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1.
氨基糖苷类药物曾经是临床最为常用而有效的抗生素,长期应用与不合理使用使得该药物效果不尽理想。本研究参照GenBank中耐药基因相关序列,设计引物,结果从禽源大肠杆菌基因组中扩增出4种抗氨基糖苷类药物基因aadA1、strA、strB、aph(3′),经pGEX-T-easy和pET32a载体克隆和序列分析,证明扩增获得的基因序列与GenBank中参考序列同源性达到97%以上。对分离保存的216株禽致病性大肠杆菌进行氨基糖苷类药物耐药基因的分子流行病学检测,结果表明:aadA1阳性率高达49.1%,strA和strB的阳性率分别为56%和65.7%,aph(3′)的阳性率为16.2%;近三分之二的被检菌株携带有2种以上氨基糖苷类药物耐药基因。药敏试验结果显示所有被检菌株对链霉素耐药率为68.9%,而对阿米卡星的耐药率为38.9%。本研究结果说明禽类细菌的耐药性与相关耐药基因的检出率基本呈正相关,临床日益严重的耐药现象与耐药基因的普遍存在有着很大的关系,提示控制禽源耐药细菌对人类健康与卫生安全有重要意义。  相似文献   

2.
为了查清新疆石河子部分规模场引起羔羊死亡的原因,本研究对采集的病羊肺脏组织进行细菌分离,同时对分离菌株进行小鼠致病性试验、特异性基因PCR检测、药物敏感性试验和耐药基因PCR检测.结果显示:从病变肺脏组织分离得到10株细菌,分离菌为大肠杆菌,致病性强;分离菌株呈现多重耐药现象,对诺氟沙星、复方新诺明、氟苯尼考等17种抗生素耐药;检测到iutA、fyuA、ireA、hlyD和afa 5种毒力因子,strA、strB、aadA1/aadA2、Bla(TEM1)、Bla(OXA1)、Tet A和Tet B 7种耐药基因.本研究为该地区规模场绵羊感染大肠杆菌的防控和临床用药提供依据.  相似文献   

3.
Rectal smears of calves, cows and young bulls, as well as cloacal smears of house sparrows (Passer domesticus), from farms at the villages of Sumice and Troskotovice, Czech Republic, were examined for E. coli resistant to 12 antimicrobials. The resistant isolates were tested for antimicrobial-resistance genes and integrons. Totals of 40% (n=183), 3% (n=95), 0% (n=33), and 9% (n=54) of Escherichia coli isolates from calves, cows, young bulls and house sparrows, respectively, were antimicrobial resistant. The following genes were identified in cattle E. coli isolates: tetA, tetB (isolates resistant to tetracycline), bla(TEM) (beta-lactams), strA, aadA (streptomycin), sul1, sul2 (sulphonamides), and cat, floR (chloramphenicol). Seven of 16 antimicrobial-resistant calf isolates from the Sumice farm possessed class 1 integrons with the aadA1 gene cassette integrated, 1 kb in size. On the Troskotovice farm, eight of 57 antimicrobial-resistant calf isolates possessed class 1 integrons. Integrons of 1.5kb with the dhfr1- aadA1 gene cassette were found in four isolates, followed by a 1kb integron with the aadA1 gene found in three isolates, and a 1.7kb integron with the dhfr17-aadA5 gene cassette and the phenotype ASSuTSxtNaCipCCfG. The prevalence of resistant E. coli in calves compared to adult cattle was much higher and probably was influenced by oral antimicrobial usage in calves, feeding with milk and colostrum from treated cows, as well as mechanisms unrelated to antimicrobial drug selection. Although house sparrows lived together with the cattle and came into contact with cattle waste on the farm, they were not infected by resistant E. coli isolates with the same characteristics as those found in cattle.  相似文献   

4.
调查来源于16种四川地区野生动物粪样的24株革兰氏阴性肠杆菌中氨基糖苷类抗生素四种修饰酶基因(am inoglycosides-mod ifying en-zym es,AMEs)的分布情况。应用K-B法进行常规药敏试验,采用多重PCR技术检测分离菌中四种AMEs(aphA3、aacC4、aadA、aacC2)基因并进行序列分析。药敏试验显示分离菌对链霉素的耐药率最高(8/24),对新霉素的耐药率最低(1/24%);多重PCR检测结果显示,aphA3、aa-dA、aacC2基因的检出率分别为:7/24、3/24和2/24,而所有菌株中均未检出aacC4基因;序列分析表明扩增产物与Genbank中的相应序列有很高的同源性(≥97.8%)。  相似文献   

5.
Pulsed field gel electrophoresis (PFGE) patterns, susceptibility to 26 antimicrobial agents used in veterinary and human medicine, and prevalence of antimicrobial resistance genes of Escherichia coli isolated from cows with mastitis were evaluated. Among 135 E. coli isolates, PFGE analysis revealed 85 different genetic patterns. All E. coli were resistant to two or more antimicrobials in different combinations. Most E. coli were resistant to antimicrobials used in veterinary medicine including ampicillin (98.4%, >or=32 microg/ml) and many E. coli were resistant to streptomycin (40.3%, >or=64 microg/ml), sulfisoxazole (34.1%, >or=512 microg/ml), and tetracycline (24.8%, >or=16 microg/ml). Most E. coli were resistant to antimicrobials used in human medicine including aztreonam (97.7%, >or=32 microg/ml) and cefaclor (89.9%, >or=32 microg/ml). Some E. coli were resistant to nitrofurantoin (38%, >or=128 microg/ml), cefuroxime (22.5%, >or=32 microg/ml), fosfomycin (17.8%, >or=256 microg/ml). All E. coli were susceptible to ciprofloxacin and cinoxacin. Almost 97% (123 of 127) of ampicillin-resistant isolates carried ampC. Eleven of 52 (21.2%) streptomycin-resistant isolates carried strA, strB and aadA together and 29 streptomycin-resistant isolates (55.8%) carried aadA alone. Among 44 sulfisoxazole-resistant E. coli, 1 isolate (2.3%) carried both sulI and sulII, 12 (27.3%) carried sulI and 10 (22.7%) isolates carried sulII. Among 32 tetracycline-resistant isolates, 14 (43.8%) carried both tetA and tetC and 14 (43.8%) carried tetC. Results of this study demonstrated that E. coli from cows with mastitis were genotypically different, multidrug resistant and carried multiple resistance genes. These bacteria can be a reservoir for antimicrobial resistance genes and can play a role in the dissemination of antimicrobial resistance genes to other pathogenic and commensal bacteria in the dairy farm environment.  相似文献   

6.
Strangles is a contagious equine disease caused by Streptococcus equi subsp. equi. In this study, clinical strains of S. equi (n=24) and Streptococcus equi subsp. zooepidemicus (n=24) were genetically characterized by sequencing of the 16S rRNA and sodA genes in order to devise a real-time PCR system that can detect S. equi and S. zooepidemicus and distinguish between them. Sequencing demonstrated that all S. equi strains had the same 16S rRNA sequence, whereas S. zooepidemicus strains could be divided into subgroups. One of these (n=12 strains) had 16S rRNA sequences almost identical with the S. equi strains. Interestingly, four of the strains biochemically identified as S. zooepidemicus were found by sequencing of the 16S rRNA gene to have a sequence homologous with Streptococcus equi subsp. ruminatorum. However, they did not have the colony appearance or the biochemical characteristics of the type strain of S. ruminatorum. Classification of S. ruminatorum may thus not be determined solely by 16S rRNA sequencing. Sequencing of the sodA gene demonstrated that all S. equi strains had an identical sequence. For the S. zooepidemicus strains minor differences were found between the sodA sequences. The developed real-time PCR, based on the sodA and seeI genes was compared with conventional culturing on 103 cultured samples from horses with suspected strangles or other upper respiratory disease. The real-time PCR system was found to be more sensitive than conventional cultivation as two additional field isolates of S. equi and four of S. zooepidemicus were detected.  相似文献   

7.
对河南省鸡源大肠杆菌Ⅰ类、Ⅱ类和Ⅲ类整合子及其携带的耐药基因盒进行了分子流行病学研究。结果表明：51株鸡源大肠杆菌中86．3％（44／51）检出Ⅰ类整合子,3．9％（2／51）检出Ⅱ类整合子,未检出Ⅲ类整合子。Ⅰ类整合子共检出5种类型的基因盒,分剐是sat基因盒（100％）,dfrAl＋aadAl基因盒（45．4％,20／44）,dfrAl7＋aadA5基因盒（22．5％,9／44）,dfrAl＋sat＋aadA2基因盒（6．8％,3／44）和4800bp未知基因盒（27．3％,12／44）,其中4800bp未知基因盒的下游携带有ESBLCTX-M基因,Ⅱ类整合子的基因盒携带dfrAl＋sat＋aadAl3种基因。  相似文献   

8.
To assess the prevalence of antimicrobial resistance and class I integrons in Escherichia coli strains (n=58) isolated from bovine mastitis in Inner Mongolia, antimicrobial susceptibility and the presence of various types of integrons were characterized. Most isolates were susceptible to amikacin, colistin, ceftazidime, gentamicin and kanamycin, while those also exhibited high resistant incidence rates to ampicillin, amoxicillin, sulfadiazine and sulfamethoxydiazine. The integrase gene of integrons was amplified by PCR using degenerate primers. The integrons were confirmed by restriction fragment length polymorphism (RFLP) analysis of positive PCR products. Neither class II nor class III integron was detected, while 56.90% (n=33) of the isolates were positive for the presence of intI1 gene. Sequencing analysis of gene cassettes revealed that seven gene cassettes were found, which encoded resistance to trimethoprim (dfrA1 and dfrA17), aminoglycosides (aacA4, aadA1 and aadA5) and chloramphenicol (catB3), respectively. Of them, the gene cassette array dfrA17-aadA5 was found most prevalent (62.96%). The percentage of positive-integron among the isolates whose resistant profile was relatively broad (n> or =7) is 100.00%, while the one in narrow-profile isolates (n=2-6) is 30.56%. The correlation analysis revealed the incidence of integrons among the isolates were highly related to the resistant profile, indicating integrons play an important role in the dissemination and spread of the antimicrobial resistant strains.  相似文献   

9.
By PCR using the ant(3")-Ia primer pair the aadA gene was detected in 34 streptomycin- and spectinomycin-resistant Salmonella enterica serotype Typhimurium strains. Out of them 12 belonged to DT104 and 22 to non-DT104 phage type. Using different primer combinations it was demonstrated that this gene was integron-associated in all cases: in the DT104 strains it was generally contained by a 1 kb integron while in the majority of the non-DT104 strains by a 2.05 kb (less often by a 1.9 or 1 kb) integron. In the case of integrons carrying multiple cassettes the cassette containing the aadA gene was located closer to the 3' end of the integron. The aadA genes of DT104 and non-DT104 strains were different: in the former group the aadA2 gene, while in the latter group (constituted by strains of five different phages types as well as unclassifiable and untypable strains) the aadA1 gene could be identified. The RH50/RH51 primer pair described by Collis and Hall (1992) proved to be suitable for rapid discrimination between the aadA1 and aadA2 genes on the basis that the RH51 primer bound exclusively to the aadA2 gene.  相似文献   

10.
查明辽宁地区整合子在猪源大肠埃希菌中的分布及整合子携带耐药基因盒的种类,可为该病的综合防控提供科学依据。本研究利用整合酶基因PCR扩增法检测整合子,并对整合子可变区进行扩增测序。结果表明,71.43%(40/56)的菌株为Ⅰ类整合子阳性,1.79%(1/56)的菌株同时为Ⅰ类和Ⅱ类整合子阳性,未检测到Ⅲ类整合子;87.8%(36/41)的菌株表现为Ⅰ类整合子可变区扩增阳性,扩增产物大小在116bp~2 307bp之间,100%(1/1)菌株表现为Ⅱ类整合子可变区扩增阳性,大小为2 106bp;整合子可变区含有编码对氨基糖苷类抗生素耐药的基因(aadA1、aadA2、aadA5、aadA22、aadB、aacA4和sat2),编码对磺胺类抗生素耐药的基因(dfrA1、dfrA12、dfrA17),编码对氯霉素抗生素耐药的基因(cmlA1、cmlA6)。因此,整合子在大肠埃希菌中广泛存在,辽宁地区大肠埃希菌中整合子主要携带编码对氨基糖苷类、磺胺类和氯霉素耐药基因盒。  相似文献   

11.
The purpose of this study was to characterize, at the molecular level, the Ehrlichia canis strains involved in naturally occurring canine monocytic ehrlichiosis (CME) in Greece, and to investigate if any sequence diversity exists between the 16S rRNA genes of those involved in the mild non-myelosuppressive or the severe myelosuppressive form of CME. To this end, amplification of the ehrlichial 16S rRNA gene was attempted by nested polymerase chain reaction (PCR) assays in bone marrow (BM) aspirates from 20 dogs tentatively diagnosed as having non-myelosuppressive (n=10, group A) or myelosuppressive (n=10, group B) CME. PCR assay using E. canis-specific primers revealed that 15 BM samples, including all group A and 5 group B dogs, were positive. Using universal PCR primers, a nearly full-length 16S rRNA gene could be amplified from 13 BM samples, including 9 group A and 4 group B dogs. The 16S rDNA analysis based on secondary structure revealed that all sequences of the Greek strains were identical to each other and indicated 100% identity among some American (Venezuelan and Brazilian), European (Greek), Middle Eastern (Turkish) and Asiatic (Thailand) strains. The results of this study suggest that the E. canis strains involved in the non-myelosuppressive and myelosuppressive forms of CME in Greece share an identical 16S rRNA genotype.  相似文献   

12.
Faecal samples of healthy dogs (n=39) and cats (n=36) obtained in Northern Portugal were seeded on Levine agar plates, and two Escherichia coli isolates per sample were recovered (78 of dogs and 66 of cats). The susceptibility to 16 antimicrobial agents was tested in this series of 144 E. coli isolates. Almost 20% of them showed tetracycline resistance and 12 and 15% presented ampicillin or streptomycin resistance, respectively. The percentage of resistance to the other antimicrobial agents was in all cases below 4%, and no resistant isolates were detected for ceftazidime, imipenem, cefoxitin or amikacin. Two isolates (from one dog) showed cefotaxime-resistance and harboured both the CTX-M-1 and OXA-30 beta-lactamases. A bla(TEM) gene was detected in 12 of 17 ampicillin-resistant isolates, the aac(3)-II gene in the three gentamicin-resistant isolates, aadA in 7 of 22 streptomycin-resistant isolates, and tet(A) and/or tet(B) gene in all 28 tetracycline-resistant isolates. The gene encoding class 1 integrase was detected in six E. coli isolates, including the four trimethoprim-sulfamethoxazole-resistant isolates and those two harbouring CTX-M-1 and OXA-30 beta-lactamases; different gene cassette arrangements were identified: dfrA1+aadA1 (two isolates), dfrA12+orfF+aadA2 (two isolates) and bla(OXA30)+aadA1 (two isolates). One amino acid change in GyrA protein (Ser83Leu or Asp87Tyr) was detected in four nalidixic acid-resistant and ciprofloxacin-susceptible isolates and two amino acid changes in GyrA (Ser83Leu+Asp87Asn) and one in ParC (Ser80Ile) were identified in one nalidixic acid- and ciprofloxacin-resistant isolate. Faecal E. coli isolates of healthy pets could be a reservoir of antimicrobial resistance genes.  相似文献   

13.
新城疫流行株GX-08全基因序列分析及其对鸭的致病性研究   总被引：1，自引：1，他引：0  
2008年从患病免疫鸡群中分离到1株新城疫病毒（Newcastle disease virus,NDV）流行株,经致病指数测定为强毒株,编号为GX-08。致病性试验表明,GX-08株对鸭具有感染性和致病性。参照GenBank发表的NDV基因序列设计10对特异性引物,采用RT-PCR方法对GX-08株全基因组序列分段进行扩增。拼接后序列分析表明GX-08株的全基因组序列总长为15192 nt,包含6个开放阅读框,分别编码6种蛋白质NP、P、M、F、HN和L。F基因裂解位点处的氨基酸序列为112R-R-Q-K-R-F117,符合强毒株氨基酸序列特点。F基因第47—420位片段基因分型分析结果表明,该流行株属于基因Ⅶ型。  相似文献   

14.
The aim of the present study was to contribute to the knowledge on extended-spectrum beta-lactamases (ESBL's), AmpC beta-lactamases and integrons in Enterobacteriaceae isolated from horses, which is still limited. The susceptibility of 1581 clinical isolates from animals to ceftiofur was tested. Most of these isolates (n=1347) originated from horses. Seven ceftiofur-resistant equine isolates (four Escherichia coli and three Klebsiella pneumoniae) were identified and all seven were multidrug-resistant. These isolates were further studied for the presence of ESBL's, AmpC beta-lactamases and class 1 integrons. The potential for the horizontal transfer of resistance genes among these clinical isolates was also studied. ESBL-type resistance genes were found in five isolates, AmpC-type genes in one isolates and integrons in six isolates. Nucleotide sequence analysis revealed that the isolates carried the bla(CTX-M-1), bla(CMY-2), bla(TEM-1) and/or bla(SHV-1) genes. This is the first report describing the in vitro conjugal transfer of the bla(CTX-M-1) genes from a clinical E. coli isolate to Salmonella isolates. Gene cassettes encoding resistance to aminoglycosides (aadA1, aadA2 and aadA5), and trimethoprim (dfrA1, drfA12 and dfrA17) were found on the integrons present in the isolates. The cassette arrays of the dfrA17-aadA5 and dfrA1-aadA1 genes in the two integrons of a single E. coli isolate have not yet been described before. To our knowledge this is the first report on ESBL's and AmpC beta-lactamases in equine E. coli and Klebsiella isolates.  相似文献   

15.
Zheng F  Lin G  Zhou J  Cao X  Gong X  Wang G  Qiu C 《Veterinary microbiology》2012,156(3-4):434-438
  相似文献   

16.
北京部分地区猪瘟病毒流行株E2基因序列分析   总被引：1，自引：0，他引：1  
为了解北京地区猪瘟病毒(CSFV)流行毒株的遗传变异情况,采用套式RT-PCR方法,对从北京部分地区近期分离的7株CSFV流行毒株的E2基因主要抗原编码区进行了扩增、克隆与序列测定,并应用DNA Star分析软件对所测定的7株毒株与国内外参考毒株的相应序列进行了同源性分析及氨基酸序列比对。结果表明,7株CSFV流行毒株之间核苷酸与氨基酸的同源性分别为96.3%～99.3%和95.6%～100%,与CSFV石门毒株(Shimen株)的核苷酸与氨基酸的同源性分别为81.7%～83.5%和82.4%～84.6%,与CSFV兔化弱毒株(HCLV株)的核苷酸与氨基酸的同源性分别为80.6%～81.7%和78.0%～80.2%,7株CSFV流行毒株均属于基因2群,且705、713、729和734位氨基酸发生置换。本研究初步揭示了北京部分地区目前流行的CSFV毒株与Shimen株和HCLV株在E2基因主要抗原区上存在较大差异。  相似文献   

17.
为了解马立克氏病病毒(MDV)强、弱毒株主要致肿瘤相关基因变异情况,本研究根据MDV GA株基因组序列,设计合成扩增基因组重复区的引物,得到MDV814疫苗株病毒基因组中约26kb的序列片段。与GenBank登录的强、弱毒株进行比较分析表明,扩增的MDV1型814疫苗株的长重复区为12774bp,预测的开放阅读框(ORF)有48个;短重复区为11426bp,预测的ORF有38个。发现了4个MDV814株特有的ORF。814株在编码Meq、RLORF6和23ku的重叠基因内具有类似于疫苗株CVI988的177bp的插入;在RLORF12基因编码区内存在69bp的缺失,该缺失位于病毒复制起始位点内。同时,发现7个814疫苗株特有的氨基酸突变,分布在6个ORF内。单核苷酸多态性(SNP)的鉴定发现,多个基因具有单核苷酸的突变,主要分布于Meq基因,其中氨基酸A115V(丙氨酸-缬氨酸),N142D(天冬酰胺-天门冬氨酸)的变异是814疫苗株所特有的。MDV814疫苗株重复区的基因序列的比较分析将有助于MDV致肿瘤机制的研究。  相似文献   

18.
Multi-drug-resistant (MDR) Salmonella enterica serovar Newport strains are increasingly isolated from animals and food products of animal origin and have caused septicemic illness in animals and humans. The purpose of this study was to determine the occurrence and the epidemiologic, phenotypic, and genotypic characteristics of S. Newport of animal origin that may infect humans, either via the food chain or directly. During the 1993-2002 period, the Office International des Epizooties Reference Laboratory for Salmonellosis in Guelph, Ontario, received 36 841 Salmonella strains for serotyping that had been isolated from animals, environmental sources, and food of animal origin in Canada. Of these, 119 (0.3%) were S. Newport. Before 2000, none of 49 S. Newport strains was resistant to more than 3 antimicrobials. In contrast, between January 2000 and December 2002, 35 of 70 isolates, primarily of bovine origin, were resistant to at least 11 antimicrobials, including the extended-spectrum cephalosporins. The blaCMY-2', flo(st'), strA, strB, sulII, and tetA resistance genes were located on plasmids of 80 to 90 MDa that were self-transmissible in 25% of the strains. Conserved segments of the integron 1 gene were found on the large MDR-encoding plasmids in 3 of 35 strains additionally resistant to gentamicin and spectinomycin or to spectinomycin, sulfamethoxazole-trimethoprim, and trimethoprim. Resistance to kanamycin and neomycin was encoded by the aphA-1 gene, located on small plasmids (2.3 to 6 MDa). The increase in bovine-associated MDR S. Newport infections is cause for concern since it indicates an increased risk of human acquisition of the infection via the food chain.  相似文献   

19.
根据已发表的５２／７０株ＩＢＤＶ基因组序列,设计并合成了一对特异扩增ＩＢＤＶ ＶＰ２基因的引物。以陕西地区分离的ＩＢＤＶ野毒ＸＮ株,ＨＺ株为材料,以其基因组为模板利用ＲＴ－ＰＣＲ技术扩增出了１．５ｋｂ的ｃＤＮＡ产物,将ＶＰ２基因克隆于ＰＵＣ１１９质粒上,得到重组ＰＵＣ１１９质粒。  相似文献   

20.
Fowl typhoid caused by Salmonella enterica subsp. enterica serotype Gallinarum biotype Gallinarum is the most important chicken disease in Korea. Due to appearance of new or multiple antibiotics resistances in the recently isolated strains, it was difficult to control the disease using antibiotics in our country. Therefore, the prevalence and genetic contents of class 1 integrons in biotype Gallinarum isolated between 1992 and 2001 were investigated by PCR and direct sequencing, respectively. Out of 90 strains, 35 (39%) carried class 1 integrons. The 1.0, 1.6 and 2.0kbp amplicons were amplified in 32 strains (36%), 2 strains (2%) and 1 strain (1%), respectively. The 1.0, 1.6 and 2.0 kbp amplicons contained one (aadA1a), two (aadB-aadA1b) and three cassettes (dhfrXII-orfF-aadA2), respectively, providing resistances against aminoglycosides (aadA1a, aadA1b, aadB, and aadA2) and trimethoprim (dhfrXII). The integron-carrying strains of biotype Gallinarum appeared in 1996 and acquired additional cassettes in 2000. Although the resistances to ampicillin, tetracycline and chloramphenicol are unrelated to class 1 integrons, relatively high prevalence of integron in biotype Gallinarum may be a dormant threat to the chemotherapy of the disease in the near future because of potency to acquire additional antibiotics resistances.  相似文献   

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