Applications and developments of gene therapy drug delivery systems for genetic diseases

Genetic diseases seriously threaten human health and have always been one of the refractory conditions facing humanity.Currently,gene therapy drugs such as siRNA,shRNA,antisense oligonucleotide,CRISPR/Cas9 system,plasmid DNA and miRNA have shown great potential in biomedical applications.To avoid the degradation of gene therapy drugs in the body and effectively deliver them to target tissues,cells and organelles,the development of excellent drug delivery vehicles is of utmost importance.Viral vectors are the most widely used delivery vehicles for gene therapy in vivo and in vitro due to their high transfection efficiency and stable transgene expression.With the development of nanotechnology,novel nanocarriers are gradually replacing viral vectors,emerging superior performance.This review mainly illuminates the current widely used gene therapy drugs,summarizes the viral vectors and non-viral vectors that deliver gene therapy drugs,and sums up the application of gene therapy to treat genetic diseases.Additionally,the challenges and opportunities of the field are discussed from the perspective of developing an effective nano-delivery system.

Drug resistance gene expression and chemotherapy sensitivity detection in Chinese women with different molecular subtypes of breast cancer

Objective:The aim of the study was to identify specific chemosensitivity drugs for various molecular subtypes of breast tumors in Chinese women,by detecting the expression of drug resistance genes and by using the drug sensitivity test on different molecular subtypes of breast cancers.Methods:The expression of drug resistance genes including Topo Ⅱ,GST-π,P-gp,LRP,and CD133 were detected with immunohistochemistry in a tissue microarray.Drug sensitivity tests included those for paclitaxel,epirubicin,carboplatin,vinorelbine,and fluorouracil and were conducted on primary cancer tissue cells and cell lines,including the T47 D,BT-474,and MDA-MB-231 cells and human breast cancer xenografts in nude mice.Results:The different drug resistant genes Topo Ⅱ,GST-π,P-gp,and LRP were differentially expressed among different molecular subtypes of breast cancers(P<0.05).Positive expression of CD133 was highest in basal-like breast cancer(P<0.05).Kaplan-Meier survival analysis showed that positive expressions of Topo Ⅱ and CD133 both correlated with shorter disease-free survival(DFS)(P<0.05)and overall survival(P<0.05),and positive expression of LRP correlated only with shorter DFS(P<0.05).BT-474 showed chemosensitivity to paclitaxel and epirubicin,while MDA-MB-231 showed chemosensitivities to paclitaxel,epirubicin,carboplatin,and fluorouracil(T/C≤50%).The basal-like and HER2+breast cancer primary cells showed chemosensitivities to paclitaxel and epirubicin with significant differences compared with luminal breast cancer primary cells(P<0.05).Conclusions:The differential expression of drug resistance genes and the differential chemosensitivities of drugs in different molecular subtype of breast cancers suggested that individual treatment should be given for each type of breast cancer.

Transduction of Fas gene or Bcl-2 antisense RNA sensitizes cultured drug resistant gastric cancer cells to chemotherapeutic drugs

AIM To compare the expression level of Fas gene and Bcl-2 gene in gastric cancer cells SGC7901 and gastric cancer MDR (multidrug resistant) cells SGC7901/VCR, to transduce Fas cDNA and Bcl-2 antisense nucleic acid into SGC7901/VCR cells respectively, and to observe the expression of two genes in transfectants and non-transfectants as well as their drug sensitivity.METHODS Eukaryotic expression vector pBK-Fas cDNA and pDOR-anti Bcl-2 were constructed and transfected into SGC7901/VCR cells by lipofectamine, respectively. Northern blot and Western blot were used to detect the expression of mRNA and protein in SGC7901/VCR and SGC7901 cells and transfectants, and drug sensitivity of transfectants for VCR, CDDP and 5-FU was analyzed with MTT assay.RESULTS After gene transfection, 80 for Fas and 120 for antisense Bcl-2 drug-resistant clones were selected from 2×105 cells, transfection rate being 0.04% and 0.06%. Two clones of SGC7901 Fas/VCR cells and SGC7901 anti Bcl-2/VCR cells were randomly selected for further incubation. Hybridization results showed that the expression level of Fas mRNA and protein in SGC7901/VCR cells was much lower, but that of Bcl-2 mRNA and protein was higher than that in SGC7901 cells. The expression of Fas mRNA and protein in SGC7901 Fas/VCR cells was higher, and of Bcl-2 mRNA and protein was lower in SGC7901 anti Bcl-2/VCR cells than that in non-transfectants. MTT assay showed that transfectants were more sensitive to VCR, CDDP, 5-FU than non-transfectants.CONCLUSION Bcl-2 gene displayed high expression while Fas gene had low expression in drug resistant gastric cancer cells. Expression of Bcl-2 protein was effectively blocked in SGC7901 anti Bcl-2/VCR cells by gene transfection. In contrast, the expression of Fas mRNA and protein in SGC7901 Fas/VCR cells increased. Fas gene and Bcl-2 antisense nucleic acid transfection sensitized drug resistant gastric cancer cells to chemotherapeutic drugs. These results suggest cell apoptosis plays an important role in the mechanism of MDR, and enhancing apoptosis might reverse MDR.

Clinical Study of Multi-drug Resistance Gene(MDR1) Expression in Primary Ovarian Cancer

This study was designed to measure the multi-drug resistance gene (MDR1) mRNA content and analyze clinical relationship between MDR1 expression and drug resistance in primary ovarian cancer. Reverse transcription PCR (RT-PCR) was used to measure MDR1 mRNA content in biopsy sample of 31 primary ovarian cancers (experimental group) and 30 gynecological tumors (control group). The level of 95.2% (20/21) MDR1 expression was relatively low, and the detected rate of MDR1 expression was 67.7%(21/31) in experimental group,which was higher than that in control group (40.0%, P<0.05). The differences of MDR1 expression between the effective group and no effect group after combined chemotherapy was significant (P<0.05). No significant relationship was found between MDR1 expression and clinical stage or histological classification or grade of differentiation in experimental group. We are led to concluded that primary ovarian cancers have drug-resistance clones which might express MDR1 spontaneously and expression of MDR1 may be used as a prognostic and predictive indicator for clinical response of ovarian cancers to combined chemotherapy.

EXPRESSION AND REVERSION OF DRUG RESISTANCE-AND APOPTOSIS-RELATED GENES OF A DDP-RESISTANT LUNG ADENOCARCINOMA CELL LINE

Objective: To investigate the co-expression of drug resistance- and apoptosis-related genes of cisplatin (CDDP)-selected lung adenocarcinoma cell line A 549 DDP for compared to the parental cell line A549, and reverse of drug resistance by antisense s-oligodeoxynucleotides (S-ODNs) of differentially expressed genes. Methods: Sense and antisense S-ODN were transferred into A 549 DDP cells by lipofectin. The expression of drug resistance and apoptosis related genes was examined by RT-PCR, immunocytochemistry and flow cytometry, respectively. Apoptostic cells were identified by DNA electrophoresis and terminal deoxynucleotidyl transferase (TdT)-mediated biotin dUTP nick end-labeling(TUNEL). Drug resistance of tumor cells was detected by a cell viability (MTT) assay. Results: The expression of bcl-2 was positive and that of multidrug resistance-associated protein (MRP) at mRNA and protein level was increased in A 549 DDP compared to A549 cells. MDR1, c-myc and topoisomeras II (TOPO II) were similarly co-expressed in two cell lines. Both cell lines were negative for c-erbB-2 expression. In A 549 DDP cells, the expression of bcl-2 and MRP was significantly inhibited by their respective antisense S-ODNs. Antisense S-ODNs could also decrease significantly drug resistance of A 549 DDP cells to CDDP by promoting cell apoptosis. Conclusion: Both intrinsic and acquired drug resistance were involved in co-expression of multiple MDR-related genes in lung adenocarcinoma. Cooperation of bcl-2 and MRP genes appeared to play an important action to confer the resistance of A 549 DDP cells to CDDP. Their antisense S-ODNs are responsible for the decrease of drug resistance of this cell line by promoting apoptosis.

Expression and Prognostic Significance of Multidrug Resistance Associated Protein (MRP) Gene in Non-small Cell Lung Cancer by in Site Hybridization

Objective: To study on the effect of MRP gene overexpression on prognosis of patients with non-small lung cancer (NSCLC). Methods: Paraffin-embedded tissues from 47 cases of NSCLC who had undergone radical tumor resection were examined for expression of MRP gene mRNA by in situ hybridization using labelled digoxigenin probes combined with immunohistochemistry. All the patients were retrospectively followed-up. Results: All of the 47 lung cancer specimens were found to have overexpression of MRP gene mRNA. It was significantly correlated with patients' survival time, response to chemotherapy, recurrence or metastases after surgery, but was not correlated with histology, tumor size, node status, TNM stage, degree of differentiation, age and sex. Conclusion: Overexpression of MRP gene is a marker of prognostic significance in patients with NSCLC.

Liposome-mediated Functional Expression of Multiple Drug Resistance Gene in Human Bone Marrow CD34^+ Cells

Summary: The expression and functional activity of multiple drug resistance (MDR1) gene in human normal bone marrow CD34+ cells was observed. Human normal bone marrow CD34+ cells were enriched with magnetic cell sorting (MACS) system, and then liposome-mediated MDR1 gene was transferred into bone marrow CD34+ cells. Fluorescence-activated cell sorter was used to evaluate the expression and functional activity of P-glycoprotein (P-gp) encoded by MDR1 gene. It was found that the purity of bone marrow CD34+ cells was approximately (91±4.56) % and recovery rate was (72.3±2.36) % by MACS. The expression of P-gp in the transfected CD34+cells was obviously higher than that in non-transfected CD34+ cells. The amount of P-gp in non-transfected CD34+ cells was (11.2±2.2) %, but increased to (23.6±2.34) % 48 h after gene transfection (P<0.0l). The amount of P-gp was gradually decreased to the basic level one week later. The accumulation and extrusion assays showed that the overexpression of P-gp could efflux Rh-123 out of cells and there was low fluorescence within the transfected cells. The functional activity of P-gp could be inhibited by 10 μg/ml verapamil. It was suggested that the transient and highly effective expression and functional activity of P-gp could be obtained by liposome-mediated MRD1 transferring into human normal bone marrow CD34+ cells.

THE CLINICAL SIGNIFICANCE OF MULTIDRUG RESISTANCE GENE(mdr1) EXPRESSION IN ACUTE LEUKEMIA

Objective: To study the clinical significance of multidrug resistance gene expression in acute leukemia. Methods: The relationships between drug resistance of leukemia cells and prognosis, multidrug resistance gene (mdr1) were examined in 85 patients with acute leukemia and 20 normal controls by reverse transcriptase polymerase chain reaction (RTPCR). Results: The mdr1 positive rate in untreated group was 44.7%. The complete remission (CR) rate of mdr1 positive patients (23.9%) was significantly lower than that of mdr1 negative patients (88.5%) (P<0.005). The mdr1 expression level in relapsedrefractory group was higher than that of CR group. A gradually increased mdr1 mRNA level in CR patients indicated early relapse. Conclusion: The mdr1 positive rate in normal control and longterm survival patients was very low. The mdr1 expression was correlated with FrenchAmericanBritish Cooperative Group (FAB) classification. The mdr1 expression level was correlated with chemotherapeutic effect and prognosis. It is an unfavorable prognostic factor for patients with acute leukemia.

Anticancer Drug Resistance of HeLa Cells Transfected With Rat Glutathione S-transferase pi Gene

To establish a cytologic expressing system of rat glutathione S-transferase pi (GST-pi) cDNA for detecting the resistance of HeLa cells to anticancer drugs. Methods The assessment was made with various anticancer drugs (adriamycin, mitomycin, cisplatinum and vincristine) that showed different cytotoxicities in transfectant HeLa cells with pSV-GT containing rat GST-pi cDNA (HeLa/pSV-GT) or control pSV-neo (HeLa/pSV-neo). Expression levels of GST-pi mRNA in HeLa/pSV-GT and HeLa/pSV-neo were measured by in situ hybridization using Digoxin-labelled cDNA probe. Results HeLa/pSV-GT expressed significantly high degree of GST-pi mRNA, whereas both HeLa/pSV-neo and HeLa cells had very low expression. Cytotoxicities of HeLa/pSV-GT and HeLa/pSV-neo with 4 anticancer drugs were measured by MTT assay. Drug concentrations for yielding 50% inhibition (IC50) in HeLa/pSV-GT by adriamycin, mitomycin and cisplatinum were 70.13 靏/mL, 10.95 靏/mL and 16.52 靏/mL, respectively. In contrast, IC50 in HeLa/pSV-neo was 10.34 靏/mL, 7.48 靏/mL and 13.70 靏/mL, respectively. The cytotoxicities of vincristine on both HeLa/pSV-GT and HeLa/pSV-neo were not significantly different. Conclusions Our findings suggest that HeLa/pSV-GT containing rat GST-pi cDNA is resistant to some anticancer drugs due to overexpression of GST-pi. Also, HeLa/pSV-GT cell line could serve as a useful cytogenetic model for further research.

In vitro study of the influence of GST-π gene transfer on drug-resistance of human cord blood CD34^+ cells

Objective:To investigate the influence of GST-π gene transfer on drug-resistance of human cord blood CD34 + cells.Methods:CD34 + cells were purified from cord blood from normal full-term pregnancy.Gene transduction into human cord blood CD34 + cells was carried out using GST-π gene containing retrovirus vector.The GST-π gene expression in transduced CD34 + cell was confirmed by RT-PCR.After confirmation of GST-π gene transfer,the transfected CD34 + cells were cultured by colony assay in the presence of carboplatin.Results:GST-π mRNA was detected in 30% of CFU-GM derived from GST-π gene transduced CD34 + cells.In vitro drug resistance test showed that the number of CFU-GM formed was significantly higher (2～3 fold) in GST-π gene transduced CD34 + cells than untransduced CD34 +cells.Conclusion:GST-π gene transfer can confer resistance to hematopoietic progenitors against carboplatin in vitro.

2019—2021年皖南地区地方品种鸡源沙门菌的血清型鉴定、毒力基因和耐药性检测

为了分析皖南地区地方品种鸡(淮南麻黄鸡、淮北麻鸡、霍邱鸡和五华鸡)来源沙门菌的分布、毒力基因和耐药性情况,本试验于2019—2021年采集该地区地方品种鸡养殖场疑似沙门菌感染病死鸡的肝脏组织、肛拭子和死胚等病料组织456份,采用细菌分离培养、形态学观察、生化试验和特异性invA基因的PCR检测进行沙门菌鉴定,采用Kauffmann-White法、PCR法和K-B纸片法分别检测分离菌株的血清型、毒力基因、耐药基因和耐药性。结果显示,共分离得到127株沙门菌,分属于13种血清型,其中以鸡白痢沙门菌、禽伤寒沙门菌和肠炎沙门菌为流行的优势血清型,分别占分离菌株的26.0%、20.5%和17.3%;127株沙门菌中携带的4种毒力基因(spvA、spvB、spvC和spvR)检出率介于7.1%~74.8%,携带的4种毒力岛基因(SPI-1 SPI-2、SPI-3和SPI-5)检出率介于38.6%~81.1%;127株沙门菌对阿莫西林、氨苄西林和磺胺二甲氧嘧啶等8种药物的耐药率介于56.7%~99.2%,以耐10(12.6%)、9(15.0%)和8(20.5%)种药物为主,耐药基因sul 1、sul 2、sul 3、tet(A)、tet(M)和tet(R)检出率介于32.3%~89.8%;经分析,地方品种鸡流行优势血清型与毒力基因和耐药基因均有一定的相关性。结果表明,从安徽省皖南地区4种地方品种鸡中分离的沙门菌具有多种血清型、携带多种毒力基因、耐药性严重,且携带多种耐药基因。

1株犬源多重耐药粪肠球菌的分离鉴定及其致病性分析

[目的]明确1例犬泌尿生殖道感染的发病原因。[方法]无菌采集病犬尿液,通过细菌分离纯化、革兰染色镜检、生化试验、16S rRNA基因扩增进行鉴定,同时经药敏试验、毒力基因和耐药基因检测及细菌致病性试验测定纯化菌株的生物学特征。[结果]分离得到1株兼性厌氧型革兰阳性球菌;菌落形态如针尖大小、表面光滑、半透明、圆形凸起、边缘整齐;生化鉴定结果显示,致病菌能发酵乳糖、果糖、蔗糖、麦芽糖、葡萄糖、山梨醇、甘露醇、精氨酸,不能发酵木糖、氧化酶、枸橼酸盐、棉籽糖和鸟氨酸脱羧酶;16S rRNA基因序列与已发表的粪肠球菌序列相似性高于99%,鉴定为粪肠球菌,并将其命名为EFGZ23GY01;药敏试验结果显示,分离株仅对万古霉素及氯霉素敏感,对新霉素中度敏感,对克林霉素等25种药物耐药;8种粪肠球菌毒力基因检测结果显示,分离株携带efaA、ace、asa 1及gelE毒力基因;14种耐药基因检测结果显示,分离株携带tetL、sulⅠ、tetC、bla TEM、bla OXA-23、aadA 1和sulⅢ耐药基因;致病性试验结果显示,5只小鼠注射菌液12 h后出现精神萎靡,被毛杂乱等症状,未出现死亡,48 h后死亡1只,表明分离菌对小鼠具有致病性。[结论]本研究成功从犬尿液种分离得到1株粪肠球菌,其携带多种毒力基因和耐药基因,且对小鼠有致病性,研究为病犬的后续治疗提供了参考。

施用猪粪水对青脆李果实品质和产量、土壤中微生物群落与抗生素耐药基因的影响

【目的】为了解猪粪浇灌对青脆李果实品质和产量、土壤性质、土壤中微生物群落与抗生素耐药基因的影响。【方法】以5年生青脆李作试材,设置猪粪水按不同比例(100%、75%、50%、25%、0%)替代化肥田间试验,通过单果重、单果横径等外观指标,李子每667 m^(2)产量,维生素C、可溶性固形物等果实品质指标测定,分析不同施肥处理对青脆李果实品质和产量的影响;基于土壤中pH、有机质含量等理化指标测定,分析不同施肥处理对李树土壤理化性质的影响;基于Illumina Hiseq 4000测序平台对李树土壤宏基因组测序,分析猪粪水浇灌对土壤中微生物群落变化、抗生素耐药基因分布的影响。【结果】不同施肥处理青脆李果实品质和产量研究结果表明,“75%猪粪+25%化肥”“50%猪粪+50%化肥”处理青脆李果实外观品质均较优,“50%猪粪+50%化肥”处理青脆李每667 m^(2)产量最高。不同施肥处理对李树土壤理化性质的研究结果显示,各处理间有机质、碱解氮等理化指标均不存在显著性差异(P>0.05)。宏基因组测序分析结果显示,“100%猪粪”组中的优势菌群为放线菌,而其余4组的优势菌群均为假单胞菌门;共鉴定39个抗生素本体(ARO),随着猪粪水用量的减少,土壤样品中检出ARO种类、ARO相对丰度和均呈依次递减趋势。【结论】本研究条件下,猪粪水部分替代化肥为青脆李树施肥是可行的,且以“50%猪粪+50%化肥”进行施肥最佳。

北京某医院感染性腹泻患者致泻性大肠埃希菌毒力和耐药特征分析

目的明确本院感染性腹泻患者致泻性大肠埃希菌毒力和耐药特征。方法采用VITEK MS微生物质谱检测系统初步鉴定,多重实时荧光PCR检测毒力基因,对本院感染性腹泻患者临床分离的致泻性大肠埃希菌(Diarrheagenic Escherichia coli,DEC)进行5种型别鉴定。微量肉汤稀释法和E-test法药敏试验检测DEC菌株的耐药表型特征。二代测序及生物信息学分析其耐药分子特征。采用Fisher确切概率法进行统计学分析。结果本院DEC检出率为11.9%,其中EAEC占比37.5%,非典型EPEC占比34.38%,ETEC占比25.0%,EIEC占比3.12%,未检出EHEC菌株。32株DEC对氨苄西林、四环素、甲氨苄啶/磺胺异恶唑耐药率最高,分别为53.12%、43.75%和37.5%。ESBLs(+)株占比18.75%,其多重耐药菌株检出率为83.83%,显著高于ESBLs(-)菌株,差异有统计学意义(P=0.042)。32株DEC共有25个ST型,优势基因型为ST10共4株(12.5%),ST28、ST31和ST3153各2株(各占比6.25%),其他21个型别各1株菌(各占比3.13%)。EAEC中检出1株携带bla_(NDM-1)的耐碳青霉烯类菌株。结论我院感染性腹泻患者的DEC以aggR、pic、astA、eae 4种毒力基因较为常见,以EAEC、EPEC为主要型别,基因型别呈高度多态性,检出多重耐药菌株。

粤北地区鸭源沙门菌分离鉴定及生物学特性研究

为了解粤北地区鸭源沙门菌流行菌株生物学特性,本研究对鉴定的鸭源沙门菌菌株进行了血清型鉴定、毒力基因检测、药敏试验及致病性初步试验。结果显示,病死鸭组织沙门菌分离率为57.14%(16/28);鉴定为鼠伤寒沙门菌占62.5%(10/16),肠炎沙门菌占31.25%(5/16)。分离菌株最少携带了12个毒力基因,最多携带了19个毒力基因。分离菌株对阿莫西林、青霉素、罗红霉素等耐药率超过90%,对多西环素、大观霉素、环丙沙星、阿米卡星等较为敏感,所有菌株呈多重耐药。分离菌株可导致小鼠发病和部分死亡。研究结果表明,沙门菌在粤北地区发病肉鸭中有较高感染率,流行的优势血清型为鼠伤寒沙门菌和肠炎沙门菌,流行菌株普遍存在多重耐药性且具有一定的致病性。

拟杆菌属临床分离株耐药性及脆弱拟杆菌肠毒素bft基因分型特征分析

目的 分析拟杆菌属临床分布及其耐药特征,并探讨脆弱拟杆菌肠毒素bft基因分型特征。方法收集2016年6月—2019年6月内蒙古医科大学附属医院分离自临床样本的拟杆菌属147株,对其进行菌种鉴定和体外药物敏感性试验。采用聚合酶链反应(PCR)检测脆弱拟杆菌肠毒素bft基因。结果 147株拟杆菌属细菌中,脆弱拟杆菌100株(68.0%)、多形拟杆菌25株(17.0%)、卵形拟杆菌9株(6.1%)。体外药物敏感性试验结果显示,拟杆菌属细菌对克林霉素的耐药率最高(78.9%);脆弱拟杆菌对克林霉素的耐药率为85.0%,高于其他拟杆菌(66.0%)。拟杆菌属细菌对亚胺培南和甲硝唑的耐药率分别为13.6%和4.7%,对其他抗菌药物呈不同程度耐药;脆弱拟杆菌对大部分抗菌药物的耐药率均高于其他类型拟杆菌。脆弱拟杆菌bft基因阳性率为38.0%,以bft-1亚型为主要基因型(28.0%),其次为bft-2亚型(10.0%),未检出bft-3亚型。分离自非腹部临床样本的脆弱拟杆菌bft基因阳性率(55.6%)高于分离自腹部临床样本的脆弱拟杆菌(36.3%)。结论 拟杆菌属临床分离株中脆弱拟杆菌分离率最高,其对临床常用抗菌药物均有一定的耐药性。分离自非腹部临床样本的脆弱拟杆菌产肠毒素的菌株流行率更高,应引起临床重视。

温州市食源性小肠结肠炎耶尔森氏菌毒力基因分布及遗传多样性研究

目的了解温州市食源性小肠结肠炎耶尔森氏菌的毒力基因分布及遗传多样性特征。方法对温州市食品及食源性疾病腹泻病人中分离得到的小肠结肠炎耶尔森氏菌进行生物型、血清型和耐药性分析,并基于全基因组测序进行菌株的毒力基因、多位点序列分型(MLST)、核心基因组多位点序列分型(cgMLST)分析。结果71株食源性小肠结肠炎耶尔森氏菌,94.4%(67/71)的分离株为生物1A型,生物血清型以1A/O∶5占比最高(29.6%,21/71)。分离株对14种抗菌药物的敏感率达到95.8%以上。共获得了16种126个毒力基因,其中2株菌株携带pYV质粒和染色体相关毒力基因。分离株以ST3型最常见(31.6%,12/38),cgMLST分析显示除具有相同ST型的菌株外,菌株无明显基因型簇形成。结论温州市食源性小肠结肠炎耶尔森氏菌存在致病性菌株,菌株呈高度遗传多态性,需加强此类菌株的监测,以预防其引起食源性疾病的暴发。

主穹窿蛋白基因在高原地区癫痫儿童中的多态性分析

目的:探讨分析主穹窿蛋白(MVP)基因三个SNP位点rs4788187、rs3815824、rs3815823多态性与高原地区儿童癫痫的相关性。方法:选取2017年1月~2020年1月青海大学附属医院收治的80例癫痫患儿为研究对象,根据耐药性不同分为耐药组与非耐药组各40例,利用PCR-RELP方法检测rs4788187、rs3815824、rs3815823三个SNP位点的多态性分布,并进行统计分析。结果:两组年龄、性别、民族以及癫痫发作类型比较差异无统计学意义(P>0.05)。癫痫患儿rs4788187、rs3815824的等位基因C频率明显低于T,而rs3815823的等位基因T频率明显低于C;两组患儿SNP位点rs4788187、rs3815824、rs3815823的基因型频率和等位基因频率比较差异均无统计学意义(P>0.05)。结论:MVP基因的SNP位点(rs4788187、rs3815824、rs3815823)与癫痫耐药性不相关。

2018—2019年辽宁省副溶血性弧菌临床和食品分离株分子流行病学分析

目的了解辽宁省副溶血性弧菌(Vibrio Parahaemolyticus,VP)的流行特征,为辽宁省食源性疾病的防控和应急事件的处置提供可靠的技术支持。方法对2018—2019年辽宁省301株临床和食品VP分离株进行血清学分型、脉冲场凝胶电泳(PFGE)分型、毒力基因(tlh、tdh、trh)鉴定及耐药试验。结果除了59株VP分离株不能进行血清学分型外,其他分离株共分为17个血清型,血清型以O3群、O1群和O2群为主。血清型O3与O1群分离株相似度高。PFGE主要分成11组优势带型,A-K组,其中A-G组为主要流行致病分子型。临床分离株毒力基因携带情况主要为tlh+、tdh+、trh-,食品分离株毒力基因携带情况主要为tlh+、tdh-、trh-。VP分离株主要对β-内酰胺类(CFZ和AMP)、大环内酯类抗菌药物(ERY)耐药。即食食品分离株耐药率远高于非即食食品分离株。结论由于即食食品的特殊性和来自于国外的特性,这些菌株具有引起食源性疾病暴发流行的高风险,应高度重视。部分食品分离株也具有引起食源性疾病的风险,应给予关注。相关部门除了应加强对国内海产品及其抗菌药物使用管控外,更应加强对进口海产品的监管。

广州地区利奈唑胺耐药结核分枝杆菌耐药基因特征分析

目的分析广州地区利奈唑胺(linezolid,LZD)耐药结核分枝杆菌临床分离株耐药基因的突变特征,为控制广州地区耐利奈唑胺结核病提供一定的研究基础和依据。方法选取2012年1月~2022年1月广州市胸科医院住院患者的60株利奈唑胺耐药结核分枝杆菌的临床分离株,分为耐多药组和广泛耐药组。采用微孔板稀释法对60株菌株进行利奈唑胺最低抑菌浓度(MIC)检测,并对60株菌株的利奈唑胺耐药基因23S rRNA、rplC、rplD进行测序,分析其突变特征。结果60株LZD耐药株对LZD的MIC值分布于2~32μg/ml之间。耐多药组的MIC50和MIC90分别为2μg/ml、32μg/ml,广泛耐药组的MIC50和MIC90分别为8μg/ml、32μg/ml。60株菌株中发现有12株23S rRNA基因发生突变,其中有1株发生两个基因位点突变;有11株rplC基因发生突变,其中有1株发生双位点突变;发现2株rplD基因发生突变。23S rRNA、rplC基因突变发生率在不同性别和年龄患者中差异无统计学意义(P>0.05)。结论广州地区利奈唑胺耐药结核分枝杆菌耐药基因23S rRNA、rplC突变发生率明显高于rplD,在不同性别和不同年龄患者中的发生率无显著差异。