Objective To establish a viable bacteria assay for Helicobacter pylori(H.pylori)by assessing the cgt gene expression,and to develop accordingly a rapid and novel testing method for clinical precision treatment.Methods Viable bacteria count was determined in bacterial cultures.The transcriptional expression level of cgt(hp0421),the conserved gene that encodes cholesterol-α-glucosyltransferase(CGT)in H.pylori,was measured by RT-PCR.The correlation between the number of colonies and cgt gene transcription expression was analyzed and the regression model was constructed.The linear range,sensitivity,and specificity of the new method were examined accordingly.The bactericidal action of clarithromycin was assessed using this method to verify the performance of the method in determining clinical bacterial drug resistance.Results The Ct values of cgt for H.pylori colony counts of 102,104,106,and 108 CFU/mL were 29.67±0.14,23.37±0.36,17.65±0.37,and 11.38±0.39,respectively.In the range of 101-108 CFU/mL,the regression equation for cgt gene expression and viable bacterial counts determined by RT-qPCR was y=-0.350 1x+12.49,with the correlation coefficient being R2=0.9992 and the sensitivity being 101 CFU/mL,showing no cross-reaction with 13 other bacteria.The lg values of live H.pylori bacteria treated with clarithromycin at 0,5,10,20,and 40 μg/mL for 12 h were 2.57±0.02,2.45±0.01,2.19±0.02,1.91±0.07,and 1.33±0.05,respectively.The corresponding cgt gene Ct values were 27.76±0.09,28.37±0.24,29.51±0.14,30.11±0.12,and 31.66±0.11.By applying the cgt gene expression in the equation,the estimated counts of viable bacteria were found to be 2.73±0.03,2.52±0.08,2.11±0.05,1.89±0.02,and 1.33±0.04,showing no significant difference in statistical analysis(P＞0.05).Conclusion The method for assessing viable bacteria account by evaluating cgt gene expression in H.pylori was successfully established,significantly reducing the time required to determine viable bacteria count and providing a new method for clinical viable bacteria testing.

Helicobacter pylori(H.pylori),for a long time,has generally been considered an extracellular bacterium.However,recent findings have shown that H.pylori can gain entry into host cells,evade attacks from the host immune system and the killing ability of medication,form stable intracellular ecological niche,and achieve re-release into the extracellular environment,thus causing recurrent infections.H.pylori intracellular infection causes cellular signaling and metabolic alterations,which may be closely associated with the pathogenesis and progression of tumors,thereby presenting new challenges for clinical eradicative treatment of H.pylori.Herein,examining this issue from a clinical perspective,we reviewed reported findings on the mechanisms of how H.pylori achieved intracellular infection,including the breaching of the host cell biological barrier,immune evasion,and resistance to autophagy.In addition,we discussed our reflections and the prospects of important questions concerning H.pylori,including the clinical prevention and control strategy,intracellular derivation,and the damage to host cells.

To carry out phylogenetic analysis for drug-resistance genes from clinical isolates of Helicobacter pylori (Hp) among patients with gastric diseases from Tibet, China.

METHODSHp strains were isolated and cultured from saliva and gastric mucosal tissues derived from patients with gastric diseases. Nine strains (including 5 isolated from oral tissues, 1 isolated from gastric tissues, and 3 representative strains of SS international standard strains used for animal models) were tested for common antibiotic resistance. Together with an ACTT 11637 international standard strain, these were subjected to re-sequencing to obtain drug-resistance genes. Such genes from various sources were compared with the resistance genes of Hp strains recorded by the NCBI website. Combined with results of drug-resistance experiments, correlation between molecular evolution and drug-resistance was analyzed.

RESULTSTesting of gastric mucosal tissues and salivary samples from 217 patients has found 89 Hp strains, which yielded a total infection rate of 41.01%. The resistance rates of 9 representative Hp strains for clarithromycin, amoxicillin, metronidazole, levofloxacin and tetracycline were 77.8%, 77.8%, 44.4%, 77.8%, and 77.8%, respectively. Compared with the reference strain, the similarity between clarithromycin-resistance genes was 99%, and that between amoxicillin- and metronidazole-resistance genes was 96%-97%. A2143G mutation was also found in clarithromycin-resistant genes of three Hp strains.

CONCLUSIONThe sensitivity of Hp to metronidazole is much higher in patients from Tibet region, and the sensitivity of Hp to clarithromycin, amoxicillin, levofloxacin and tetracycline is poor. Resistance mutations are consistent with drug resistance.

Objective To investigate the epidemiological characteristics of imported malaria in Suining City from 2011 to 2016,so as to provide the evidence for improving the surveillance,risk assessment and control measures of imported malaria. Methods The malaria epidemic data in Suining City from 2011 to 2016 were collected from the reporting system,and statisti-cal analyzed. Results Totally 71 malaria cases were reported in Suining City from 2011 to 2016,and they were all imported cases. Sixty cases(84.51％)were imported from Africa. There were cases reported throughout the year,and no obvious seasonal distribution characteristics. The median time from onset to seeing a doctor was 2 days;the median time from seeing a doctor to confirmed diagnosis of malaria was 4 days. Totally 63.38％of the cases were diagnosed of malaria in initially seeing a doctor. For-ty-three cases were diagnosed by county and municipal medical institutions,accounting for 60.56％. All cases were laboratory confirmed cases,and they all received the standardized treatment. Conclusion The imported malaria epidemic situation in Su-ining City is still severe,and it is necessary to strengthen the management of floating population and professional training of med-ical staff so as to prevent the second malaria cases and deaths.

ObjectiveTo probe the feasibility and therapeutic effect of the lateral thigh perforator flap for repairing soft tissue defects in extremity limbs.MethodsDissected six fresh Chinese adult lower limbs specimens which were injected with red latex via arterial cannula. Obseved the number, distribution,course, category of the flaps'perforators and measured their length of pedicle, external diameter on the specimens.Combinafing the results of the anatomy study,thirteen lateral thigh perforator flaps,whose size were 3.0 cm × 3.0 cm - 19.0 cm× 8.0 cm, were applyied to repair different defect, in which 1 was pedieled flap and 12 were free flaps.Two of them applied vessel anastomosis in series with anterlateral thigh perforator flap. The axis was from the fibulae capitulum to greater trochanter of femur.ResultsAll 13 flaps surived com-pletely.In 3-6 months follow-up,all flaps were with thin and good appearance,and they were characterized by soft texture and good colour.No chimatlon and ulcer appeared in any flap.There were no effect of motor function in donor site because of muscle conglutination and scar contracture. There were plainness figure in donor site where skin- grafing was used, and only linearity scar remained where sewed directly.ConclusionThe lateral thigh flap had provided with thin and pleasing appearance, soft texture and minor morbidity in donor site and correspondingly easy dissection. It was feasiblely to repair middling to little skin and soft tissue defect in extremity limbs as a free flap, and to repair skin and soft tissue defect in knee and popliteal space as a pedicel flap.It could apply vessel anastomosis in series with anterlateral thigh perforator flap for repairing large defect.

Objective To explore the method of sensory reconstruction after the operation of reversed island pedicled sural flap and evaluate its therapeutic effect of clinical application. Methods Thirteen clinical cases with traumatic soft tissue defects in heel had recepted the treatment of reversed island pedicled sural flap. All flaps were innervated by anastomosing the distal end of the sural nerve in the flaps and the recipient nerve (superficial peroneal nerve) in end to end or end to side. All patients were evaluated at 9-15 months on the postoperative follow-up parameters, including flap contour, flap stability, locomotor activity,touch sensation, pain sensation, static two-point discrimination, thermal sensibility, and the skin sensory recovery level in lateral dorsutn of foot. Results Thirteen cases flaps had good blood supply and primary healing. All cases were followed up 9-15 months, the rate of good sensory recovery was 53.85%. All pa tients had protective sensory in lateral dorsum of foot, the rate of good sensory recovery was 61.54%. Conclusion Anastomosing the proximal end of sural nerve and superficial peroneal nerve together will be good for the sensory recovery in flap and lateral foot in repairing soft tissue defects in heel with reversed island pedicled sural flap.

Objective To investigate the effects of acidosis on the current change of transient receptor potential channel M7 (TRPM7) and collagen production in mouse cardiac fibroblast (MCFs), and to explore the pathophysiological function of TRPM7 on the cardiac fibrosis. Methods (1) The model of MCFs was established and isolated. (2) MCFs was subcuhured. (3) Patch clamp technique was used to observe the current characteristics of TRPM7 in low PH solutions. (4) The influence of acidic condition on Ca2+ influx in MCFs was recorded by calcium fluorescent indicators. Results (1) There was a high level expression of TRPM7 in MCFs and the electrophysiological characteristics of TRPM7-like (TRPM7L) was similar to that of TRPM7. (2) Ca2+ influx was increased in acidic condition, and the F340/F380 ratio was increased from 1.0 to 4. 6 at pH 4.0 compared with pH 7.0 (t=2.72, P<0.01). Conclusions (1) TRPM7 is the molecular basis of the native TRPM7L in MCFs and TRPM7L plays an important role in Ca2+ influx. (2) The pathophysiological function of MCFs is influenced by regulation of Ca2+ influx mediated by TRPM7L in the condition of acidosis.

Objective To investigate the changes of TRPM 7-like current in mouse cardiac fibroblast(CFs) after myocardial infarction and the effect of myocardial ischemia on the TRPM 7 expression and current. Methods (1) The model of myocardial infarction was made and CFs were isolated;(2) CFs were cultured and infected by TRPM 7 siRNA;(3) The effects of myocardial ischemia on TRPM 7 current were recorded by whole cell patch clamp technique;(4) The influences of myocardial isehemia on Ca2+ influx in CFs were recorded by Ca2+ fluorescence imaging. (5) The effects of ischemia on total collagen content of CFs were studied. Results (1) Ca2+ inward current of CFs was increased after myocardial infarction [(16.2±1.7) vs. (7.4±0.7) pA/pF, P<0.053];(2) TRPM 7 current was reduced by 90% after siRNA infection;(3) The total collagen content of CFs after ischemia was approximately 2.3-fold higher than basic value. Conclusions Ca2+ influx in CFs plays an important role in the pathophysiological process of myocardial fibrosis.

Objective To investigate the pathological contribution of serological markers of collagen metabolism including matrix-metalloproteinases(MMPs)and its tissue inhibitors(TIMPs),the markers for type I collagen degradation(ICTP)and synthesis(PICP)to cardiac remodeling of ischemic cardiomyopathy(ICM)as well as the pharmacological regulation effect.Methods Plasma levels of MMP-9,TIMP-1,ICTP and PICP were determined by ELISA and RIA in 86 consecutive ICM patients and 25 age-matched control subjects.Patients were divided into two subgroups according to ACEI-taking or not.Echocardiography were performed in all cases.Results The plasma concentration of MMP-9、TIMP-1 and MMP-9/TIMP-1 was significantly increased in ICM group(all P