ABSTRACT
Objective To investigate antibiotics resistance of Proteus mirabilis isolated from stools of patients with acute diarrhea for the prevention and treatment of its infection and the rational use of antibiotics. Methods Stool samples of acute diarrhea patients were collected in the diarrhea outpatient clinic of the Second Hospital of Tianjin Medical University and Tianjin Medical University General Hospital from 2013 to 2014. Enrichment culture and biochemical identification were used to isolate and identify Proteus mirabilis, which were further performed antimicrobial susceptibility testing and class 1 integron detection. Extended spectrum β-lactamases (ESBLs) phenotype and ESBLs genes (TEM, OXA and CTX-M) were amplified by polymerase chain reaction (PCR), and sequencing were carried on in parts of suspected isolates. ESBLs-positive strains were analyzed by pulsed-field gel electrophoresis (PFGE). Results A total of 277 strains of non-repetitive Proteus mirabilis were isolated, and 268 of them were performed antimicrobial susceptibility testing (the remaining 9 strains failed to recover). Relative higher resistant rates were trimethoprim/sulfamethoxazole (30.2%), ampicillin (25.4%), nalidixic acid (25.7%), streptomycin (21.6%) and chloramphenicol (21.3%). The multiple drug resistance rate was 24.6% (66/268). The positive rate of class 1 integron was 22.8%(61/268). Resistance rates to third-generation cephalosporin, ciprofloxacin and imipenem were less than 10%, but 4 isolates were resistant to imipenem, third-generation cephalosporin, fluoroquinolones, trimethoprim/sulfamethoxazole, and chloramphenicol simultaneously. Three cefotaxime-resistant strains (1062, 1505 and 1650) were positive for ESBLs phenotype and harbored CTX-M extended-spectrum β-lactamase genes, among them 2 strains also carried TEM and/or OXA β-lactamase genes. The clustering analysis of pulsed-field gel electrophoresis (PFGE) displayed that the similarities between 1505 and 1650 were 85.7%, and the similarity with 1062 was 58.1%. Conclusion Proteus mirabilis isolated from patients with acute diarrhea in our city show significant multidrug resistance, high positive rate of class 1 integron, and emergence of ESBLs-positive strains resistant to imipenem and fluoroquinolones, which pose a threat to public health. Rational use of antibiotics is important in both clinical and nonclinical settings.
ABSTRACT
<p><b>OBJECTIVE</b>To establish an improved three-dimension (3D) and serum-free approach to differentiate human embryonic stem cells (hESCs) into endothelial cells, and detect the endothelial functions of the obtained cells.</p><p><b>METHODS</b>We cultured undifferentiated H9 human embryonic stem cell line in low-adhesion dishes to form embryonic bodies (EBs). After 12 days, EBs were harvested, re-suspended into rat tail collagen type I, and put into the incubator (37℃). After 30 minutes, EGM-2 culture medium was added to the solidified collagen, and the EBs were cultured for another 3 days to form embryonic body-sproutings (EB-sproutings). EB-sproutings were digested with 0.25% collagenase I and 0.56 U/ml Liberase Blendzyme for 20 minutes respectively, and the CD31(+) cells were sorted by FACS. The endothelial functions were tested by Dil-ac-LDL uptake assay and tube formation assay.</p><p><b>RESULTS</b>This approach raised the efficiency of endothelial differentiation to 18%, and also avoided the contamination with animal materials. The obtained hESC-derived endothelial cells (hESC-ECs) had the similar pattern of surface biomarkers as human umbilical vein endothelial cells (HUVECs), and their endothelial functions were confirmed by the uptake of Dil-ac-LDL and the tube formation on Matrigel.</p><p><b>CONCLUSIONS</b>The improved 3D approach can enhance the efficiency of differentiation from hESCs into endothelial cells. Furthermore, serum free differentiation system may be applied in future hESC-based therapies for various ischemic diseases.</p>