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1.
Plasmid transfer and plasmid-mediated genetic exchange in Brucella abortus. 总被引：2，自引：0，他引：2 
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下载免费PDF全文 Naturally-occurring plasmids and gene transfer mechanisms have not yet been reported in brucellae. Here we show that Brucella abortus is capable of maintaining and transferring the broad-host-range plasmids pTH10 (IncP), pSa (IncW) and R751 (IncP), and describe pTH10-mediated transfer of B. abortus chromosomal genes to Escherichia coli. All three plasmids transferred by conjugation from E. coli to B. abortus S19, and from B. abortus S19 to B. abortus 292 (biovar 4). They were stably maintained with no effect on biotyping characteristics. Plasmid pTH10 is a Tn1-containing derivative of RP4. It confers temperature-sensitive resistance to kanamycin, tetracycline and ampicillin to E. coli, but its tetracycline resistance and temperature sensitivity were poorly expressed in B. abortus. Plasmids pTH10 and pSa both transferred from B. abortus to E. coli DP50, a strain that is auxotrophic for diaminopimelic acid (DAP) Plasmid pTH10 (but not pSa) mobilized Brucella chromosomal gene(s) for DAP synthesis to DP50, yielding non-DAP-requiring (NDR) transconjugants. Neither plasmid transferred the NDR marker from their original E. coli host strains, nor did pTH10 transfer it from NDR transconjugants. Escherichia coli NDR transconjugant EP8.11 was cured of pTH10 by passage at the nonpermissive temperature, but retained the NDR marker and the Tn1-encoded resistance to ampicillin, indicating Tn1-mediated integration of Brucella chromosomal DNA into the E. coli chromosome. 相似文献
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D Klimuszko Z M Szynkiewicz A Piekarowicz M Binek U Wójcik 《Comparative immunology, microbiology and infectious diseases》1989,12(1-2):29-38
The results of our study suggest the in vivo transfer of Hly plasmid from native pathogenic and enterotoxigenic Escherichia coli strain to autochtonous Escherichia coli, using ileal loop test. To confirm this hypothesis pHly::Tn5 and PHly::Tn3 were obtained using an in vitro recombination method, and introduced to Escherichia coli laboratory strain. For experiments the laboratory strain, carrying pHly::Tn5 and pHly::Tn3 and pHly::Tn5 strain which acquired K88(F4) by means of conjugation, were used. In the study in which the donor Escherichia coli pHly::Tn5 strain, carrying antigen K88(F4), was injected into the ileum, pHly conjugants were isolated from faeces after 48 h in 2 out of 5 pigs, which was a low frequency. After the oral introduction of 10(9) cells of pHly::Tn5 and pHly::Tn3 Escherichia coli strains without the colonizing factor K88(F4), conjugants were not isolated from faeces of experimental animals. However when the pigs received donor CSH55 pHly::Tn5 Escherichia coli strain orally, which were also carrying plasmid K88(F4), transconjugants were obtained in a low frequency of 3 out of 9 pigs. Our experiments confirmed the suggestion of Smith that in vivo transfer of plasmid in the intestine of animals is only possible when the donor transfers the plasmid with high frequency and readily colonizes the intestine. The pHly::Tn5 plasmid acquired by in vitro recombination does not spread with time throughout the autochtonic population of Escherichia coli present in the intestine of swine. The results of our study showed the in vivo transfer in pigs intestine of Hly pathogenicity marker from both native pathogenic strains carrying antigen K88(F4) and constructed donor laboratory strain of Escherichia coli pHly::Tn5 also carrying antigen K88(F4) to autochtonous intestinal strains. 相似文献
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Fifty six Danish streptomycin (Sm) resistant isolates of Salmonella enterica serotype Typhimurium from pigs (n=34), calves (n=3) and humans (n=19) were characterised with respect to co-resistances (14 drugs), transferability of Sm-resistance by conjugation, genetic determinants encoding Sm-resistance and diversity with respect to localisation of genes in the genome and DNA-sequences. Forty-six strains carried resistance(s) other than Sm-resistance. Nineteen different co-resistance patterns were observed and tetracycline was the most commonly observed resistance in these patterns. In 22 of the strains, Sm-resistance was transferred by conjugation. Eleven strains contained the gene aadA only, six strains contained aadA+strA+strB, and 35 strains contained strA+strB. Partial sequences of aadA were obtained from four strains. Three strains showed identical sequences to a published aadA sequence from the transposon Tn7, and in one strain the sequence showed one synonymous substitution compared to this sequence. Partial sequences were obtained of strA and strB in seven strains. The sequence of strB was identical to the published sequence of the plasmid RSF1010 in all strains. All seven sequences of strA were identical and differed from the sequence of strA in RSF1010 by two non-synonymous substitutions. 相似文献
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Development of a system for genetic and molecular analysis of Streptococcus agalactiae 总被引：4，自引：0，他引：4
DNA-DNA hybridisation was used to compare the genetic relation of human and bovine strains of Streptococcus agalactiae. All strains showed significant homology under very stringent hybridisation conditions. The extent of relatedness did not correlate with the serological type. It was demonstrated that the S faecalis transposon Tn916 could be inserted randomly into the S agalactiae chromosome when introduced by conjugation. The ability of Tn916 insertion to cause genetic changes in S agalactiae was confirmed by identification of a mutation in lactose and trehalose fermentation associated with acquisition of the transposon. This system should be useful in genetic analysis of the pathogenicity of S agalactiae. 相似文献
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Slater JD Allen AG May JP Bolitho S Lindsay H Maskell DJ 《Veterinary microbiology》2003,93(3):197-206
Genetic tools for studying streptococci are much less sophisticated than those that are available for many other bacterial genera. In this paper, we describe the development of a transposon mutagenesis system that we have used to mutate two important veterinary streptococci, Streptococcus equi and Streptococcus suis. The system uses a temperature-sensitive suicide vector to deliver Tn917 via electroporation, transposing Tn917 into the chromosomal DNA of the two streptococci. The transposon insertions can be rescued from the streptococcal chromosomes by plasmid rescue and selection in E. coli, with subsequent insertion site analysis by DNA sequencing. Transposition appeared to have occurred in an essentially random fashion when chromosomal DNA of S. suis and S. equi mutants was analysed by Southern blotting. However, when analysis of 60 S. equi mutants was carried out using the S. equi genome sequence database, 60% of transposon insertions had occurred within a 15 kb region of the genome whereas the other insertions appeared to have occurred essentially randomly. This finding suggests that Southern blot analysis for assessing the randomness of transposon libraries may need to be interpreted with caution. However, this observation notwithstanding, the Tn917 based system described in this paper will facilitate the study of S. suis and S. equi. 相似文献
6.
在构建的小肠结肠耶氏菌毒性质粒DNA基因文库pYB1～8与pYP1～6的基础上,筛选出了pYB7和pYP6克隆株.用限制性内切酶Bam HI消化pYB7,Pst消化pYP6,可分离出3.8kb和6.4kb的插入性DNA片段.以这两个基因片段为目的基因,用生物素化dUTP和光敏生物素标记,获得了生物素标记的基因探针.该探针能检出10pg以上的强毒小肠结肠耶氏菌DNA,不与无毒小肠结肠耶氏菌及大肠杆菌、鼠伤寒沙门氏菌、金黄色葡萄球菌等18种对照菌反应,具有高度的特异性和敏感性.pYB7与pYP6探针对不同血清型及来源的小肠结肠耶氏菌检测,其结果与自凝性试验、依钙试验等结果相符;对小肠结肠耶氏菌强毒株与无毒株检定的准确率为100%. 相似文献
7.
Y Wasteson O Olsvik 《Zentralblatt für Veterin?rmedizin. Reihe B. Journal of veterinary medicine. Series B》1991,38(6):445-452
Ten porcine strains of enterotoxigenic Escherichia coli possessing the K88 (F4) adhesion fimbriae, were selected for study of enterotoxin- and fimbriae-encoding plasmids. Plasmid DNA, separated according to size by gel electrophoresis was transferred to nylon membranes by Southern blotting, and hybridized with enzyme-labelled oligonucleotide probes for ST1 and LT1 enterotoxins, and a 32P-labelled probe for the F4 fimbriae. Plasmids possessing the enterotoxin genes ranged from 50 MDa to 78 MDa in size. The ST1 genes were located on a common 8-MDa EcoR1 restriction endonuclease fragment, while the LT1 genes were located on a 4.5-MDa EcoR1 fragment from the different plasmids. Plasmids with the F4 genes ranged from 50 MDa to 118 MDa in size, but the F4 encoding genes were located on a common 3-MDa HindIII restriction endonuclease fragment. ST1 and LT1 genes were found on the same plasmid in only one strain, LT1 and F4 genes on the same plasmids in 5 strains, while no plasmid contained genes for both ST1 and F4. 相似文献
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Vo AT van Duijkeren E Fluit AC Heck ME Verbruggen A van der Zwaluw K Gaastra W 《Veterinary microbiology》2006,117(2-4):192-200
Fifty-nine Salmonella enterica serovar Dublin (Salmonella Dublin) isolates from clinical cases of bovine salmonellosis between 1993 and 2004 were tested for their susceptibility to 15 antimicrobial agents and the presence of class 1 integrons. Integrons were further analyzed by conserved segment PCR-RFLP. DNA sequencing was used to identify the inserted gene cassette. Twelve (20.3%) isolates were multidrug-resistant. A combination of resistance against chloramphenicol, streptomycin and sulphonamides was the most common phenotype observed. Multidrug-resistance (MDR) was found to be strongly associated with the presence of integrons, since a class 1 integron with the aadA1 gene cassette encoding resistance to streptomycin and spectinomycin was found in all 12 multidrug-resistant isolates. The presence of the aadA1 gene in Salmonella Dublin has not been reported before. None of the integron carrying Salmonella Dublin isolates could transfer its antimicrobial resistance to E. coli K12 by conjugation. Analysis of plasmid profiles and pulsed field gel electrophoresis (PFGE) patterns showed at least some clonality among the Salmonella Dublin isolates, but 11 different types could be distinguished based on both XbaI and BlnI-PFGE patterns. Thus, the Dutch Salmonella Dublin strains were closely related but not clonal. 相似文献
11.
Survival of smooth, rough and transposon mutant strains of Brucella abortus in bovine mammary macrophages 总被引：3，自引：0，他引：3
Transposon mutants offer a unique way to evaluate the role of lipopolysaccharide (LPS) by producing a theoretical single-gene difference between the original strain and the transposon mutant strain. Comparative survival of Brucella abortus smooth strain 2308, rough RB51, smooth strain 19, and two transposon mutant strains (rough strain 2308::Tn5 Lac Z [m106] and rough strain 19::Tn5 Lac Z [m3], was tested in restrictive bovine mammary macrophages that were able to effectively reduce the percentage of intracellular bacterial survival and permissive bovine mammary macrophages that were unable to control the intracellular replication of B. abortus. The theoretical single-gene difference between strain 19 and strain 19::Tn5 lac Z [m3] and between smooth virulent strain 2308 and rough transposon mutant 2308::Tn5 lacZ [m106] is likely related to differences in LPS content or structure. Significant (P less than 0.05) reduction in the survival of rough strain 19::Tn5 Lac Z [m3] with no significant reduction in the rough transposon mutant strain 2308::Tn5 lacZ [m106] indicated that at least one factor other than LPS contributes to the intracellular survival of B. abortus in bovine macrophages. 相似文献
12.
The purpose of this study was to characterize E. coli isolates from canine pyometra which were isolated in pure culture. The E. coli strains were obtained in 128 cases, from 143 animals which were submitted to ovariohisterectomy. Biochemical analysis of all strains examined was possible on separation of 10 primary biotypes. The majority of the strains (87.5%) belonged to biotype 9, 1, 13 and 15. Dulcitol was fermented by 93% of all isolates. Haemolysin and colicin production was found in 53.9% and 26.6% of the strains, respectively. Approximately 37% of strains expressed resistance to two or more antibacterial agents. No plasmid was detected in 4.6% of the isolates. Plasmid profiles of all plasmid-containing isolates revealed plasmid bands corresponding to molecular weight ranging from 1 kb to 160 kb. Many of the strains examined had a single plasmid of 110 kb (46.1%), or two plasmids 110:65 kb (18.8%). Both plasmids appearing alone or in combination with other plasmids were detected in 90.1% of isolates with plasmid content. It was established that among haemolytic, colicinogenic and motile strains, the presence of both plasmids was 91.3, 94.1 and 91.4%, respectively. The appearance of both plasmids among dulcitol-positive and raffinose-negative strains was estimated at 88.2 and 88.3%, respectively. In a group of colicinogenic strains the presence of a single plasmid of molecular weight 110 kb was estimated at 5.9%. When both plasmids were present (profile 110:65), the percentage of these strains was 70.6%. 相似文献
13.
Cloning and comparison of heat-stable enterotoxin genes from Escherichia coli strains of bovine, porcine, and avian origins 总被引：1，自引：0，他引：1
The Ent plasmid encoding Escherichia coli heat-stable enterotoxin (ST) isolated from bovine, porcine, and avian strains were used for the cloning of ST genes. The ST+ DNA regions were finally cloned into pBR322 as the 1.75 kilobases PstI fragment. The electron microscopic analysis of self-annealed molecules indicated that ST+ recombinant plasmid had a stem-loop structure of a size the same as that observed in their wild type Ent plasmids. The stem-loop structures and the restriction enzyme cleavage mappings indicated that 4 kinds of ST genes cloned in this experiment may be identical to Tn1681. The ST production levels of the recombinant plasmids were higher than those of the original plasmids. 相似文献
14.
猪源大肠杆菌耐药性质粒的监测研究 总被引：1，自引：1，他引：0
在１１５株猪大肠杆菌耐药性监测的基础上,提取耐药类型最普遍的５株耐药菌的质粒进行质粒指纹图谱分析。结果表明：同一来源、耐药类型相同的菌株有相似的质粒图谱,来源不同者,酶切图谱有提示出它们之间的同源性,说明质粒指纹图谱可作为流行病学调查的工具。 相似文献
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The incompatibility plasmids of 101 strains Salmonella typhimurium were detected by PCR-based replicon typing. Eighteen pairs of primers were designed to perform 5 triplex-PCRs and 3 simplex-PCRs, recognizing FIA, FIB, FIC, HI1, HI2, I1, L/M, N, P, W, T, A/C, K, B/O, X, Y, F and FII. 94 strains (93.1%) Salmonella typhimurium carried a total of 10 Inc groups. The results suggested that the method was applicable for a large number of strains to trace the diffusion of specific multi-drug resistance plasmids in different environments. 相似文献
16.
LI Chen ZHANG Shuang-xiang ZHOU Bi-jun CHENG Zhen-tao WEN Ming MA Guang-qiang WANG Wei HU Xing-yi ZHANG Hai DING Zun-e WANG Kai-gong 《中国畜牧兽医》2017,44(12):3625-3633
In order to find out the serotype, resistant phenotype and genotype of Escherichia coli (E. coli) and Salmonella in piglets, this study collected 128 samples of diarrhea piglets from seven large-scale pig farms in five cities in Guizhou province, and the E. coli and Salmonella were isolated and identified. The pathogenicity of the strain was identified by animal test. The drug resistance of the main pathogen was tested by drug susceptibility paper. The resistance gene of each pathogen was detected by PCR. The drug resistance and genotype correlation of the bacterial were analyzed. The results showed that 78 strains of pathogenic E. coli and 21 strains of Salmonella were isolated and identified in this study. The serotypes of pathogenic E. coli were predominantly O138 and O87. Salmonella Typhimurium and Salmonella Enteritidis were predominant serotypes. The susceptibility test showed that the resistant strains of 78 strains of E. coli were more than 80% resistant to β-lactams and more than 40% for other antibacterials. The resistance rate of 21 strains of Salmonella to aminoglycosides was more than 50% and more than 20% to other types of antibacterials; 12 and 10 kinds of drug resistance-related genes of E. coli and Salmonella were detected, respectively; The coincidence rate of resistant genotype and phenotype of two kinds of bacteria were above 60%, and both were multiple drug resistance. This study provided a theoretical basis for comprehensive prevention and control of piglets diarrhea. 相似文献
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O Sk?ld 《Veterinary research》2001,32(3-4):261-273
Sulfonamides and trimethoprim have been used for many decades as efficient and inexpensive antibacterial agents for animals and man. Resistance to both has, however, spread extensively and rapidly. This is mainly due to the horizontal spread of resistance genes, expressing drug-insensitive variants of the target enzymes dihydropteroate synthase and dihydrofolate reductase, for sulfonamide and trimethoprim, respectively. Two genes, sul1 and sul2, mediated by transposons and plasmids, and expressing dihydropteroate synthases highly resistant to sulfonamide, have been found. For trimethoprim, almost twenty phylogenetically different resistance genes, expressing druginsensitive dihydrofolate reductases have been characterized. They are efficiently spread as cassettes in integrons, and on transposons and plasmids. One particular gene, dfr9, seems to have originally been selected in the intestine of swine, where it was found in Escherichia coli, on large plasmids in a disabled transposon, Tn5393, originally found in the plant pathogen Erwinia amylovora. There are also many examples of chromosomal resistance to sulfonamides and trimethoprim, with different degrees of complexity, from simple base changes in the target genes to transformational and recombinational exchanges of whole genes or parts of genes, forming mosaic gene patterns. Furthermore, the trade-off, seen in laboratory experiments selecting resistance mutants, showing drug-resistant but also less efficient (increased Kms) target enzymes, seems to be adjusted for by compensatory mutations in clinically isolated drug-resistant pathogens. This means that susceptibility will not return after suspending the use of sulfonamide and trimethoprim. 相似文献
18.
为探明仔猪细菌性腹泻肠道致病性大肠埃希氏菌和沙门氏菌流行的血清型、耐药表型及耐药基因型,本试验采集了贵州省5个地（州）市7个规模化养猪场的128份腹泻仔猪肠道样本,并对采集的样本进行了大肠埃希氏菌和沙门氏菌分离与鉴定,通过动物试验鉴定菌株的致病性,利用血清学方法鉴定其血清型,并通过药敏纸片法对主要致病菌进行耐药性研究,采用PCR技术检测各致病菌株耐药相关基因,分析细菌耐药表型和耐药基因型相关性。结果显示,本研究共分离鉴定到78株致病性大肠埃希氏菌与21株沙门氏菌,致病性大肠埃希氏菌血清型以O138、O87为主,沙门氏菌血清型以鼠伤寒沙门氏菌、肠炎沙门氏菌居多;药敏试验结果表明,本试验分离到的78株致病性大肠埃希氏菌对β-内酰胺类药物耐药率达80%以上,对其他种类的抗菌药耐药率均超过40%,分离鉴定的21株沙门氏菌对氨基糖苷类药物耐药率达50%以上,对其他种类的抗菌药耐药率均达20%以上;本试验分离鉴定的致病性大肠埃希氏菌共检出12种耐药相关基因,沙门氏菌共检出10种耐药相关基因,两种细菌耐药基因型与耐药表型符合率均达60%以上,且均为多重耐药。本研究为仔猪腹泻的综合防控提供了理论依据。 相似文献
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Plasmids of Salmonella enterica vary in size from 2 to more than 200 kb. The best described group of plasmids are the virulence plasmids (50-100 kb in size) present in serovars Enteritidis, Typhimurium, Dublin, Cholerae-suis, Gallinarum, Pullorum and Abortus-ovis. They all encode spvRABCD genes involved in intra-macrophage survival of Salmonella. Another group of high molecular weight plasmids are plasmids responsible for antibiotic resistance. Since most of these plasmids are conjugative, besides storage of genetic information, they contribute to the spread of genes in bacterial populations. The low molecular weight plasmids are the last group of plasmids found in S. enterica. Some of them have been shown to increase resistance to phage infection due to the presence of restriction modification systems. Despite limited knowledge on their function, their presence or absence is frequently used for strain differentiation in epidemiological studies. 相似文献