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PubMed1980

Comparative analysis of plasmids and some metabolic characteristics of Escherichia coli K1 from diseased and healthy individuals.

All 62 Escherichia coli strains possessing the K1 capsular polysaccharide contained plasmid deoxyribonucleic acid, and most (51 of 62) had multiple plasmid species. The incidence of hemolysins, colicins, hemagglutinins for human erythrocytes, and plasmids did not differ among K1 strains isolated from the cerebrospinal fluids of neonates with meningitis or among those strains isolated from the stools of healthy individuals of all ages. There was an association between E. coli serotype and the distribution of plasmids, hemolysins, and colicins among the K1 strains. A common plasmid of about 65 megadaltons was found in all of the O18 serotypes; the similarity of these plasmids was confirmed by analysis with the restriction endonuclease EcoRI. Plasmids of similar molecular weight were also present in E. coli strains of the O7:K1 and O75:K100 serogroups. These data are consistent with the hypothesis that E. coli strains of the same serotype may be descendents of a single bacterial clone.

Adult↗
PubMedDate not supplied

Molecular epidemiology of bacterial infections: examples of methodology and of investigations of outbreaks.

Plasmid profile analysis, plasmid and chromosomal bacterial restriction endonuclease DNA analysis, and DNA hybridization are increasingly used in clinical microbiology and epidemiology. These techniques have been applied, singly and in combination, to investigations of outbreaks, including those of diarrheal diseases caused by pandemic Vibrio cholerae, enterotoxigenic and enterohemorrhagic Escherichia coli, Salmonella muenchen, and Salmonella typhimurium. The techniques have been critical in the unraveling of outbreaks of disease caused by nosocomial and community-acquired pathogens. Although there are limitations to these methods, their applications have important ramifications for basic science and public health.

Bacteria↗
PubMed1992

Plasmid pattern analysis of Staphylococcus species isolated from bovine mammary secretions.

Plasmid profiles of staphylococci isolated from bovine mammary secretions were heterogeneous as shown by the study of 94 isolates representing six species. Plasmids were identified in 19 of 94 staphylococcal isolates. Number of plasmids per isolate varied from 1 to 4. Size of plasmids ranged from 1.2 to 45 MDa; however, most were between 1.8 and 4.8 MDa. Some isolates with identical plasmid profiles were observed within and between species. Plasmid profiles observed in this study suggest that no specific plasmid pattern occurs within a species. Ability to differentiate isolates was not enhanced when antibiograms were used in conjunction with plasmid profiles. Plasmid pattern analysis does not appear to be an adequate method for discriminating between isolates of a species and would likely provide limited epidemiological information regarding staphylococci of bovine origin.

Ampicillin↗
PubMed1987

Antimicrobic susceptibility and plasmid profile analysis as identity tests for multiple blood isolates of coagulase-negative staphylococci.

We compared a disk diffusion antimicrobic susceptibility panel with plasmid DNA profiles as tests for identity of 106 isolates of coagulase-negative staphylococci cultured from the blood of 45 patients on multiple occasions. The antimicrobic panel included penicillin, oxacillin, clindamycin, trimethoprim-sulfamethoxazole, chloramphenicol, tetracycline, tobramycin, kanamycin, and gentamicin. Nineteen patterns of antimicrobic susceptibility were found. The most common pattern was present in 25% of the isolates, and at least one isolate from 31% of the patients had this pattern. Forty-seven distinct plasmid DNA profiles were found. The most common plasmid profile was present in 8.5% of the isolates, and at least one isolate from 15% of the patients had this profile. Twenty-eight patients had multiple isolates that were identical by plasmid profile analysis. Twenty-seven (96%) of these patients had isolates that were also identical by antimicrobic susceptibility. Nineteen patients had multiple isolates that were different by plasmid profile analysis. In 18 (95%) of these patients, the isolates were also different by antimicrobic susceptibility. Although plasmid DNA profile analysis is a more discriminating tool, these data confirm that a selected disk diffusion antimicrobic susceptibility panel may be used to screen multiple blood isolates of coagulase-negative staphylococci for identity or differences.

Anti-Bacterial Agents↗
PubMed1993

Evaluation of the molecular epidemiology of an outbreak of multiply resistant Shigella sonnei in a day-care center by using pulsed-field gel electrophoresis and plasmid DNA analysis.

Outbreaks of diarrhea in child day-care centers (DCC) are common. This study was undertaken to evaluate the molecular epidemiology of an outbreak of diarrhea due to Shigella sonnei. This outbreak involved 25 of 52 (48%) DCC children and 14 of 132 (11%) teachers and household contacts. S. sonnei isolates from nine children and five contacts were characterized by antimicrobial susceptibility, plasmid content, plasmid DNA restriction fragment pattern, and pulsed-field gel electrophoresis (PFGE) of total genomic DNA; 33 isolates from Houston, Tex., Chicago, Ill., and Mexico City, Mexico, also were studied. All outbreak isolates were resistant to ampicillin and trimethoprim-sulfamethoxazole and shared five to six plasmids ranging from 3.3 to 70 MDa. A total of 8 of 12 temporally associated nonoutbreak Houston isolates had plasmid profiles and restriction fragment patterns similar to those of the outbreak strain, despite possessing different antibiotic susceptibility patterns. PFGE demonstrated identical DNA patterns among outbreak isolates and similar or identical patterns among temporally associated sporadic Houston isolates with plasmid profiles similar to that of the outbreak strain. All other nonoutbreak strains from Houston, Chicago, and Mexico had plasmid profiles, restriction fragment patterns, and PFGE patterns different from those of the outbreak strain. DCC outbreak isolates could be distinguished from most sporadic isolates by antimicrobial susceptibility testing, but plasmid analysis and PFGE could not differentiate common-source isolates from sporadic isolates in the same location during the same time period, indicating that isolates present in the community were genetically similar to those producing outbreaks in the DCC.

Child Day Care Centers↗
PubMedDate not supplied

Phenotypic and plasmid pattern analysis of Staphylococcus epidermidis in bacterial keratitis.

BACKGROUND: Staphylococcus epidermidis, a commensal of the conjunctival sac has been incriminated as the commonest etiological agent of bacterial keratitis. However, the pathogenic potential of this commensal organism is not clearly known. AIM: To determine any phenotypic, molecular markers of S. epidermidis pathogenicity in bacterial keratitis. MATERIALS AND METHODS: A total of 382 corneal ulcer isolates of S. epidermidis and 87 S. epidermidis isolates from healthy eyes (controls) were studied. Speciation, biotyping and antibiotic sensitivity testing were performed by conventional methods. Tube slime and adherence tests were carried out by recommended techniques. Plasmid analysis was conducted by a standard protocol. STATISTICAL ANALYSIS: Chi-square test was employed for calculations. RESULTS: Out of 382 corneal ulcer isolates (Pathogens) 284 (74.3%) belonged to biotypes I and II. Slime was detected in 164 (42.9%) of 382 pathogens vs. 21 (24.1%) of 87 controls (P<0.001). Sixty-five (39.6%) of 164 slime positive isolates were multidrug-resistant as compared to only 49 (22.4%) of 218 slime negative isolates (P<0.001). A significantly higher number i.e, 73.1% (120/164) of slime-producers possessed a 21 Kb plasmid in contrast to only 53.2% (116/218) of nonslime-producers (P<0.001). Presence of this plasmid had a statistical correlation of low significance with multidrug resistance (P=0.04). One hundred and seventy-two (45.0%) of 382 pathogens and 24 (27.6%) of the 87 controls were adherent to artificial surfaces (P=0.003) and the majority of the adherent organisms (99/172, 57.6%) were slime producers (P<0.001). CONCLUSIONS: Slime was associated with multidrug resistance in corneal ulcer isolates of S. epidermidis. The 21 Kb plasmid could determine virulence as it was responsible for slime production and adherence.

Corneal Ulcer↗
PubMed1986

Plasmid profile analysis in epidemiological studies of animal Salmonella typhimurium infection in Japan.

Plasmid profiles were investigated in 65 isolates of Salmonella typhimurium derived from animal outbreaks during the period of 1978 through 1983 in Japan. Incidence of plasmids, drug-resistance, and conjugative R plasmids were extraordinarily high in these isolates. This high incidence reflects the prophylactic and therapeutic use of antibiotics. Most isolates from diseased animals, cohabiting animals, and each farm showed the same or similar plasmid patterns. However, there was a difference in plasmid patterns within strains isolated from each of several animals. It may be that one or two plasmids were introduced or deleted in these strains, leading to the difference discerned in strains isolated from the same animal. It was also shown that during one epidemic, two strains of S. typhimurium were involved that could be distinguished by plasmid profile analysis. Our conclusion is that when S. typhimurium strains isolated from animals reared in limited areas exhibit identical or similar plasmid patterns, they are derived from the same source and that when strains isolated in a limited area exhibit quite a different plasmid pattern, these strains are derived from independent sources.

Animals↗
PubMed1998

PCR methods and plasmid profile analysis for characterisation of Histophilus ovis strains.

The value of polymerase chain reaction (PCR)-based DNA fingerprinting and plasmid profile analysis for differentiation of Histophilus ovis isolates was assessed. Nineteen isolates of H. ovis were typed by PCR-ribotyping, repetitive extragenic palindromic element (REP)-PCR and enterobacterial repetitive intergenic consensus (ERIC)-PCR. These methods distinguished five types by PCR-ribotyping, 11 types by REP-PCR and seven types by ERIC-PCR. The ribotyping method produced a relatively simple pattern and a small number of distinct types and was useful for differentiation of H. ovis from the phenotypically similar organism, Haemophilus somnus. REP- and ERIC-PCR both produced complex banding patterns, but increased the discrimination between strains. Plasmids were found in 12 of the 19 isolates and there were four different plasmid profiles. A combination of the PCR methods and plasmid profile analysis provided a high resolution typing method for H. ovis.

Animals↗
PubMed1993

Restriction endonuclease analysis and ribotyping differentiate Pasteurella haemolytica serotype A1 isolates from cattle within a feedlot.

Pasteurella haemolytica serotype A1 isolates were collected from cattle within a feedlot during an outbreak of bovine respiratory disease. Genetic heterogeneity among the isolates was examined by restriction endonuclease analysis (REA), ribotyping, and analysis of plasmid content. The susceptibilities of isolates to several antibiotics were also examined. Five different REA patterns and three different ribotypes were observed among the isolates. Fifty percent of the isolates had an identical REA type, ribotype, and plasmid profile. Examination of the plasmid content of the isolates revealed that most (73%) carry a single plasmid which encodes beta-lactamase, 13.5% carry two plasmids, and 13.5% carry no plasmid. The data reveal the presence of genetic differences among isolates of P. haemolytica A1, associated with shipping fever pneumonia within a closed feedlot, and suggest that a combination of REA, ribotyping, plasmid analysis, and antibiotic susceptibility determination will be useful in analyzing the molecular epidemiology of this disease.

Animals↗
PubMedDate not supplied

Conventional and molecular typing of Salmonella typhi strains from Brazil.

Phenotypic and genotypic characteristics of Salmonella Typhi were studied in 30 strains, isolated in different years, from some areas in Brazil. Conventional typing methods were performed by biochemical tests, Vi phage-typing scheme, and antimicrobial susceptibility test. Molecular typing methods were performed by analysis of plasmid DNA and by random amplified polymorphic DNA (RAPD-PCR). For the latter, an optimization step was performed to ensure the reproducibility of the process in genetic characterization of S. Typhi. The predominance of 76.7% of biotype I (xylose +, arabinose -) was noticed in all studied areas. Three phage types were recognized, with prominence for the phage types A (73.3%) and I+IV (23.3%). All the strains were susceptible to the drugs used. However, 36.7% of the strains contained plasmids, with predominance of the 105 Kb plasmid. RAPD was capable of grouping the strains in 8 genotypic patterns using primer 784, in 6, using primer 787 and in 7, using primer 797. Conventional phenotypic typing methods, as well as the DNA plasmid analysis, presented nonsignificant discriminatory power; however, RAPD-PCR analysis showed discriminatory power, reproducibility, easy interpretation and performance, being considered as a promising alternative typing method for S. Typhi.

Bacterial Typing Techniques↗
PubMed1998

Computer-assisted restriction endonuclease analysis of plasmid DNA in field strains of Salmonella enteritidis.

Computer-assisted restriction endonuclease analysis of plasmid DNA in field strains of Salmonella enterica serovar Enteritidis (S. enteritidis) is described. The procedure consists of plasmid DNA purification, its digestion with restriction endonuclease TaqI, electrophoresis, charge-coupled device camera scanning of the gels, and an analysis of the restriction patterns with the software Gel Manager. The system allowed us to analyse, in detail, results of plasmid profiling in more than 600 field strains of S. enteritidis. In addition to plasmid-free and virulence plasmid only containing strains, 15 additional plasmid types were detected. All the images and detailed protocols are available at the Web site http://www.clark.cz/vri/salmon.htm.

Animals↗
PubMedDate not supplied

Clonal distribution of multiple-drug-resistant Shigella sonnei strains: identification by means of plasmid pattern analysis.

The spread of particular clones of S. sonnei throughout the territory of the German Democratic Republic within one year was evaluated by means of plasmid pattern analysis of the bacterial isolates. Such epidemic clones are described by their "clone formulae" comprising the phage type, biotype, colicinogenotype, antibiogram and the plasmid pattern type. They were not observed before 1982, but potential progenitor strains could be identified as having occurred sporadically in the years before. The additional application of plasmid pattern analysis revealed a better discrimination of the epidemic distribution of distinct bacterial clones than the use of conventional methods such as phage-typing, biotyping and colicinogenotyping only.

Bacteriological Techniques↗
PubMed1993

Molecular technology for hospital epidemiology.

Hospital-acquired infections in the United States contribute to approximately 80,000 deaths per year, with an associated cost of > $4 billion. Some of these infections are associated with outbreaks and clusters occurring within the hospital. The hospital infection control team must respond quickly and decisively to recognize and curtail these outbreaks, but their response often depends on critical laboratory data that characterize or "fingerprint" suspected microbial isolates. This presentation summarizes the simple, easily performed tests and the more molecular approaches that a hospital laboratory may take in support of infection control efforts. In addition to phenotypic data, such as biotype and antibiograms, hospitals that elect to incorporate molecular protocols should limit their first experiences to plasmid analysis and plasmid restriction endonuclease profiles.

Cross Infection↗
PubMed1988

Occurrence of multiple antibiotic resistance and R plasmids in Enterobacteriaceae isolated from children in the Sudan.

The prevalence of resistance to six commonly-used antimicrobial agents in faecal coliforms from children in Khartoum, Sudan was studied. A relatively high prevalence of resistance was found, ranging from 96% of children with isolates resistant to ampicillin to 70% of children with isolates resistant to chloramphenicol. Seventy-seven percent of children had isolates with high-level resistance to trimethoprim (MIC greater than 1000 micrograms/ml). Twenty-nine different resistance patterns were found. Thirty-nine percent of the children had isolates resistant to all six antibiotics studied, and 80% of children had isolates resistant to at least four. Transfer of resistance to each of the antimicrobials, in varying combinations, was demonstrated, but did not occur for all resistance patterns. Plasmid analysis showed plasmids ranging from 160 MDa to 2.8 MDa and isolates contained from one to five plasmids of different sizes. There were no consistent relationships between resistance pattern and plasmid profile, but multiple resistance transfer was mediated commonly by plasmids with a molecular weight of 62 MDa. The high prevalence of potentially transferable antibiotic resistance in gut commensals of children in the Sudan may be of importance in the management of enteric and other infections requiring antimicrobial treatment.

Ampicillin Resistance↗
PubMed2000

Molecular methods for the epidemiological typing of Salmonella enterica serotype Typhi from Hong Kong and Vietnam.

A total of 217 and 73 strains of Salmonella enterica serotype Typhi isolated from 1985 to 1997 in Hong Kong and in 2 months of 1989 and 1990 in Vietnam, respectively, were studied. These isolates were typed by plasmid profile analysis, plasmid fingerprinting, ribotyping with PstI, and total DNA fingerprinting with NarI. There appeared to be no major outbreak of typhoid fever in Hong Kong during the study period since there was considerable heterogeneity among the isolates. Isolates from Hong Kong were different from those from Vietnam. Thirty-seven percent of Vietnamese isolates belonged to two predominant clones, with the rest being heterogeneous in nature. Total DNA fingerprinting supplemented with ribotyping could be a reliable and rapid method for epidemiological typing of S. enterica serotype Typhi.

Bacterial Typing Techniques↗
PubMedDate not supplied

[Reliability of plasmid profile analysis in the identification of epidemic strains of bacteria causing an outbreak of intestinal infections].

The aim of the study was to determine the plasmid profile of isolates originated from the outbreaks of gastrointestinal tract infection in the attempt to identify the outbreak strains. Plasmid profile analysis was performed on 163 isolates; 82 isolates originated from foodborne outbreakes and 81 isolates originated from waterborne outbreakes or outbreakes spread by contact. Outbreak strain was identified in 1 (10%) of 10 foodborne outbreakes and in 7 (70%) of 10 waterborne outbreakes or outbreakes spread by contact. According to the obtained results, the method of plasmid profile analysis was more reliable in the investigation of outbreaks of gastrointestinal tract infection spread by water or contact compared to the outbreakes spread by food.

Bacterial Typing Techniques↗
PubMed1993

Differentiation of Salmonella enteritidis and S. typhimurium by plasmid profile analysis and restriction endonuclease analysis of chromosomal DNA.

Infections with Salmonella enteritidis and S. typhimurium are frequent causes of food-borne diseases in man and are responsible for considerable economic losses in the poultry industry (Fantasia et al., 1991; Pohl et al., 1991). Methods for the more careful differentiation and typing of these two serotypes are necessary for the investigation of the spread and transmission of the infection. Conventional methods of Salmonella differentiation (bioassays, phage-typing, resistance to antibiotics) often lack the necessary resolution potential (Wray et al., 1987). Plasmid profile analysis and restriction analysis of chromosomal DNA, based on restriction fragment length polymorphism (RFLP) have been used for the differentiation of Salmonellae (Wray et al., 1987; Nastasi et al., 1988; Franklin et al., 1990; Helmuth von et al., 1990; Martinetti and Altwegg, 1990). Rapid and unsophisticated analysis of the content of plasmid DNA tends to be preferred. However, some strains lack plasmids or may lose them during laboratory passages (Nastasi et al., 1988; Hartstein et al., 1991). On the other hand, restriction analysis of chromosomal DNA yields more constant and reliable information on bacterial strains (Tveten et al., 1991). The aim of this study was to compare 18 field strains of S. enteritis and 12 strains of S. typhimurium on the basis of plasmid profile analysis and restriction endonuclease analysis of chromosomal DNA. Plasmid DNA content was determined and chromosomal DNA, isolated from bacterial cells immobilized in low-melting agarose, was digested with restriction endonuclease Pst I in 18 and 12 field strains of S. enteritidis and S. typhimurium, respectively. The resulting fragments were separated by pulse-field electrophoresis in agarose gel.(ABSTRACT TRUNCATED AT 250 WORDS)

DNA, Bacterial↗