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1.
Strains of Staphylococcus aureus obtained from bovine (n = 117) and caprine (n = 114) bulk milk were characterized and compared with S. aureus strains from raw-milk products (n = 27), bovine mastitis specimens (n = 9), and human blood cultures (n = 39). All isolates were typed by pulsed-field gel electrophoresis (PFGE). In addition, subsets of isolates were characterized using multilocus sequence typing (MLST), multiplex PCR (m-PCR) for genes encoding nine of the staphylococcal enterotoxins (SE), and the cloverleaf method for penicillin resistance. A variety of genotypes were observed, and greater genetic diversity was found among bovine than caprine bulk milk isolates. Certain genotypes, with a wide geographic distribution, were common to bovine and caprine bulk milk and may represent ruminant-specialized S. aureus. Isolates with genotypes indistinguishable from those of strains from ruminant mastitis were frequently found in bulk milk, and strains with genotypes indistinguishable from those from bulk milk were observed in raw-milk products. This indicates that S. aureus from infected udders may contaminate bulk milk and, subsequently, raw-milk products. Human blood culture isolates were diverse and differed from isolates from other sources. Genotyping by PFGE, MLST, and m-PCR for SE genes largely corresponded. In general, isolates with indistinguishable PFGE banding patterns had the same SE gene profile and isolates with identical SE gene profiles were placed together in PFGE clusters. Phylogenetic analyses agreed with the division of MLST sequence types into clonal complexes, and isolates within the same clonal complex had the same SE gene profile. Furthermore, isolates within PFGE clusters generally belonged to the same clonal complex.  相似文献   

2.
Strains of Staphylococcus aureus obtained from bovine (n = 117) and caprine (n = 114) bulk milk were characterized and compared with S. aureus strains from raw-milk products (n = 27), bovine mastitis specimens (n = 9), and human blood cultures (n = 39). All isolates were typed by pulsed-field gel electrophoresis (PFGE). In addition, subsets of isolates were characterized using multilocus sequence typing (MLST), multiplex PCR (m-PCR) for genes encoding nine of the staphylococcal enterotoxins (SE), and the cloverleaf method for penicillin resistance. A variety of genotypes were observed, and greater genetic diversity was found among bovine than caprine bulk milk isolates. Certain genotypes, with a wide geographic distribution, were common to bovine and caprine bulk milk and may represent ruminant-specialized S. aureus. Isolates with genotypes indistinguishable from those of strains from ruminant mastitis were frequently found in bulk milk, and strains with genotypes indistinguishable from those from bulk milk were observed in raw-milk products. This indicates that S. aureus from infected udders may contaminate bulk milk and, subsequently, raw-milk products. Human blood culture isolates were diverse and differed from isolates from other sources. Genotyping by PFGE, MLST, and m-PCR for SE genes largely corresponded. In general, isolates with indistinguishable PFGE banding patterns had the same SE gene profile and isolates with identical SE gene profiles were placed together in PFGE clusters. Phylogenetic analyses agreed with the division of MLST sequence types into clonal complexes, and isolates within the same clonal complex had the same SE gene profile. Furthermore, isolates within PFGE clusters generally belonged to the same clonal complex.  相似文献   

3.
Outbreaks of hypersensitivity pneumonitis (HP) among industrial metal-grinding machinists working with water-based metalworking fluids (MWF) have frequently been associated with high levels of mycobacteria in the MWF, but little is known about these organisms. We collected 107 MWF isolates of mycobacteria from multiple industrial sites where HP had been diagnosed and identified them to the species level by a molecular method (PCR restriction enzyme analysis [PRA]). Their genomic DNA restriction fragment length polymorphism (RFLP) patterns, as determined by pulsed-field gel electrophoresis (PFGE), were compared to those of 15 clinical (patient) isolates of the recently described rapidly growing mycobacterial species Mycobacterium immunogenum. A total of 102 of 107 (95%) MWF isolates (from 10 industrial sites within the United States and Canada) were identified as M. immunogenum and gave PRA patterns identical to those of the clinical isolates. Using genomic DNA, PFGE was performed on 80 of these isolates. According to RFLP analysis using the restriction enzymes DraI and XbaI, 78 of 80 (98%) of the MWF isolates represented a single clone. In contrast, none of the 15 clinical isolates had genetic patterns the same as or closely related to those of any of the others. Given the genomic heterogeneity of clinical isolates of M. immunogenum, the finding that a single genotype was present at all industrial sites is remarkable. This suggests that this genotype possesses unusual features that may relate to its virulence and its potential etiologic role in HP and/or to its resistance to biocides frequently used in MWF.  相似文献   

4.
The root voles intestinal strains of Bacillus thuringiensis, were characterised by pulsed-field gel electrophoresis (PFGE). For 14 isolates, three pulsotypes were found, with the use of SmaI or NotI as restriction enzymes. Strains in each pulsotypes presented identical DNA patterns, indicating that the population structure of B. thuringiensis from root voles is clonal. The similarities in banding patterns were estimated at 56% and 33% for SmaI and NotI digests, respectively. The strains under study differed significantly in the size of their entire genome, which varied between 2.4 and 4.2 Mb. No significant differences were detected among the isolates subjected to biochemical properties determined by API tests. Present study showed that genomic diversity is a common feature of B. thuringiensis originating from one ecological niche. PFGE appears to be a useful technique for use in studies on the spread of B. thuringiensis in the environment.  相似文献   

5.
Proteus mirabilis isolates (n = 177), collected between 1996 and 2000 in four hospitals in the West Pomeranian area of Poland, were characterized by antibiotype and pulsed-field gel electrophoresis (PFGE). The selected isolates were collected from different wards (intensive care unit, surgery, internal medicine, and urology). The strains were cultured from various specimen types, mostly from urine, wound samples, bronchial exudates and sputa. The identification was done by biochemical test ID 32E ATB (bioMerieux). Analysis of PFGE patterns was based on comparison of the banding patterns obtained by PFGE of chromosomal DNA digested with SfiI enzyme. Among all P. mirabilis isolates tested three major genotypes A (A1-A7), B (B1-B4), C (C1-C5) and 71 unique patterns were identified. The same genotypes were obtained from different patients, treated in different wards and hospitals during a 5-year period. The strains which belonged to the genotypes A and B were multiresistant and most of them produced ESBL; genotype C was more sensitive to antibiotics.  相似文献   

6.
Thirty-five Finnish Campylobacter jejuni strains with five SmaI/SacII pulsed-field gel electrophoresis (PFGE) genotypes selected among human and chicken isolates from 1997 and 1998 were used for comparison of their PFGE patterns, amplified fragment length polymorphism (AFLP) patterns, HaeIII ribotypes, and heat-stable (HS) serotypes. The discriminatory power of PFGE, AFLP, and ribotyping with HaeIII were shown to be at the same level for this selected set of strains, and these methods assigned the strains into the same groups. The PFGE and AFLP patterns within a genotype were highly similar, indicating genetic relatedness. The same HS serotypes were distributed among different genotypes, and different serotypes were identified within one genotype. HS serotype 12 was only associated with the combined genotype G1 (PFGE-AFLP-ribotype). These studies using polyphasic genotyping methods suggested that common Finnish C. jejuni genotypes form genetic lineages which colonize both humans and chickens.  相似文献   

7.
Although campylobacters have been isolated from a wide range of animal hosts, the association between campylobacters isolated from humans and animals in the farm environment is unclear. We used flagellin gene typing and pulsed-field gel electrophoresis (PFGE) to investigate the genetic diversity among isolates from animals (cattle, sheep, and turkey) in farm environments and sporadic cases of campylobacteriosis in the same geographical area. Forty-eight combined fla types were seen among the 315 Campylobacter isolates studied. Six were found in isolates from all four hosts and represented 50% of the total number of isolates. Seventy-one different SmaI PFGE macrorestriction profiles (mrps) were observed, with 86% of isolates assigned to one of 29 different mrps. Fifty-seven isolates from diverse hosts, times, and sources had an identical SmaI mrp and combined fla type. Conversely, a number of genotypes were unique to a particular host. We provide molecular evidence which suggests a link between campylobacters in the farm environment with those causing disease in the community.  相似文献   

8.
The genomic stability of 12 Campylobacter jejuni strains consisting of two groups of human and chicken isolates was studied by analysis of their PFGE (pulsed-field gel electrophoresis) patterns after passage through newly hatched chicks’ intestines. The patterns of SmaI, SalI, and SacII digests remained stable after intestinal passage, except for those of two strains. One originally human strain, FB 6371, changed its genotype from II/A (SmaI/SacII) to I/B. Another strain, BTI, originally isolated from a chicken, changed its genotype from I/B to a new genotype. The genomic instability of the strains was further confirmed by SalI digestion and ribotyping of the HaeIII digests. In addition, heat-stable serotype 57 of strain FB 6371 changed to serotype 27 in all isolates with new genotypes but remained unchanged in an isolate with the original genotype. Serotype 27 of strain BTI remained stable. Our study suggests that during intestinal colonization, genomic rearrangement, as demonstrated by changed PFGE and ribopatterns, may occur.  相似文献   

9.
Pulsed-field gel electrophoresis (PFGE) was used to investigate the dissemination and diversity of ampicillin-resistant (Ampr) and nalidixic acid-resistant (Nalr) commensal Escherichia coli strains in a cohort of 48 newborn calves. Calves were sampled weekly from birth for up to 21 weeks and a single resistant isolate selected from positive samples for genotyping and further phenotypic characterization. The Ampr population showed the greatest diversity, with a total of 56 different genotype patterns identified, of which 5 predominated, while the Nalr population appeared to be largely clonal, with over 97% of isolates belonging to just two different PFGE patterns. Distinct temporal trends were identified in the distribution of several Ampr genotypes across the cohort, with certain patterns predominating at different points in the study. Cumulative recognition of new Ampr genotypes within the cohort was biphasic, with a turning point coinciding with the housing of the cohort midway through the study, suggesting that colonizing strains were from an environmental source on the farm. Multiply resistant isolates dominated the collection, with >95% of isolates showing resistance to at least two additional antimicrobials. Carriage of resistance to streptomycin, sulfamethoxazole, and tetracycline was the most common combination, found across several different genotypes, suggesting the possible spread of a common resistance element across multiple strains. The proportion of Ampr isolates carrying sulfamethoxazole resistance increased significantly over the study period (P < 0.05), coinciding with a decline in the most common genotype pattern. These data indicate that calves were colonized by a succession of multiply resistant strains, with a probable environmental source, that disseminated through the cohort over time.  相似文献   

10.
This study investigates whether there is a predominant Staphylococcus aureus strain in retail foods and healthy human hands, and examines the relationship between pulsed-field gel electrophoresis (PFGE) banding patterns and the S. aureus characteristics of staphylococcal enterotoxin (SE) type, coagulase type, and β-lactamase activity. Ninety-four strains of S. aureus isolated from retail foods and healthy human hands were analyzed by PFGE. Several strains isolated from the same shop or a chain store showed identical patterns, indicating that the origins of these strains were identical. After excluding these strains showing identical patterns, 54 strains were used for the PFGE analysis. No spread of a particular clone in the environment surrounding the food was apparent. The PFGE analysis of these 54 strains was classified in 6 lineages (L1-L6). There was no relationship between the PFGE banding pattern and coagulase type or SE type. Eleven (84.6%) of the 13 isolates in PFGE banding pattern L5 did not produce β-lactamase, suggesting that the production of β-lactamase influenced a specific PFGE banding pattern.  相似文献   

11.
The root voles intestinal strains of Bacillus thuringiensis, were characterised by pulsed-field gel electrophoresis (PFGE). For 14 isolates, three pulsotypes were found, with the use of SmaI or NotI as restriction enzymes. Strains in each pulsotypes presented identical DNA patterns, indicating that the population structure of B. thuringiensis from root voles is clonal. The similarities in banding patterns were estimated at 56% and 33% for SmaI and NotI digests, respectively. The strains under study differed significantly in the size of their entire genome, which varied between 2.4 and 4.2 Mb. No significant differences were detected among the isolates subjected to biochemical properties determined by API tests. Present study showed that genomic diversity is a common feature of B. thuringiensis originating from one ecological niche. PFGE appears to be a useful technique for use in studies on the spread of B. thuringiensis in the environment. Received: 14 May 2002 / Accepted: 21 June 2002  相似文献   

12.
The genomic stability of 12 Campylobacter jejuni strains consisting of two groups of human and chicken isolates was studied by analysis of their PFGE (pulsed-field gel electrophoresis) patterns after passage through newly hatched chicks' intestines. The patterns of SmaI, SalI, and SacII digests remained stable after intestinal passage, except for those of two strains. One originally human strain, FB 6371, changed its genotype from II/A (SmaI/SacII) to I/B. Another strain, BTI, originally isolated from a chicken, changed its genotype from I/B to a new genotype. The genomic instability of the strains was further confirmed by SalI digestion and ribotyping of the HaeIII digests. In addition, heat-stable serotype 57 of strain FB 6371 changed to serotype 27 in all isolates with new genotypes but remained unchanged in an isolate with the original genotype. Serotype 27 of strain BTI remained stable. Our study suggests that during intestinal colonization, genomic rearrangement, as demonstrated by changed PFGE and ribopatterns, may occur.  相似文献   

13.
Outbreaks of hypersensitivity pneumonitis (HP) among industrial metal-grinding machinists working with water-based metalworking fluids (MWF) have frequently been associated with high levels of mycobacteria in the MWF, but little is known about these organisms. We collected 107 MWF isolates of mycobacteria from multiple industrial sites where HP had been diagnosed and identified them to the species level by a molecular method (PCR restriction enzyme analysis [PRA]). Their genomic DNA restriction fragment length polymorphism (RFLP) patterns, as determined by pulsed-field gel electrophoresis (PFGE), were compared to those of 15 clinical (patient) isolates of the recently described rapidly growing mycobacterial species Mycobacterium immunogenum. A total of 102 of 107 (95%) MWF isolates (from 10 industrial sites within the United States and Canada) were identified as M. immunogenum and gave PRA patterns identical to those of the clinical isolates. Using genomic DNA, PFGE was performed on 80 of these isolates. According to RFLP analysis using the restriction enzymes DraI and XbaI, 78 of 80 (98%) of the MWF isolates represented a single clone. In contrast, none of the 15 clinical isolates had genetic patterns the same as or closely related to those of any of the others. Given the genomic heterogeneity of clinical isolates of M. immunogenum, the finding that a single genotype was present at all industrial sites is remarkable. This suggests that this genotype possesses unusual features that may relate to its virulence and its potential etiologic role in HP and/or to its resistance to biocides frequently used in MWF.  相似文献   

14.
Following the identification of a case of severe clinical mastitis in a Saanen dairy goat (goat A), an average of 26 lactating goats in the herd was monitored over a period of 11 months. Milk microbiological analysis revealed the presence of Pseudomonas aeruginosa in 7 of the goats. Among these 7 does, only goat A showed clinical signs of mastitis. The 7 P. aeruginosa isolates from the goat milk and 26 P. aeruginosa isolates from environmental samples were clustered by RAPD-PCR and PFGE analyses in 3 genotypes (G1, G2, G3) and 4 clusters (A, B, C, D), respectively. PFGE clusters A and B correlated with the G1 genotype and included the 7 milk isolates. Although it was not possible to identify the infection source, these results strongly suggest a spreading of the infection from goat A. Clusters C and D overlapped with genotypes G2 and G3, respectively, and included only environmental isolates. The outcome of the antimicrobial susceptibility test performed on the isolates revealed 2 main patterns of multiple resistance to beta-lactam antibiotics and macrolides. Virulence related phenotypes were analyzed, such as swarming and swimming motility, production of biofilm and production of secreted virulence factors. The isolates had distinct phenotypic profiles, corresponding to genotypes G1, G2 and G3. Overall, correlation analysis showed a strong correlation between sampling source, RAPD genotype, PFGE clusters, and phenotypic clusters. The comparison of the levels of virulence related phenotypes did not indicate a higher pathogenic potential in the milk isolates as compared to the environmental isolates.  相似文献   

15.
Staphylococcus aureus subsp. anaerobius is the etiological agent of the Morel’s disease in sheep and goats. The disease presents with subcutaneous abscesses, located mainly in the superficial lymph nodes. Forty-one isolates of S. aureus subsp. anaerobius were collected from two outbreaks of the Morel’s disease in Poland in years 2006–2008. Analysis of DNA SmaI digests by PFGE showed that 35 of 41 isolates belonged to the same PFGE type, identical to the type strain of S. aureus subsp. anaerobius ATCC 35844, confirming high level of clonality of the species. The DNA patterns of the remaining identical 6 isolates, different from the reference strain only by two bands, were found closely related. Genotyping performed with AFLP technique revealed two clonal groups including 16 and 25 isolates, respectively. The study indicated that AFLP technique might be a better discriminatory tool for genetic analysis of S. aureus subsp. anaerobius isolates, when compared to PFGE.  相似文献   

16.
The clonal relationship among Salmonella enterica serotype Typhimurium isolates from selected pig production units in Denmark was investigated by the pulsed field gel electrophoresis (PFGE) typing method to determine environmental survival and spread of Salmonella in different herds. Thirty-four Typhimurium isolated during 1996-1998 from porcine faeces and environmental samples from three pig farms designated 1, 3 and 5 were characterised by PFGE using two restriction enzymes. Farm 5 supplied piglets to farm 1 and the herds were located close to each other. Results of PFGE analysis showed both intra- and inter-relationships, i.e. identical PFGE patterns among the faecal and environmental isolates from farm 1 and farm 5. All the isolates from farm 3 irrespective of the source showed identical PFGE patterns, but were different from samples from farms 1 and 5. This study indicates spread between farms and survival of a farm-specific clone. Furthermore, identical PFGE patterns of isolates from piglet supplier and finisher herds indicate that the farrow-to-grower herd of farm 5 was sub-clinically infected prior to delivery to farm 1 and thereby caused the transmission of Salmonella.  相似文献   

17.
Consumption and handling of chicken meat are well-known risk factors for acquiring campylobacteriosis. This study aimed to describe the Campylobacter jejuni population in Finnish chickens and to investigate the distribution of C. jejuni genotypes on Finnish chicken farms over a period of several years. We included 89.8% of the total C. jejuni population recovered in Finnish poultry during 2004, 2006, 2007, 2008, and 2012 and used multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) to characterize the 380 isolates. The typing data was combined with isolate information on collection-time and farm of origin. The C. jejuni prevalence in chicken slaughter batches was low (mean 3.0%, CI95% [1.8%, 4.2%]), and approximately a quarter of Finnish chicken farms delivered at least one positive chicken batch yearly. In general, the C. jejuni population was diverse as represented by a total of 63 sequence types (ST), but certain predominant MLST lineages were identified. ST-45 clonal complex (CC) accounted for 53% of the isolates while ST-21 CC and ST-677 CC covered 11% and 9% of the isolates, respectively. Less than half of the Campylobacter positive farms (40.3%) delivered C. jejuni-contaminated batches in multiple years, but the genotypes (ST and PFGE types) generally varied from year to year. Therefore, no evidence for a persistent C. jejuni source for the colonization of Finnish chickens emerged. Finnish chicken farms are infrequently contaminated with C. jejuni compared to other European Union (EU) countries, making Finland a valuable model for further epidemiological studies of the C. jejuni in poultry flocks.  相似文献   

18.
Pulsed-field gel electrophoresis (PFGE) was used to investigate the dissemination and diversity of ampicillin-resistant (Amp(r)) and nalidixic acid-resistant (Nal(r)) commensal Escherichia coli strains in a cohort of 48 newborn calves. Calves were sampled weekly from birth for up to 21 weeks and a single resistant isolate selected from positive samples for genotyping and further phenotypic characterization. The Amp(r) population showed the greatest diversity, with a total of 56 different genotype patterns identified, of which 5 predominated, while the Nal(r) population appeared to be largely clonal, with over 97% of isolates belonging to just two different PFGE patterns. Distinct temporal trends were identified in the distribution of several Amp(r) genotypes across the cohort, with certain patterns predominating at different points in the study. Cumulative recognition of new Amp(r) genotypes within the cohort was biphasic, with a turning point coinciding with the housing of the cohort midway through the study, suggesting that colonizing strains were from an environmental source on the farm. Multiply resistant isolates dominated the collection, with >95% of isolates showing resistance to at least two additional antimicrobials. Carriage of resistance to streptomycin, sulfamethoxazole, and tetracycline was the most common combination, found across several different genotypes, suggesting the possible spread of a common resistance element across multiple strains. The proportion of Amp(r) isolates carrying sulfamethoxazole resistance increased significantly over the study period (P < 0.05), coinciding with a decline in the most common genotype pattern. These data indicate that calves were colonized by a succession of multiply resistant strains, with a probable environmental source, that disseminated through the cohort over time.  相似文献   

19.
A total of 98 vancomycin-resistant Enterococcus faecium (VREF) isolates (58 isolates from patients and 40 isolates from poultry) were compared based on their antimicrobial susceptibility, Tn1546 element organization, and pulsed-field gel electrophoresis (PFGE) patterns. This comparison aided in determining the relationships between the groups of isolates. All the VREF isolates harbored the vanA gene; however, 29 (29.6%) of the isolates exhibited the VanB phenotype-vanA genotype. Furthermore, the VREF isolates from humans and poultry exhibited distinct antimicrobial resistance patterns. The PCR mapping of the Tn1546 elements exhibited 12 different transposon types (A to L). The VREF isolates of poultry were classified into types A to D, whereas the human isolates were classified into types E to L. A PFGE analysis demonstrated a high degree of clonal heterogeneity in both groups of isolates; however, the distinct VREF clones appeared in each group of isolates. The deletion of the vanX-vanY genes or insertion of IS1216V in the intergenic region from the vanX-vanY genes is directly associated with the incongruence of the VanB phenotype-vanA genotype in human VREF isolates. These data suggest that the VREF isolates exhibit distinct phenotypic and genotypic traits according to their origins, which suggests that no evidence exists to substantiate the clonal spread or transfer of vancomycin resistance determinants between humans and poultry.  相似文献   

20.
Thirty-five Finnish Campylobacter jejuni strains with five SmaI/SacII pulsed-field gel electrophoresis (PFGE) genotypes selected among human and chicken isolates from 1997 and 1998 were used for comparison of their PFGE patterns, amplified fragment length polymorphism (AFLP) patterns, HaeIII ribotypes, and heat-stable (HS) serotypes. The discriminatory power of PFGE, AFLP, and ribotyping with HaeIII were shown to be at the same level for this selected set of strains, and these methods assigned the strains into the same groups. The PFGE and AFLP patterns within a genotype were highly similar, indicating genetic relatedness. The same HS serotypes were distributed among different genotypes, and different serotypes were identified within one genotype. HS serotype 12 was only associated with the combined genotype G1 (PFGE-AFLP-ribotype). These studies using polyphasic genotyping methods suggested that common Finnish C. jejuni genotypes form genetic lineages which colonize both humans and chickens.  相似文献   

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