新疆克拉玛依市某猪场不同生长期猪源大肠杆菌耐药性调查

为调查新疆克拉玛依市某规模化猪场不同生长期猪源大肠杆菌对临床常用抗菌药物的耐药情况,从该猪场的550份粪样中分离出大肠杆菌549株,其中,哺乳猪源菌200株、妊娠猪源菌100株、保育猪源菌100株、育肥猪源菌99株和哺乳母猪源菌50株。试验采用微量肉汤稀释法测定不同生长期猪粪源大肠杆菌对9种临床常用抗菌药物的最小抑菌浓度。并通过卡方检验比较不同生长期猪粪源大肠杆菌耐药率的差异。结果发现,不同生长期的猪粪源大肠杆菌均对氨苄西林和阿莫西林/克拉维酸两种抗菌药物高度耐药。其中,保育猪粪源大肠杆菌对安普霉素、恩诺沙星、阿莫西林/克拉维酸和头孢噻呋的耐药率显著高于其他生长期猪粪源大肠杆菌(P0.05),哺乳猪和保育猪粪源大肠杆菌耐药情况严重,对多种抗菌药物高度耐药,以7耐为主;育肥猪、哺乳母猪和妊娠猪粪源大肠杆菌以3耐为主。不同生长期的猪粪源大肠杆菌对临床常用抗菌药物的耐药程度不同。     

我国部分地区鸡、猪源沙门氏菌血清型与耐药性比较

本研究将2012—2017年分离自山东等6省份的194株鸡源和89株猪源沙门氏菌进行了血清型鉴定和9大类13种药物的药敏试验。血清型鉴定结果显示:鸡源沙门氏菌血清型以肠炎(59.69%)、婴儿(13.61%)和鸡白痢(8.90%)为主,猪源沙门氏菌血清型以鼠伤寒(58.43%)、肠炎(12.36%)、印第安纳(11.24%)和德尔卑(7.87%)为主,两者优势血清型差异极显著(P0.01)。最小抑菌浓度(MIC)分布结果显示:沙门氏菌对氨苄西林、头孢噻呋、庆大霉素、大观霉素、四环素、多西环素、氟苯尼考、磺胺甲基异噁唑、复方新诺明、恩诺沙星和氧氟沙星耐药的问题较为突出,对黏菌素E的耐药率相对较低;猪源菌株比鸡源菌株耐药严重,对11种药物的耐药率均高于鸡源菌株。2种来源的分离菌株对阿莫西林/克拉维酸钾、头孢噻呋、庆大霉素、大观霉素、四环素、多西环素、氟苯尼考、恩诺沙星、氧氟沙星的耐药率差异均极显著(P0.01),对黏菌素E的耐药率差异显著(P0.05);猪源分离菌株的耐药问题较欧盟国家突出,与鸡源菌株5耐以上的菌株比例差异极显著(P0.01)。肠炎和鼠伤寒沙门氏菌对阿莫西林/克拉维酸、庆大霉素、大观霉素、四环素、氟苯尼考、磺胺异噁唑和黏菌素E的耐药率差异极显著(P0.01),对氨苄西林耐药率差异显著(P0.05)。不同年份间沙门氏菌分离株对氨苄西林、阿莫西林/克拉维酸、头孢噻呋、庆大霉素、大观霉素、四环素、氟苯尼考、复方新诺明、恩诺沙星和黏菌素E的耐药率差异极显著(P0.01),对多西环素和氧氟沙星耐药率差异显著(P0.05)。结果表明:肠炎、鼠伤寒分别是我国部分地区鸡、猪源沙门氏菌的优势血清型;沙门氏菌耐药性与动物来源、血清型和分离年份相关,且鸡、猪源沙门氏菌耐药情况比欧盟国家严重。     

新疆奎屯地区不同团场牛源大肠杆菌耐药情况比较

为了解新疆奎屯地区不同团场牛源大肠杆菌对临床常用抗菌药物的耐药情况,比较不同养殖场之间牛源大肠杆菌的耐药差异,分别从新疆奎屯地区位于123团、127团和128团的牛场,用肛拭子在直肠采集粪样各30份,并分离大肠杆菌。采用微量肉汤稀释法,对分离出的大肠杆菌进行10种抗菌药物最小抑菌浓度的测定,通过卡方检验对不同养殖场牛源大肠杆菌耐药差异性进行比较分析。结果显示,128团对阿莫西林/克拉维酸（33.3%）和安普霉素（23.3%）耐药率相对较高;127团对氨苄西林（20.0%）、阿莫西林/克拉维酸（16.7%）和氟苯尼考（16.7%）耐药率相对较高;123团场对氨苄西林（13.3%）和安普霉素（13.3%）耐药率相对较高。卡方检验结果显示,127团对氟苯尼考和恩诺沙星的耐药率显著高于123团和128团（P<0.05）;128团对阿莫西林/克拉维酸和安普霉素的耐药率分别显著高于123团和127团（P<0.05）。不同团场多重耐药均以1～2耐为主。结果表明,新疆奎屯地区不同团场牛源大肠杆菌对多数被检药物的耐药率不高,仅对常用药物如阿莫西林/克拉维酸和安普霉素等具有较高耐药率,需在临床治疗细菌疾病时更换或避免使用此类抗菌药物。此外,各养殖场对被检抗菌药物的耐药情况不同,说明养殖场的用药极大程度影响细菌的耐药性。     

新疆不同动物源大肠埃希菌耐药性比较

为了比较新疆不同动物源大肠埃希菌对临床常用抗菌药物的耐药情况,从猪场、羊场和牛场分别分离猪源大肠埃希菌454株、羊源大肠埃希菌638株和牛源大肠埃希菌89株,用微量肉汤法对上述细菌进行临床常用β-内酰胺类、氟喹诺酮类、氨基糖苷类和酰胺醇类抗菌药物最小抑菌浓度测定。结果表面,猪源大肠埃希菌对氨苄西林(67.0%)和阿莫西林/克拉维酸(63.7%)耐药率较高,其他药物耐药率在10.4%~41.2%之间;羊源大肠埃希菌对安普霉素(33.9%)和阿莫西林/克拉维酸(21.2%)耐药率较高,其他药物耐药率在3.1%~15.6%之间;牛源大肠埃希菌对氨苄西林(24.4%)和阿莫西林/克拉维酸(8.9%)耐药率较高,其他药物耐药率在1.1%~6.7%之间。多药耐药结果,猪源大肠埃希菌以2耐~5耐为主,羊源大肠埃希菌以0耐~2耐为主,牛源大肠埃希菌以0耐~1耐为主。新疆猪源大肠埃希菌对临床常用抗菌药物耐药情况最严重,羊源菌次之,牛源菌最轻;猪源大肠埃希菌多药耐药现象严重。     

新疆伊犁某牛场大肠杆菌耐药性调查

为了调查新疆伊犁某牛场饮水、饲料和粪样中分离的大肠杆菌对临床常用抗菌药物的耐药情况。本试验采用微量肉汤稀释法,对饮水源、饲料源、牛粪源样品中分离出的大肠杆菌进行最小抑菌浓度测定。结果显示,25份牛场饮水源样品,大肠杆菌分离率为100.0%（25/25）,对阿莫西林/克拉维酸（12.0%）、氨苄西林（4.0%）、诺氟沙星（4.0%）、恩诺沙星（8.0%）和安普霉素（8.0%）5种抗菌药物耐药;72份牛场饲料源样品,大肠杆菌分离率为65.3%（47/72）,对阿莫西林/克拉维酸（36.2%）、氨苄西林（19.1%）、诺氟沙星（4.3%）和安普霉素（4.3%）4种抗菌药物耐药;80份牛粪源样品,大肠杆菌分离率为100.0%（80/80）,对阿米卡星（12.5%）、氨苄西林（7.5%）、恩诺沙星（7.5%）、庆大霉素（5.0%）、诺氟沙星（2.5%）、环丙沙星（2.5%）、阿莫西林/克拉维酸（1.3%）和头孢噻呋（1.3%）8种抗菌药物耐药,仅对安普霉素敏感。该牛场分离的大肠杆菌对常用抗菌药物耐药情况一般,但中介率较高,须在临床治疗细菌性疾病中避免使用不敏感和中介率高的抗菌药物,养殖场饮水和饲料有被耐药大肠杆菌污染的风险。     

大肠杆菌转座子及整合子携带类型与其多重耐药谱型相关性的研究

为探究猪源大肠杆菌转座子及整合子携带类型与其多重耐药相关性，试验采集贵州省12个规模化养猪场猪肛门拭子，分离鉴定到259株大肠杆菌。采用微量肉汤稀释法测定分离菌株对7类（16种）抗菌药物的最小抑菌浓度（MIC）值，并用PCR方法检测转座子及整合子的携带情况。结果发现，受试菌株对7类（16种）抗菌药物呈现出不同程度的耐药，对大观霉素、庆大霉素、四环素为高度耐药，MIC₉₀的耐药倍数为32倍；对亚胺培南、多黏菌素B表现敏感，MIC₉₀的耐药倍数为2倍，其他药物的耐药倍数介于二者之间。受试菌株多重耐药最高达7重，2重以上耐药率达到100%，多重耐药谱型共计14种，以β-内酰胺类+氨基糖苷类+四环素类+酰胺醇类+磺胺类+氟喹诺酮类、β-内酰胺类+氨基糖苷类+四环素类+酰胺醇类+磺胺类+氟喹诺酮类+多肽类为主，占56.97%。受试菌株转座子和整合子检出率较高，转座子及整合子表型共计20种，其中IScp1+IS26+tnpA+tnp513+intI1和IS26+tnpA+tnp513+intI1的组合类型占比最多，分别为48.48%和26.06%。本试验结果表明，插入元件IScp1与β-内酰胺类药物耐药呈显著或极显著相关（P<0.05，P<0.01），插入元件IS26与氨苄西林耐药相关性极显著（P<0.01），与磺胺异唑耐药相关性显著（P<0.05）；整合子intI1与氨苄西林耐药相关性极显著（P<0.01）；intI2与头孢他啶耐药相关性极显著（P<0.01），与头孢噻呋、恩诺沙星、亚胺培南耐药相关性显著（P<0.05）；IScp1+IS26+tnpA+tnp513+intI1组合类型与复方新诺明耐药相关性极显著（P<0.01）；IS26+tnpA+tnp513+intI1组合类型与复方新诺明耐药相关性显著（P<0.05），与头孢噻呋耐药相关性极显著（P<0.01），但与庆大霉素耐药呈负相关关系（P<0.05）。综上，受试菌株对β-内酰胺类、氟喹诺酮类、磺胺类、氨基糖苷类的多重耐药与转座子及整合子携带类型呈显著或极显著相关。     

新疆某牛场不同日龄犊牛粪源大肠杆菌耐药性分析

观察不同日龄犊牛粪源大肠杆菌耐药性趋势动态变化,筛选新疆一某牛场进行分离粪源大肠杆菌株并分析临床常用抗菌药物耐药性情况,了解养殖场对抗生素治疗安全状况。选择新疆某一集约化程度较高的肉牛养殖场,分别采集0-30日龄、31-60日龄、61-90日龄犊牛的粪样。按照实验室常用大肠菌株分离法分离菌株,细菌药敏试验采用世界卫生组织(WHO)推荐的Kirby-Bauer法对分离出的大肠杆菌进行耐药性测定。200份不同日龄犊牛粪源大肠杆菌分离率为100%;犊牛日龄与耐药率成正比关系,耐药趋势为阿米卡星恩诺沙星诺氟沙星庆大霉素阿莫西林克拉维酸钾阿莫西林环丙沙星氨苄西林。大肠杆菌的耐药率较高,养殖户要正确使用抗菌药,通过药敏试验合理使用抗菌药物,要针对性用药以减少耐药菌株的产生。     

徐州地区猪源致病性大肠杆菌耐药性调查

为了掌握徐州地区猪场大肠杆菌耐药菌株的产生情况和耐药程度,在2016-2018两年间对徐州地区猪源大肠杆菌进行耐药监测。从猪场分离鉴定致病性大肠杆菌,采用纸片扩散法测定46株分离菌株对15种抗菌药物的敏感性。结果表明,猪源大肠杆菌分离株多重耐药现象严重,至少耐药10种以上,其中10耐和11耐的菌株所占比例最大,分别为28.3%和23.9%;分离菌株对阿莫西林、氟苯尼考、强力霉素耐药率最高,达到80%以上,对头孢噻呋耐药率最低,为17.4%。两年间痢菌净耐药率下降幅度最大,丁胺卡那霉素耐药率增长幅度最大。     

新疆某猪场不同生长期的猪源大肠杆菌对抗生素耐药性调查

为了调查新疆某规模化养猪场不同生长期的猪源大肠杆菌对临床常用抗生素的耐药情况,从该猪场分别采集育肥猪、妊娠猪、保育猪和哺乳猪的粪样。试验采用微量肉汤稀释法,对从粪样中分离出的大肠杆菌进行最小抑菌浓度测定。结果显示,育肥猪、妊娠猪、保育猪和哺乳猪源大肠杆菌分离率分别为85.2%、83.0%、81.0%和83.0%。不同生长期的猪源大肠杆菌均对氨苄西林和阿莫西林/克拉维酸2种抗生素高度耐药,对头孢噻呋都较为敏感。哺乳猪和保育猪源大肠杆菌耐药情况严重,对多种抗生素高度耐药,以5耐为主;妊娠猪源大肠杆菌轻度耐药,以0耐为主;育肥猪源大肠杆菌以3耐为主。不同生长期的猪源大肠杆菌对临床常用抗菌药物的耐药程度不同。     

30日龄犊牛粪源大肠杆菌耐药性调查

旨在调查新疆某牛场犊牛粪源大肠杆菌对临床常用抗菌药物的耐药情况。对该牛场92头30日龄犊牛进行直肠粪样采集,采用常规方法进行大肠杆菌的分离鉴定,利用纸片扩散法对分离菌株进行药物敏感性试验。结果表明,从92份犊牛粪便样本中共分离到92株大肠杆菌,分离率为100%;分离菌株对阿米卡星(93.75%)、恩诺沙星(64.50%)、诺氟沙星(59.30%)、庆大霉素(57.14%)4种抗菌药物的耐药性较强,耐药率均超过50%;对阿莫西林/克拉维酸钾(36.30%)、阿莫西林(26.00%)、环丙沙星(13.60%)3种抗菌药物的耐药率也相对较高;对氨苄西林的敏感性最高,耐药率仅为3.03%。综上提示,该牛场犊牛粪源大肠杆菌对不同抗菌药物产生了一定程度的耐药性,建议该牛场应在药物敏感性试验结果的指导下合理选择抗菌药物。     

Comparison of Drug Resistance of E.coli from Pigs in Different Regions under the Same Breeding Mode

In order to compare the resistance of E.coli from pigs of different regions under same breeding mode in Xinjiang, we collected fecal samples from pig farms of Hutubi (207), Manasi (210) and Changji (210) in a certain scale, respectively, a total of 627 samples, and isolation rate of E.coli from fecal samples were all 100.0%.The broth dilution method was used to detect resistance of E.coli to antimicrobials, and we compared the differences of resistance rate of E.coli from pig farms in three regions by chi square test. E.coli from pig farms of Hutubi region to apramycin, amikacin, amoxicillin-clavulanic acid and ceftiofur existed extremely significant differences comparing with another two regions (P< 0.01).E.coli from pig farms of Manasi region to ciprofloxacin existed significant differences comparing with another two regions (P< 0.05), and existed extremely significant differences to norfloxacin (P< 0.01).E.coli from pig farms of Changji region to norfloxacin had extremely significant differences comparing with Hutubi region (P< 0.01), and existed significant differences to amoxicillin-clavulanic acid and ampicillin comparing with Manasi region (P< 0.05).3 to 8 resistant E.coli strains accounted for 94.7% in Hutubi region, 3 to 7 resistant E.coli strains accounted for 89.5% in Manasi region and 4 to 8 resistant E.coli strains accounted for 82.4% in Changji region, respectively.There was no significant difference among different regions on multi-drug resistance (P> 0.05).The results indicated that E.coli from pigs of different regions under the same breeding mode had different characteristics of resistance.In addition, the resistance of E.coli from pigs were very serious and mainly multi-drug resistant, drug resistance patterns were diversified.     

Relationship Between Expression Levels of Porcine m6A Demethylases FTO and ALKBH5 Genes and Resistance to E.coli F18 Infection

To investigate the relationship between the mRNA expression level of m⁶A demethylase and E.coli F18 resistance in piglets,Real-time quantitative PCR was used to detect the mRNA expression differences of m⁶A demethylase FTO and ALKBH5 genes in the duodenum and jejunum tissues of E.coli F18-resistant and -sensitive individuals from 35-day Sutai weaned piglets.In E.coli F18ab,F18ac bacteria-stimulated and endotoxin LPS-induced porcine small intestinal epithelial cells (IPEC-J2),the expression levels of FTO and ALKBH5 genes were detected,respectively.The results showed that the expression levels of FTO and ALKBH5 genes in the duodenum and jejunum of resistant individuals were extremely significantly or significantly higher than those of sensitive individuals (P<0.01,P<0.05),and the expression of FTO gene were not significantly changed in E.coli F18 bacteria-stimulated IPEC-J2 cells (P>0.05),but the expression levels of ALKBH5 gene were significantly up-regulated after F18ac stimulation (P<0.05).After LPS induction for 4 hours,the expression levels of FTO and ALKBH5 genes showed significant up-regulation in IPEC-J2 cells (P<0.05).This study preliminarily verified and indicated that the expression levels of m⁶A demethylases FTO and ALKBH5 genes were closely related to E.coli resistance of piglets at the cellular and individual levels,which will provide a theoretical basis for future in-depth study of the regulation mechanism of RNA demethylation modification on bacterial diarrhea in piglets.     

猪m6A去甲基化酶FTO、ALKBH5基因表达水平与大肠杆菌F18感染抗性的关系分析

为了探讨猪6-甲基腺嘌呤（m⁶A）去甲基化酶mRNA表达水平与仔猪大肠杆菌F18抗性的关系,本研究运用实时荧光定量PCR方法检测m⁶A去甲基化酶FTO和ALKBH5基因在35日龄苏太猪断奶仔猪大肠杆菌F18抗性型和敏感型个体十二指肠和空肠组织中的mRNA表达差异,同时分别利用F18ab、F18ac大肠杆菌菌体刺激和内毒素LPS诱导猪小肠上皮细胞（IPEC-J2）,检测FTO、ALKBH5基因表达变化。结果表明,FTO、ALKBH5基因在抗性型个体十二指肠和空肠中的表达量均极显著或显著高于敏感型个体（P<0.01,P<0.05）;不同F18大肠杆菌菌体刺激IPEC-J2细胞后,FTO基因表达水平均无显著变化（P>0.05）,而ALKBH5基因在F18ac刺激后表达量显著上调（P<0.05）;LPS诱导4 h时,FTO和ALKBH5基因表达量均显著上调（P<0.05）。本研究在细胞和个体水平上初步验证并发现了m⁶A去甲基化酶FTO和ALKBH5基因表达水平与大肠杆菌感染仔猪密切相关,为今后深入研究RNA去甲基化修饰在仔猪细菌性腹泻调控中的作用机制提供了理论基础。     

石榴皮水提物对猪源多重耐药大肠杆菌的抑菌作用研究

【目的】筛选腹泻仔猪中携毒力基因且多重耐药的大肠杆菌菌株,评估中药水提物对该菌株的抑菌活性,并探索中药抑菌机制。【方法】通过PCR方法与Kirby-Bauer(K-B)药敏纸片法评估临床大肠杆菌菌株的肠毒素基因携带情况和耐药性;通过微量肉汤稀释法评估中药水提物对多重耐药大肠杆菌菌株的抑菌活性,确定最小抑菌浓度(minimal inhibitory concentration, MIC)和最小杀菌浓度(minimal bactericidal concentration, MBC);通过电导率仪以及相关试剂盒检测评估石榴皮水提物对多重耐药菌株作用不同时间后菌液电导率、碱性磷酸酶(alkaline phosphatase, AKP)含量及菌体内ATP含量的变化;通过SDS-PAGE和蛋白含量检测评估菌体内可溶性蛋白的含量变化;通过扫描电镜观察大肠杆菌形态变化。【结果】PCR检测14株临床大肠杆菌中肠毒素基因携带率达到78.57%,抗生素药敏试验结果显示,所有菌株至少对2种抗生素耐药,均属于多重耐药菌株。中药药敏试验结果显示,黄连、黄芩、石榴皮水提物对多重耐药菌株抑菌活性较好,其中石榴皮水提...     

猪m6A甲基化酶WTAP基因与F18大肠杆菌感染的关系

本研究旨在探讨猪m⁶A甲基化酶WTAP表达水平与大肠杆菌（E. coli）感染抗性的关系。选取35日龄苏太断奶仔猪（Sus scrofa）大肠杆菌抗性型和敏感型个体各4头,采集十二指肠和空肠组织,利用RT-qPCR检测WTAP在E. coli抗性型和敏感型个体十二指肠、空肠的表达差异,并分别利用产肠毒素大肠杆菌（F18ab、F18ac）刺激和内毒素（LPS）诱导猪小肠上皮细胞（IPEC-J2）,检测WTAP基因的表达变化。同时构建WTAP基因干扰载体并转染IPEC-J2细胞,通过菌毛定量、菌落计数以及间接免疫荧光试验检测该基因沉默对大肠杆菌黏附能力的影响。结果显示：在十二指肠和空肠组织中,WTAP基因在E. coli抗性型个体中的表达量显著高于敏感型个体（P<0.01）;并且在F18ab和F18ac刺激后表达量显著下降,与LPS诱导6 h后结果相一致（P<0.01）。沉默WTAP基因后,大肠杆菌黏附能力极显著上升（P<0.01）。本研究在细胞和个体水平上验证发现,m⁶A甲基转移酶WTAP的高表达可能有助于仔猪抗大肠杆菌感染,为进一步揭示仔猪抗大肠杆菌感染的RNA甲基化调控机制奠定基础。     

Resistance Analysis and Genotype Detection of ESBLs-producing Swine E.coli Strains in Parts of Guizhou Province

To find out the drug resistance and the distribution and prevalence of ESBLs genotypes of E.coli strains on scale pig farms in Guizhou, the antimicrobial susceptibility of 164 E.coli strains were determined.And the genotype of ESBLs was identified and determined by CLSI standard method, DNA was amplified and the 4 genotypes of drug resistance plasmid of ESBLs were analyzed by PCR.The results showed that the drug resistance rates of 164 E.coli strains to ten kinds of antimicrobial agents were ceftiofur 93.29%, ampicillin 87.19%, tetracycline 86.59%, gentamicin 81.10%, streptomycin 53.66%, polymyxin 51.83%, ciprofloxacin 53.05%, kanamycin 47.56%, chlortetracycline 34.76% and florfenicol 21.95%, respectively, and most of them were multiple drug resistance strains, during which there were 137 strains ESBLs positive, detection rate was 83.54%, and the detection rates of 4 regions were different.The detection rates of TEM, CTX-M-1, SHV and OXA-1 genes of 137 ESBLs-producing E.coli strains were 90.51%, 70.07%, 51.82% and 43.07%, respectively.And the detection rates were various in 4 regions.The results showed that the drug resistance of E.coli strains from scale pig farms was serious in Guizhou, and the detection rate of ESBLs strains was very high, the drug resistance gene detection rate was extremely high, and most drug resistance strains were the composite genotype drug resistant.To prevent and cure colibacillosis in veterinary clinic effectively, we should reinforce monitoring and studying about the ESBLs-producing drug resistance bacteria.     

贵州部分地区猪源大肠杆菌耐药性分析及ESBLs基因型检测

为了解贵州地区规模养猪场大肠杆菌菌株耐药情况和ESBLs基因型的流行情况,试验采用CLSI推荐的方法对采自贵州省4个地区规模养猪场的164株大肠杆菌进行药物敏感性试验和产ESBLs菌的检测,并用PCR方法对TEM、SHV、OXA-1和CTX-M-1 4种常见ESBLs基因进行检测。结果显示,164株大肠杆菌对10种常用抗菌药物耐药率分别是头孢噻呋93.29%、氨苄西林87.19%、四环素86.59%、庆大霉素81.10%、链霉素53.66%、多黏菌素51.83%、环丙沙星53.05%、卡那霉素47.56%、金霉素34.76%和氟苯尼考21.95%,且大多为多重耐药,其中检测出ESBLs阳性菌株137株,阳性率为83.54%,各个地区的检出率不同;137株产ESBLs大肠杆菌中,TEM、SHV、OXA-1和CTX-M-1基因的检出率分别为90.51%、70.07%、51.82%和43.07%,且多为复合基因型耐药菌株,各地区的各种基因检出率不同。试验结果表明,贵州部分地区的猪源大肠杆菌耐药现象严重,ESBLs菌株的检出率很高,耐药基因的检出率也极高,且多为复合基因型耐药菌株,应加强当地产酶耐药菌的监测和研究,有效防制此类细菌引发的疾病。     

六十二株水貂源大肠杆菌分离株耐药表型与耐药基因及克隆菌群分析

为了调查诸城地区某水貂养殖场粪便源大肠杆菌的表观及其分子特征,采集某个水貂养殖场的水貂粪便进行大肠杆菌分离鉴定,对分离鉴定的大肠杆菌进行血清型鉴定和对14种常见抗菌药物的耐药表型鉴定;使用PCR检测耐药基因以及Ⅰ整合子基因盒的携带情况,利用多位点序列分型（MLST）来分析菌株的克隆关系并构建系统发育树来分析相同克隆群菌株的遗传相似性。结果显示,自82份水貂粪便样品分离到62株大肠杆菌,分离率75.61%;大肠杆菌分离株对AMP和TET的耐药率超过90%,多重耐药菌株（MDR）占比为85.48%。PCR检测到5类耐药基因的存在,qnrS检出率最高,为61.29%（38/62）;aaC2、aaC4、sul1和aac（6'）-Ib-cr耐药基因与菌株产生相应的耐药抗性存在一致性（P<0.01）。分离菌株中Ⅰ类整合子可变区域的优势结构为dfrA27-aadA2-qnrA。鉴定出致病性血清型的存在,且对应菌株都具有多重耐药性,优势血清型为O104：H4。分离株中存在33个STs,ST46为优势STs（16.13%）,具有3个主要克隆群,依次为CC10、CC46和CC176;与致病性相关菌株的STs和人源大肠杆菌具有共同的遗传背景。本研究表明,养殖场的水貂受到致病性和多重耐药性大肠杆菌的污染,相同克隆群菌株的耐药基因分布具有多态性,表观特征差异明显。     

板蓝根微粉水提物抗大肠杆菌活性及其机制的探究

为了探究板蓝根微粉水提物的抗菌活性及其抗菌机制，试验通过扫描电镜、透射电镜检测板蓝根微粉水提物对大肠杆菌形态和结构影响；酶标仪测定板蓝根微粉对大肠杆菌电导率、胞内物质总漏出率影响；测定大肠杆菌培养液中碱性磷酸酶含量以及大肠杆菌菌体内、外蛋白质含量；通过DAPI染色DNA、RNA，检测板蓝根微粉水提物对大肠杆菌核酸的影响；检测板蓝根微粉水提物对大肠杆菌菌体内、外谷丙转氨酶（ALT）、谷草转氨酶（AST）、丙酮酸以及三磷酸腺苷（ATP）等菌体内代谢的影响。结果显示，板蓝根微粉水提物作用大肠杆菌10 h后，扫描电镜检测可见菌体出现溢缩，菌体长度明显变短，断裂形成许多残体，有的中间凹陷，发生变形；透射电镜下可观察大肠杆菌的胞壁界限模糊不清，壁膜呈现锯齿状，弯弯曲曲，变形，有的菌体破碎。总漏出率、电导率以及碱性磷酸酶含量测定结果显示，板蓝根微粉水提物各组D_{600 nm}均高于空白对照组，且呈剂量依赖性。菌体外蛋白质含量从8 h开始与空白对照组差异极显著（P<0.01）；菌体内蛋白质含量从4 h开始与空白对照组差异极显著（P<0.01）。DNA含量在12 h前与空白对照组无显著差异（P>0.05），从16 h开始与空白对照组差异极显著（P<0.01）；RNA含量在8 h开始降低，在12 h时与空白对照组差异显著（P<0.05），从16 h开始差异极显著（P<0.01）。ALT和AST浓度测定无显著性差异（P>0.05）。培养液和菌体内的丙酮酸含量均高于空白对照组，且从4 h开始与空白对照组差异极显著（P<0.01）。培养液中的ATP含量与空白对照组差异极显著（P<0.01）；菌体内ATP含量从4 h开始与空白对照组差异极显著（P<0.01）。综上，板蓝根微粉水提物可以通过破坏细胞壁、细胞膜的完整性，抑制细菌遗传物质合成和代谢，影响丙酮酸和ATP含量从而实现抗大肠杆菌作用。     

Study on the Anti-Escherichia coli Activity and Mechanism of Water Extract of Radix isatidis Powder

In order to explore the antibacterial activity and mechanism of the Radix isatidis powder water extract,the effects of the Radix isatidis powder water extract on the morphology and structure of Escherichia coli (E.coli) were tested by scanning electron microscopy and transmission electron microscopy.The effects of the Radix isatidis powder water extract on the conductivity and total leakage rate of E.coli and the content of alkaline phosphatase in the culture medium of the content of protein,DNA and RNA were stained by DAPI to detect the effect of the Radix isatidis powder water extract on the nucleic acid of E.coli,and the effect of the Radix isatidis powder water extract on the metabolism of E.coli in vivo,in vitro,ALT,AST,pyruvic acid and ATP.The results showed that after the Radix isatidis powder water extract acted on E.coli for 10 h,it was observed by SEM that the bacteria appeared to overflow and shrink,the length of the bacteria became shorter obviously,many residues were formed due to breakage,some of them were sunken in the middle and deformed.Under TEM,it was observed that the boundary of the cell wall of E.coli was unclear,the wall membrane was zigzag,deformed and some of the bacteria were broken.The protein content outside the cell was significantly different from that in the blank control group from 8 h (P<0.01), and that in the cell from 4 h (P<0.01). DNA content had no significant difference with blank control group before 12 h (P>0.05), but had significant difference with blank control group from 16 h (P<0.01); RNA content began to decrease at 8 h, and was significantly different from blank control group at 12 h (P<0.05), and was extremely significant from 16 h (P<0.01). There was no significant difference between ALT and AST (P>0.05). The pyruvate content in culture medium and bacteria was higher than that in blank control group, and the difference was very significant from 4 h (P<0.01). The ATP content in the culture medium was significantly different from that in the blank control group (P<0.01), and the ATP content in the cell was significantly different from that in the blank control group from 4 h (P<0.01).In conclusion,the Radix isatidis powder water extract could inhibit the synthesis and metabolism of bacterial genetic material and the content of pyruvate and ATP by destroying the integrity of cell wall and cell membrane.