AIM:To investigate the effect of doublecortin-like kinase 1(DCLK1)on the biological properties of gastric cancer stem cells,and to explore its possible mechanism.METHODS:Serum-free suspension culture of gastric cancer stem cells and targeted inhibition of DCLK1 activity in gastric cancer stem cells with DCLK1 inhibitor DCLK1-IN-1 were performed.The expression levels of DCLK1,stemness-related proteins(SOX2 and OCT4),proliferation-related pro-teins(cyclin D1 and c-MYC),drug resistance-related proteins(ABCG2 and TOP2A),epithelial-mesenchymal transition-related proteins(E-cadherin,vimentin and Snail),and PI3K/AKT/mTOR signaling pathway-related proteins in gastric cancer stem cells were examined by Western blot.The effects of DCLK1 on viability and drug resistance of gastric cancer stem cells were determined by CCK-8 assay,and the effects of DCLK1 on self-renewal of gastric cancer stem cells were de-termined by methylcellulose spheroid-forming assay.Wound-healing and Transwell assays were performed to assess the ef-fect of DCLK1 on the migration and invasion of gastric cancer stem cells.RESULTS:The expression levels of DCLK1 and stemness-related proteins SOX2 and OCT4 in gastric cancer stem cells were significantly higher than those in parental cells(P＜0.01).The proliferation,drug resistance,migration and invasion of gastric cancer stem cells in DCLK1 inhibi-tion group were significantly lower than those in Sphere cell group(P＜0.01).The expression levels of proliferation-related proteins(c-MYC and cyclin D1)and drug resistance-related proteins(TOP2A and ABCG2)were down-regulated,the ex-pression of epithelial marker E-cadherin was up-regulated,the expression of mesenchymal markers vimentin and Snail was down-regulated,and the expression levels of PI3K/AKT/mTOR signaling pathway-related proteins and their phosphoryla-tion levels were reduced in DCLK1 inhibition group(P＜0.05).CONCLUSION:DCLK1 is highly expressed in gastric cancer stem cells,which may be involved in the proliferation,drug resistance and invasion of gastric cancer stem cells by regulating PI3K/AKT/mTOR signaling pathway.It suggests that DCLK1 can be used as a potential target for gastric cancer stem cells.

Objective:This work aims to investigate the phenotype-characteristics of drug resistance and the possible mechanisms of extensively drug-resistance Klebsiella pneumoniae(XDRKP). Methods:Screened by the previous drug susceptibility results, 116 clinical Klebsiella pneumoniae isolates were collected from Shanxi Bethune Hospital from January 2018 to December 2020. Matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS) rapid microbial identification system and VITEK-compact 2 were used. The modified carbapenem inactivation method (mCIM) combining with EDTA carbapenem inactivation method (eCIM) was used to identify the strains′ carbapenemase phenotypes, which were compared with subsequent qPCR results. The qPCR amplification combining with agarose gel electrophoresis were carried out to detect various drug-resistant related genes, including: carbapenemase genes: blaKPC, blaNDM, blaVIM, blaIMP, blaOXA; aminoglycosides resistance genes: ① 16S rRNA methylase genes: rmtA, rmtC, rmtD, rmtG, rmtH, armA, npmA, rmtB, rmtE, rmtF, ② variant of aminoglycosides acetyltransferase gene: aac(6′)-Ib-cr; quinolone resistance genes: DNA gyrase protection protein qnr family: qnrA, qnrB, qnrC, qnrD, qnrS, efflux pump protein gene: oqxAB, qepA, variant of aminoglycoside acetyltransferase gene: aac(6′)-Ib-cr; and tigecycline-resistant Tet protein genes: efflux pump protein gene: tet (A), tet (L), ribosome protection protein gene: tet (M), tigecycline modified enzyme gene: tet (X). Each isolate′s phenotype and resistance gene result were compared and analyzed correspondingly. Results:A total number of 116 XDRKP isolates were collected in 3 years, 115 of which are identified as carbapenem resistant. Both cephalosporins and quinolones resistant rate were 100%, while the resistant rate of aminoglycosides antibiotic gentamicin, tobramycin and amikacin was 95.69% (111/116), 94.83% (110/116), or 88.79% (103/116) respectively. Sulfonamide antibiotics and tigecycline showed a relatively lower resistant rate. Compared with PCR amplification results, mCIM combining with eCIM phenotype testing had a high conformity, up to 95.65% (110/115). Positive rate of each resistance related gene was: blaKPC 90.52% (105/116), blaNDM 10.34% (12/116), rmtB 81.90% (95/116), armA 2.59% (3/116), oqxAB 65.52% (76/116), qnrB 6.03% (7/116), qnrS 12.93% (15/116), aac(6′)-Ib-cr 7.76% (9/116), or tet(A) 21.55% (25/116), respectively. Other resistance related genes were not detected. Corresponding analysis between the resistant phenotypes and resistance related genes indicated that a total of 65 XDRKP didn′t have a matched pairs, i.e. bacteria′s resistance to specific antibiotic could not be interpreted by carrying some associated resistant genes.Conclusions:The wide distribution of resistant genes and multiple-antibiotic-inactivated trait of some genes(such as aac(6′)-Ib-cr and oqxAB) in XDRKP are potential causes of the generation of extensively drug resistant phenotype. Different XDRKP isolates may carry one or more resistant genes in responding to specific antibiotic. In addition, there are some bacteria with an unmatched phenotype-gene feature indicating that both resistance genes′ regulation and some other mechanisms also play a role in development of XDR.

Objectives:To investigate the proportion of MSM among males over 15 years old and analyze its related factors to provide a reference for estimation of MSM size.Methods:Using cross-sectional survey design, multi-stage sampling method, and street interception survey method, a survey was conducted on males over 15 years old in Kunming from October to December 2019, with an estimated sample size of 9 908.Results:Totally, 10 707 males were recruited from 30 sites in 5 counties, and 10 283 were effectively surveyed with a response rate of 96.0%. Respondents aged 16 to 40 accounted for 75.3% (7 748), senior high school or above 71.1% (7 312), and unmarried 49.8% (5 121). The proportion of homosexual behavior in the past half-year was 1.06% (95% CI: 0.86%-1.26%), and the age-adjusted rate was 0.97% (95% CI: 0.78%-1.16%). And multivariate logistic regression showed the associated factors for homosexual behavior as following: proportion of main urban area was 2.217 times (95% CI:1.004-4.895) that of the outer suburbs, registered residence outside Kunming was 0.421 times (95% CI:0.260-0.682) that of in Kunming, having been in Kunming ≤6 months was 2.282 times (95% CI:1.262-4.126) that of >6 months, senior middle school or above was 0.336 times (95% CI:0.228-0.495) that of junior middle school and below, and being married was 0.462 times (95% CI:0.303-0.705) that of unmarried. Conclusions:The proportion of over 15-year-old males who have recently practiced male-male behavior was close to 1.00% in Kunming. The relevant factors included survey areas with a permanent residency of Kumming, short-time residency, education level, and marital status. This study obtained the data and related factors, which provided a reference for estimating MSM size in Yunnan province.

Objective:To investigate the biological characteristics of monoclonal antibodies against avian influenza virus (AIV).Methods:Monoclonal antibodies (mAbs) against AIV H5N1 were prepared and it′s characteristics were identified including subtype,titer,hemagglutination inhibition activity and cross-reactivity with other influenza viruses.Besides,Western blot and immuno-histochemical staining methods were conducted to test the combination of antibodies and antigen ( H5N1 ) and human normal tissues.Results:Immunohistochemical analysis showed that 2 mAbs (H5-32 and H5-35) cross-reacted with human tissues kidney and pancreas respectively.Conclusion:These data indicated that there have some association between the AIV H 5N1 with human tissues, which may provide reference for the study on avian influenza virus infection and pathogenicity .

Objective To evaluate the in vitro synergistic effect of tetrandrine on ketoconazole against Candida parapsilosis complex.Methods According to the Clinical and Laboratory Standards Institute (CLSI) M27-A3 guidelines,the microdilution checkerboard method was used to evaluate in vitro antifungal activities of ketoconazole alone and in combination with tetrandrine against 21 clinical isolates of Candida parapsilosis complex based on the fractional inhibitory concentration index (FICI).Antifungal effects of the above drugs at different time points were evaluated by the XTT assay,and then time-killing curves were drawn and assessed to investigate the in vitro dynamic antifungal activity.Results The minimum inhibitory concentrations (MICs) of tetrandrine and ketoconazole alone against 21 clinical isolates of Candida parapsilosis complex were 32-64 mg/L and 0.031 25-2 mg/L,respectively.When ketoconazole was combined with tetrandrine,MICs of tetrandrine and ketoconazole were reduced to 2-8 mg/L and 0.008-0.25 mg/L respectively,and the FICI ranged from 0.09 to 0.5.The time-killing curves revealed that the fungal growth was delayed obviously in the combination group compared with the ketoconazole alone group and tetrandrine alone group.Conclusion Tetrandrine has obvious synergistic effects on ketoconazole against Candida parapsilosis complex in vitro.

Objective To summarize the efficacy of extracorporeal membrane oxygenation (ECMO)utilization in Emergency Department (ED),as well as the establishment of emergency ECMO team.Methods A retrospective analysis was carried out in 16 patients treated with ECMO between April 2015 to December 2016 in ED.The clinical data including demographics,diagnosis,initiating ECMO timing,place of ECMO establishment,intubation approaches,duration of ECMO,complications and outcomes were collected and analyzed.Results Eight patients were successfully weaned from ECMO,and 7 of them survived to discharge from hospital.The duration of ECMO support was 4 to 384 hours.The emergency ECMO team was set up.Conclusions Emergency medical team can successfully operate the ECMO process.The emergency medical team-initiated ECMO can provide effectively adjuvant measures to support patients with respiratory failure,circulatory failure and cardiac arrest.

A 76?year?old female patient complained of right chest pain for three months. CT scan showed a clump?like high?density shadow measuring 4.8 cm × 3.0 cm in size in the dorsal portion of the right lower lobe of the lung. Aspiration biopsy was performed, and biopsy samples were subjected to fungal culture and histopathological examination. Histopathological examination showed chronic granulomatous inflammation with hyaline septate hyphae. After 4?day culture, white villous dense colonies were formed on the Sabouraud′s agar medium. The center of the colonies was slightly elevated with wrinkles or radiating striae on the surface, and the bottom of the colonies was faint yellow in color. Microculture yielded abundant septate branched hyphae, and very few colorless hyaline quasi?circular spores. DNA sequencing of rDNA internal transcribed spacer (ITS) regions and β?tubulin genes was performed to identify the isolate, and antifungal susceptibility testing was carried out in vitro. The MEGA7.0 software was used to build phylogenetic trees of Aspergillus fumigatus complex and its closely related species. The isolate was identified as Aspergillus fumigatus by molecular biologic sequencing. The patient was diagnosed with pulmonary aspergilloma. After administration of itraconazole oral solution and vorionazole tablets, the condition got better obviously.

In this study,adults of Armillifer agkistrodontis (A.agkistrodontis) were collected from Agkistrodon acutus,and then the eggs were separated to feed mice.In the next step,when the infection model was established,blood serum of infected mice were collected after 1,2 and 3 weeks,respectively.Furthermore,ELISA and dot- ELISA were used to detect the dynamic change of specific antibodies and circulating antigens respectively.The specific antibodies increased from 8th week,reached the top at 12th week,decreased from 16th week,and then maintain at the same level constantly.Meanwhile,the specific antibodies were typed.It is evident that IgM antibody appeared first.However,it was substitute by IgG1 after 16 weeks.Moreover,the circulating antigens have been detected in the 1st week by dot-ELISA.Then,the dilution between 1:8 to 1:128were founded in 3rd week.The highest dilution with 1:256 appeared at 8th week,maintained before 11th week and then decreased gradually,which might provide a significant clinical implication for early diagnosis of circulating antigens.

ObjectiveComparing gastric perforation repair with traditional contrast,to explore the feasibility and superiority of the laparoscopic gastric perforation repair.Methods68 cases were randomly divided into two groups for laparoscopic gastric perforation repair and traditional repair,then compared two groups of treatment.Results Both operations were successful ( including laparoscopic repair in 34 cases) and surgery time,blood loss,postoperative drainage,length of stay,and cosmetic results of the comparison.ConclusionCompared with the traditional open surgery,the laparoscopic surgery had less trauma,leas pain,faster recovery,shorter hospital stay,high efficacy and good cosmetic results and other advantages.

Objective To estimate the application value of colony PCR in the detection of pathogenic filamentous fungi. Methods Colony PCR was established and performed to amplify the internal transcribed spacer (ITS) region of 19 species (strains) of filamentous fungus followed by sequencing analysis. At the same time, DNA extracts from 8 of the 19 species of filamentous fungus were subjected to conventional PCR. Hha I and Hinf I endonucleases were used for restriction fragment length polymorphism (RFLP) analysis of the conventional and colony PCR products. Comparison analysis was carried out between the colony and conventional PCR. Results Of the 19 strains, 16(84.2%) yielded positive results by colony PCR; sequence analysis of the PCR products of ITS region revealed a 96% - 100% similarity with the reference sequence (NCBI database)of corresponding fungi. The amplification product length and RFLP profile of these products from the 8 species of filamentous fungus, except for those from Aspergillus nidulans, were consistent between the colony and conventional PCR. Conclusions Compared with conventional PCR, colony PCR-based detection of filamentous fungi is easy to operate, time and labor-saving, with high accuracy and reliability, and can be applied to the rapid identification of filamentous fungi.