Since 1990 multiresistant (MR) Salmonella enterica serotype Typhimurium definitive phage-type (DT) 104 (MR DT104) and closely related phage types have emerged as a worldwide health problem in humans and food animals. In this study the presence of the blaCARB-2 (ampicillin), cmlA (chloramphenicol), aadA2 (streptomycin/spectinomycin), sul1 (sulphonamide), and tetG (tetracycline) resistance genes in isolates of one such phage type, U302, have been determined. In addition blaTEM primers have been used for the detection of TEM-type beta-lactamases. Isolates have also been characterized by plasmid profile and pulsed field gel electrophoresis (PFGE). Thirty-three of 39 isolates were positive for blaCARB-2, cmlA, aadA2, sul1 and tetG, four for blaTEM, aadA2 and sul1, one for aadA2 and sul1, and one for blaTEM only. blaTEM-mediated ampicillin resistance was transferred to Escherichia coli K12 from three isolates along with other resistance markers, including resistance to chloramphenicol, streptomycin, spectinomycin, sulphonamides, and tetracyclines. Strains carried up to 6 plasmids and 34 plasmid profiles were identified. Although the majority of strains (33/39) produced a PFGE profile identical to that predominant in MR DT104, six different patterns were generated demonstrating the presence of various clones within MR U302. The results show that the majority of the MR U302 strains studied possessed the same antibiotic resistance genes as MR DT104. However, isolates with distinctive PFGE patterns can have different mechanisms of resistance to ampicillin, chloramphenicol, streptomycin, sulphonamides, and tetracyclines. Such resistance genes may be borne on transmissible plasmids.
Aims: To assess the degree of genetic diversity among animal Salmonella Dublin UK isolates, and to compare it with the genetic diversity found among human isolates from the same time period. Methods and Results: One hundred isolates (50 human and 50 animal) were typed using plasmid profiling, XbaI-pulsed field gel electrophoresis (PFGE) and PstI-SphI ribotyping. Antimicrobial resistance data to 16 antibiotics was presented, and the presence of class-I integrons was investigated by real-time PCR. Seven different plasmid profiles, 19 ribotypes and 21 PFGE types were detected. A combination of the three methods allowed clear differentiation of 43 clones or strains. Eighteen isolates were resistant to at least one antimicrobial; five of them were multi-resistant and of these, only three presented class I integrons. Conclusions: Ribotyping data suggest the existence of at least three very different clonal lines; the same distribution in well-defined groups was not evident from the PFGE data. The existence of a variety of clones in both animals and humans has been demonstrated. A few prevalent clones seem to be widely disseminated among different animal species and show a diverse geographical and temporal distribution. The same clones were found in animals and humans, which may infer that both farm and pet animals may act as potential vehicles of infection for humans. Some other clones seem to be less widely distributed. Clustering analysis of genomic fingerprints of Salmonella Dublin and Salm. Enteritidis isolates confirms the existence of a close phylogenetic relationship between both serotypes. Significance and Impact of the Study: This paper describes the utility of a multiple genetic typing approach for Salm. Dublin. It gives useful information on clonal diversity among human and animal isolates.
Pulsed-field gel electrophoresis of Escherichia coli O157:H7 isolates (n ؍ 228) from 122 healthy animals on 11 farms discriminated 57 types. Most clones were found only on individual farms. Numerous clones were found within each farm, with a prevalent clone normally found in several animals. A variety of clones were found within the different phage types.Escherichia coli strains that produce Shiga-like toxins may produce two toxins (Shiga toxin 1 [ST1] and/or ST2) with similar biological activities. A subset of Shiga-toxigenic E. coli strains designated as enterohemorrhagic (EHEC) carry sets of virulence genes (e.g., eaeA) that encode factors for attachment to host cells. Recent studies have revealed the existence of two divergent lineages (EHEC 1 and 2) (25). The EHEC 1 lineage consists solely of a few very closely related and geographically disseminated multilocus genotypes each bearing the O157 phenotype (15). Despite this similarity, pulsed-field gel electrophoresis (PFGE) and other molecular techniques have shown a considerable degree of genetic heterogeneity among O157 isolates in several countries, suggesting large evolutionary distances between strains. PFGE is considered the "gold standard" for fingerprinting of O157 strains and forms the basis of a large surveillance database (PulseNet) (24).Cattle are important reservoirs for this agent. Limited information is available about the genetic diversity of O157:H7 animal strains in the United Kingdom. Several studies have looked at human outbreak isolates, in most cases from Scotland (2,3,16,23). Some studies have described diversity in plasmid profiles in a limited number of animal isolates in comparison with isolates from food and humans (6,8,9). To our knowledge, one recent study (5) was the first to investigate the diversity of genotypes among animal isolates. The aim of the present study was to evaluate the genotypic diversity of O157:H7 isolates on a variety of cattle farms.Bacterial strains. E. coli O157:H7 (666 isolates) was cultured (7) (14) at 100 ng each, 2.5 mM MgCl 2 , and 2 l of bacterial DNA template. The PCR run consisted of 95°C for 10 min (one cycle), followed by 35 cycles of 95°C for 10 s, 65°C for 5 s, and 72°C for 25 s. The nature of the amplicon was determined by melting analysis over a temperature range from 70 to 97°C (0.1°C/s), and the melting temperature (T m ) was 87.5°C. The size of the product (625 bp) was verified by gel electrophoresis.
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