oxygen consumption rate was detected by Seahorse cell energy metabolism detection system.Results The decreased MRC-5 cell proliferation,up-expression of p21 protein,increased β-galactosidase activity were observed in D-Gal-treated cells,which indicated the cell premature senescence.When treated with D-Gal,the significantly increased ROS and MDA level,decreased SOD activity and increased oxidized mtDNA proved that the cells kept higher oxidative stress.D-Gal induced-mitochondrial impairment was evidenced by the dimming of mitochondrial cristae and double membrane structure,decrease of transmembrane potential and ATP synthesis,and decrease of its oxygen consumption rate(OCR).Conclusions The 55 mmol/L D-Gal causes an impairment of mitochondrial structure and a decrease of function of energy metabolism,which is associated with cellular senescence induced by D-Gal.

Prevalence and plasmid characterization of the qnrD determinant in Morganella morganii isolates / 中华微生物学和免疫学杂志

Objective To investigate the prevalence and plasmid size of qnrD determinant in Morganella morganii (M.morganii) isolates.Methods A total of 100 non-duplicated M.morganii clinical isolates were collected from inpatients.Standard ager dilution method was used to determine the minimum inhibitory concentrations (MICs) of fluoroquinolones against M.morganii isolates.PCR were performed to detect plasmid-mediated quinolone resistance determinants (PMQRs) in M.morganii isolates and the prevalence of extended-spectrum β-lactamase (ESBL) genes and AmpC β-lactamase genes in PMQRs-positive M.morganii strains.The homology analysis among qnrD-positive M.morganii strains were conducted by using pulsed-field gel electrophoresis (PFGE).The location of qnrD gene and the size of plasmid carrying it were determined by southern hybridization.The transferability of qnrD gene was determined by conjugation experiment.Results Thirty out of 100 M.morganii isolates (30％) were found carrying PMQRs including 17 qnrD-positive strains,14 aac (6')-Ib-cr-positive strains and 5 qepA-positive strains.PCR and sequencing confirmed that thirty PMQRs-positive isolates carried blaDHA-1.Among them,six isolates were positive for ESBLs genes (four for blaCTX-M-14,one for blaCTX-M-3 and one for blaCTX-M-24) and four isolates were positive for blaTEM-1.Almost all PMQRs-positive M.morganii isolates showed reduced susceptibility to fluoroquinolones.Moreover,seventeen qnrD-positive M.morganii isolates harbored blaDHA-1 including five (29.4％) harboring aac(6')-Ib-cr gene,four (23.5％) harboring blaCTX-M-14,two (11.8％) harboring blaTEM-1 and one harboring aac(6')-Ib-cr gene,blaCTX-M-14 and blaDHA-1.PFGE analysis showed that the 17 qnrD-positive M.morganii isolates were divergent from each other and not clone-related.Southern hybridization analysis showed that qnrD genes of all M.morganiiis isolates were mainly located in a 2.7 kb plasmid,but only a few of them were located in a size of 5.1 kb plasmid.M.morganiiis isolates failed to transfer qnrD gene to E.coli EC600 through conjugation.Conclusion PMQRs were widely distributed in M.morganiiis isolates.qnrD gene was the predominant determinants with a high prevalence rate of 17.0％,followed by aac(6')-Ib-cr gene.qnrD gene was located on a non-conjugative plasmid of approximately 2.7 kb or 5.1 kb.One qnrD-positive M.morganii isolate carrying aac(6')-Ib-cr gene,blaCTX-M-14 and blaDHA-1 was detected.

Study on the resistant genes and virulence gene of vancomycin-resistant Enterococci / 中华微生物学和免疫学杂志

Objective To study the vancomycin-resistant genes and the virulence factors genes in vancomycin-resistant Enterococci (VRE),and to analyze the drug-resistance character and epidemic characteristics of VRE strains and provide the basis for clincal selection of drugs and infection control.Methods VRE were screened by agar dilution sieving plate (ADSP) containing 6 μg/ml of vancomycin,drug resistance of VRE to other common antibiotics were detected by VITEK-60 automatic microbial analyzer.The gene types and virulence factor genes of VRE were determined by PCR.And the genetic relationships among VRE were determined by multilocus sequence typing.Results Seven vancomycin-resistant Enterococcus faecium strains were found in 360 enterococcus strains.All the VRE strains exhibited high-level vancomycin resistance ; some of them were medium or senstive to teicoplanin.They all carried vanA gene and esp gene and one of them carried 4 kinds of virulence factor genes.The ST type of the 7 VRE strains were diffused distribution.Conclusion We found vanB phenotype vanA genotype vancomycin-resistant Enterococcus faecium isolates in Wenzhou; these VRE strains were multidrug resistance and carried various virulence factor genes.Linezolid could be used as a recommend drug for treatment of VRE infection.The protection of antibiotics sensitivity should be strengthened.

Reflections on offering and readjusting specialties of medical sciences in the catalog for regular college program / 中华医学教育探索杂志

The catalogue for regular college programs is the guide to offering specialties and academic degrees in medical colleges and universities.It serves as the orienting framework under which to develop human resources at the institution of higher learning.Thus,it is an important task concerning the overall reform and development of tertiary education to revise the catalogue for regular college programs.This paper discusses the adjustment of such kind of catalogs in terms of their special aspects and distinctive qualities of medical education with an attempt to draw the attention of more experts and scholars to this research direction in order to ensure a stable,scientific,and continuous development of the catalog for medical sciences.

99Tc(m) -N(NOEt)2 uptake kinetics difference among KMB17 human embryonic lung diploid fibroblast and different human lung cancer cells / 中国肺癌杂志

<p><b>BACKGROUND AND OBJECTIVE</b>PET/CT imaging is expensive, so searching the tumor imaging agent for SPECT/CT is necessary. 99Tc(m) -N(NOEt)2 [bis (N-ethoxy-N-ethyl dithiocarbamato) nitrido 99Tc(m) (V)] can be uptaken by lung cancer cells and other cells alike. The aim of this study is to evaluate the distinctive value in lung tumor with 99Tc(m) -N(NOEt)2, the difference in its uptake kinetics in human embryonic lung diploid fibroblasts KMB17 and several kinds of lung cancer cells lines.</p><p><b>METHODS</b>Firstly, six different cell culture medium which contained YTMLC Gejiu human lung squamous carcinoma cell, SPC-A1 human lung adenocarcinoma cell, AGZY low metastatic human lung adenocarcinoma, 973 high metastatic human lung adenocarcinoma cell, GLC-82 Gejiu human lung adenocarcinoma cell, and KMB17 human embryonic lung diploid fibroblast, respectively with equal cell density of 1 x 10(6)/mL and the same volume were prepared; secondly, the same radioactive dose of 99Tc(m) -N(NOEt)2 was added into each sample and then 300 microL mixed sample was taken out respectively and cultured in 37 degrees C culture box; Finally, 5 min, 15 min, 30 min, 45 min, 60 min, 75 min, 90 min after cultivation, centrifuged each cultured sample and determined the intracellular radiocounts of each sample, calculated each cell sample's uptake rate of 99Tc(m) -N(NOEt)2 at different time.</p><p><b>RESULTS</b>Statistical difference was found among six cell samples, and the uptake rate sequence from high to low is 973 and SPC-A1 > YTMLC > GLC-82 > AGZY > KMB17 respectively; furthermore, 30 min-45 min after culture, the uptake rate reached stability, and the 45 min uptake rate of each sample was higher than its 96.7% uptake peak.</p><p><b>CONCLUSION</b>Based on the results above mentioned, it is supposed that there are discriminative clinical value when using 99Tc(m) -N(NOEt)2 as a tumor targeting imaging agent, and 30 min or so after injection may be the best imaging time in the early imaging stage.</p>

Argon-helium cryoablation combined with transcatheter arterial chemoembolization for the treatment of primary hepatocellular carcinoma / 介入放射学杂志

Objective To evaluate argon-helium cryoablation combined with transcatheter arterial chemoembolization (TACE) in treating primary hepatocellular carcinoma by comparing the changes of AFP level and T cell immunity after the therapy with that obtained after the treatment of argon-helium cryoablation alone and with that obtained after the treatment of TACE alone. Methods (1) Ninety-nine patients with primary hepatocellular carcinoma were randomly divided into three groups: group A (n=30), treated with argon-helium cryoablation; group B (n=34), treated with TACE; and group C (n=35), treated with argon-helium cryoablation together with TACE. The patients' gender, age and pathology of three groups were comparable with each other. (2) The peripheral blood T cell immunity and AFP level both before and after the treatment were determined and the results were statistically compared. Results After the treatment the AFP level in all 3 groups was significantly reduced when compared to that determined before the treatment (P < 0.05). And the difference in the decrease of AFP level between group C and A, also between group C and B, was statistically significant (P < 0.05). After the treatment the T cell immunity, including Th, TS and Th / TS, in all 3 groups was significantly different from that determined before the treatment (P < 0.01), and significant difference also existed between group C and A and between group C and B (P < 0.01). Conclusion The statistic analysis of AFP and T cell immunity, which are regarded as the index of therapeutic efficacy, indicates that argon-helium cryoablation combined with TACE is superior to simple argon-helium cryoablation and also to simple TACE in the treatment of primary hepatocellular carcinoma.

Investigation on Whole Blood Lead Levels in 2956 Children of 1/12～13 Years old in Wenzhou / 医学研究杂志

Objective To Inves tigate the blood lead levels of children of 1/12～13 years old in Wenzhou and early diagnose lead poisoning in them.Methods The whole blood lead levels of 2956 children aged 1/12～13 years were determined by atom absorbspectral analysis.Results The mean of all blood lead levels was(63.11?30.17)?g/L (2037 boys were 65.55?31.23?g/L,919 girls were 57.72?26.78?g/L),318 children(10.77%) were with a blood lead ≥100?g/L(259 boys,59 girls).The prevalence of lead poisoning in boys(12.7%)as higher than those in girls 6.4%(?~2=18.99,P

Early impact of cryosurgery ablation on the function of T cellular immunity in tumor-bearing rabbits / 介入放射学杂志

Objective To discuss the early impact of cryosurgery ablation on the function of T cellular immunity in tumor-bearing rabbits through observing the changes of T cell subsets after cryosurgery procedure in experimental rabbits.Methods ① Thirty tumor-bearing rabbits were randomly and equally divided into 3 groups:group A,receiving cryosurgical treatment;group B,receiving surgical resection;and group C,used as control group.②Both the preoperative and the postoperative peripheral blood T cell subsets were determine in all experimental rabbits of three groups,the results were compared and statistically analyzed.Results After the procedure,CD8 was significantly decreased in all three groups(P

DSA diagnosis and interventional therapy in acute massive hemorrhage of abdomen and pelvis / 介入放射学杂志

Objective To evaluate the diagnostic angiography and interventional therapy for the different acute massive hemorrhage of abdomen. Methods 80 cases of acute hemorrhage of abdomen and pelvis with clinical data of selective arteriography and endovascular interventional therapy were collected and analized retrospectively. Seldinger technique was adopted for selective visceral angiography via femoral approach with lipiodal, gelfoam and spring coils as the embolic materials. Results All bleeding sites in 80 cases could be confirmed and 68 cases of them were successfully embolized, 9 cases occurred with rehaemor- rhagia and 3 cases were ineffective. Conclusion Interventional therapy not only ascertain the bleeding site, but also stop bleeding. The effect is certainly positive.

Application of radionuclide ~(90)Sr in prevention and treatment of skin scar / 中华医学美学美容杂志

Objective To study the ultrastructural changes of hypertrophic scars and animal wound healing models induced by radionuclide 90 Sr exposure and to get the most effective dosage and time in the prevention and treatment of scars. Methods The clinical hypertrophic scars and animal wound models were exposed using 90 Sr applicator in this study. The exposure doses were 200 800 cGy and 200 4 000 cGy. Then the fibroblastic ultrastructure of the tissues from the experimental and control groups were observed with transmission electron microscope. Results Compared with the control groups, capillaries and fibroblasts obviously increased in small and medium doses (200 600 cGy) groups and fibroblastic function was activated. The fibroblasts decreased and fibroblastic function was inhibited in large dose (800 2 000 cGy) groups. Conclusions Small and medium dose of 90 Sr can accelerate wound healing, and can therefore be used in the treatment of early wounds (2 3 days after wounded) ; large dose of 90 Sr can prevent scars from hyperplasia, and can be used in the wounds of the first week after operation; 1 000 2 000 cGy 90 Sr can cure the old hypertrophic scars or keloids; It is useless that 90 Sr exposes before operation for prevention of scars.The most effective method to prevent scars from hyperplasia is large dose of 90 Sr exposure after operation.

A new spectrophotometric method to determing serum magnesium / 中华检验医学杂志

0 05, The reference inteval ( ?2s ) was 0 70～1 15 mmol/L ( n =67) Conclusions This method is simple, sensitive, rapid and accurate,and suitable for the routine manual or automatic analysis of magnesium in serum

Interventional Therapy of Ischemic Necrosis of Femoral Head / 实用放射学杂志

Objective To study the curative effect of ischemic necrosis of femoral head by interventional thrombolysis.Methods By applying Seldinger's technique,the catheter was guided via arteria femoralis with super-selection into circumflexa femoris medialis,lataralis,and acetabuli to performed the interventional thrombolysis in 50 cases(60sides)with necrosis of femoral head.Results All patients were followed-up for 6~48 months,combined with angiography,the clinical symptoms and the change of bone.The curative effect was evaluated.The ratio of excellent and better effect was 91.4%,the ratio of the improvement of angiography was 87%.the angiography showed that after treatment,the supply artery of femoral head was increased in number;the stained area of femoral head was extended,the capillaries were incereased and the reflux of vein was remarkably improved.The ratio of the improvement of clinical symptom was 96.6%.90% of the necrotic area of femoral head showed hyperplasia,sclerosis,and reduction of cystic changes.Conclusion The interventional therapy by leading to target vascular with thrombolysis in treating necrosis of femoral head has remarkable therapeutic effect.