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Pulsed-field gel electrophoresis as an epidemiologic tool in the investigation of laboratory acquired Salmonella typhi infection.

Strains of Salmonella typhi implicated in two separate cases of laboratory acquired infection from patients and the medical laboratory technologists who processed the patients' samples were analysed by pulsed-field gel electrophoresis. Although all four isolates were of bacteriophage type E1, PFGE was able to demonstrate that the strains responsible for the two laboratory acquired cases were not genetically related. The PFGE patterns of the isolates from the MLTs were found to be identical to those of the corresponding patients after digestion with restriction enzyme AvrII. This provided genetic as well as epidemiological evidence for the source of the laboratory acquired infections.

Bacteriophage Typing↗

The naturally transformable marine bacterium WJT-1C formally identified as "Vibrio" is a pseudomonad.

A marine bacterial isolate, previously identified as Vibrio WJT-1C (ATCC 55351) and used as a model for investigating the process of natural transformation in the marine environment, has been further examined to determine its taxonomic identity. API 20E test strips, phenotypic testing, and flagellar staining had previously assigned the strain to the genus Vibrio, most closely related to V. campbelli. 16S rRNA analysis indicated that WJT-1C was in the Pseudomonas subgroup of the gamma proteobacteria. Bacteriophage typing and natural transformation with chromosomal DNA indicated that it was distinct from previously described marine transforming pseudomonads including Pseudomonas stutzeri strain JM300. The importance and abundance of the Pseudomonas subgroup of the gamma proteobacteria in the environment suggest that these marine strains are well suited as model organisms for describing the process and importance of natural transformation in nature.

Bacteriophage Typing↗

The measurement of C-reactive protein and immune complexes in endocarditis caused by coagulase-negative staphylococci.

Coagulase-negative staphylococci are a major cause of early endocarditis following the insertion of prosthetic heart valves but rarely cause endocarditis in natural valves. From a 2-year prospective study we report seven cases: six with endocarditis related to prosthetic valves and one with endocarditis related to a natural valve. Each isolate of coagulase-negative staphylococcus was identified biochemically (six Staphylococcus epidermidis; one Staphylococcus hominis) and characterised by bacteriophage typing. Six isolates were also examined for slime production and for extracellular toxins and enzymes: all produced toxin but enzyme and slime production was variable. Concentrations of immune complexes and C-reactive protein in the serum were measured in six of the patients. Our results suggest that measuring C-reactive protein may be useful but measuring immune complexes is not helpful and takes more time.

Antigen-Antibody Complex↗

ABSCESS-FORMING FACTOR(S) PRODUCED BY STAPHYLOCOCCUS AUREUS. II. ABSCESS FORMATION AND IMMUNITY BY A STAPHYLOCOCCUS AND ITS MUTANTS.

Lam, Gow T. (Jefferson Medical College, Philadelphia, Pa.), Francis J. Sweeney, Jr., Charlotte M. Witmer, and Robert I. Wise. Abscess-forming factor(s) produced by Staphylococcus aureus. II. Abscess formation and immunity by a Staphylococcus and its mutants. J. Bacteriol. 86:87-91. 1963.-Three mutants were derived from a virulent parent strain of Staphylococcus aureus by ultraviolet irradiation. One mutant retained coagulase but lost its hemolytic activity; a second had no coagulase but retained its hemolysin; and a third contained hemolysin and bound coagulase but no free coagulase. The parent strain was coagulase-positive, hemolysin-positive, and possessed all the known extracellular products. All four strains were compared by bacteriophage typing, mannitol utilization, and antibiotic sensitivity tests. The four isolates were studied for their pathogenicity in Swiss albino mice and four strains of inbred mice. Their immunological characteristics as studied by repeated subcutaneous injections are reported. The importance of hemolysin and the coagulase in abscess formation is discussed. Local immunity was produced only by the parent strain, but abscesses were produced by all the strains containing coagulase.

Abscess↗

Bacteriophage and bacteriocin typing scheme for Clostridium difficile.

The study of the epidemiology of infection with Clostridium difficile would be aided by a way to type individual bacterial isolates. We therefore sought bacteriophages for use in typing. With mitomycin C exposure (3 micrograms/ml), filtrates from 10 strains of C. difficile had plaque-forming lytic activity on other C. difficile strains. Individual phage were passaged and made into high-titer stock preparations for typing. Electron microscopy revealed tailed phage particles from one such preparation. In addition to phage, inhibitory activity without distinct plaque formation consistent with bacteriocins was observed for 20 strains. C. difficile isolates from 16 patients taken 1 to 14 days apart were similar in their phage sensitivity pattern, whereas isolates from separate geographic locations showed a great variety of patterns. We conclude that bacteriophage should be useful for typing strains of C. difficile.

Bacteriocins↗

[Outbreak of methicillin resistant Staphylococcus aureus in a central hospital].

Until 1994, methicillin resistant Staphylococcus aureus (MRSA) were isolated from less than 60 patients annually in Denmark. We describe an outbreak of MRSA involving eight patients within a timespan of six weeks. Bacteriophage-typing and DNA typing using pulsed field gel electrophoresis revealed that three different strains of MRSA were involved in the outbreak. This emphasizes the importance of typing MRSA in order to clear up the spread in an outbreak. Of the eight patients four died, and in one case MRSA was thought to be the major cause of death. The expenses of the extra hygienic measures i.e. isolation of patients, medication and bacteriological surveillance were estimated to approximately DKK 80.000 pr. patient. Patients, who have been admitted to or worked at a hospital outside Scandinavia within the last six months, are potentially colonized with MRSA. Specific guidelines for hospital hygiene are necessary to prevent the spread of such strains.

Adult↗

Staphylococcus aureus in patients with cystic fibrosis: an epidemiological analysis using a combination of traditional and molecular methods.

An epidemiological analysis of Staphylococcus aureus was conducted in a study group of 157 cystic fibrosis patients cultured over a 30-month period. The resulting S. aureus isolates were categorized by bacteriophage type, plasmid profile, and (in some instances) chromosomal restriction fragment pattern of the culture-positive patients with S. aureus (34 of 157) 44% only were sporadically infected while 68% shared identical strains with one or more other patients. Six patients exhibited persistent infection (for up to ten months) which, in three individuals, occurred as cycles of carriage and reappearance. By contributing toward our understanding of the persistence and spread of S. aureus in cystic fibrosis patients these data should aid in clarifying the role this organism may play in the course of the disease.

Bacteriophage Typing↗

Phage pattern-specific oxacillin-resistant and borderline oxacillin-resistant Staphylococcus aureus in U.S. hospitals: epidemiological significance.

For a 13-year period (1978 through 1990), oxacillin-resistant (MIC, greater than 4 micrograms/ml) Staphylococcus aureus (ORSA) strains were collected from Clinical Center (National Institutes of Health) patients and patients from five other U.S. hospitals. From Clinical Center patients, 251 of 253 isolates (99%) were bacteriophage typed as phage group III. Five other hospitals contributed 203 ORSA strains, of which 188 (93%) were group III. The group III ORSA strains predominantly included a characteristic core pattern of phages, 7/47/53/54/75/77. For the low-level (borderline) oxacillin-resistant strains (MIC, 2 to 4 micrograms/ml), amoxicillin-clavulanic acid combination (Augmentin) testing disclosed 62 hyper-beta-lactamase producers, of which 59 (95%) were of a separate, distinct S. aureus strain, with the phage pattern 92/94/96/292/D-11 (group V). Thus, ORSA and hyper-beta-lactamase producing S. aureus are distinct epidemic strains.

Bacteriophage Typing↗

Outbreak of Salmonella typhimurium gastroenteritis due to an imported strain resistant to ampicillin, chloramphenicol, and trimethoprim-sulfamethoxazole in a nursery.

An outbreak caused by a highly resistant strain of Salmonella typhimurium occurred in a nursery at a university medical center. The outbreak strain, which was resistant to ampicillin, chloramphenicol, and trimethoprim-sulfamethoxazole, was apparently imported from the Far East by a Cambodian refugee. The five patients involved had severe underlying diseases, and bacteremia and meningitis developed in one of these patients. The only reservoir identified was the gastrointestinal tracts of the infected patients, and infection was probably transmitted by the contaminated hands of hospital personnel. The outbreak was rapidly brought under control by isolating cases outside of the nursery and by instituting enteric precautions for infants who remained in the nursery. When compared by disk diffusion susceptibility tests with 353 strains of S. typhimurium tested at the Centers for Disease Control, the imported strain had a unique antibiogram. Bacteriophage typing of the strains revealed that all were untypable; this, in itself, was a good marker, because only 5 to 10% of S. typhimurium isolates in this country have this property. Agarose gel electrophoresis of isolates from the five patients revealed an identical plasmid banding pattern consisting of three large and three small plasmids. Highly resistant strains of S. typhimurium imported from the Far East may spread rapidly when introduced into a hospital nursery. Prompt institution of control measures may limit the outbreak and prevent systemic infections for which there are few effective therapeutic agents.

Ampicillin↗

Ciprofloxacin-resistant methicillin-resistant Staphylococcus aureus in an acute-care hospital.

Use of ciprofloxacin as an alternative to vancomycin for treatment of methicillin-resistant Staphylococcus aureus infection has been paralleled by the emergence of resistant strains. This phenomenon has also been noticed in our hospital. To confirm our observation, methicillin and ciprofloxacin susceptibilities were tested by disk diffusion and broth microdilution techniques. We studied 83 methicillin-resistant Staphylococcus aureus isolates obtained from various sources over a 4-month period. Ciprofloxacin resistance (MIC, greater than 2 micrograms/ml) was detected in 69 isolates (83%). Prior use of ciprofloxacin was reported for 24 of 69 patients with ciprofloxacin-resistant strains and 0 of 14 patients with ciprofloxacin-susceptible strains. The day of detection during the hospital stay and the location of the source patient were not significantly different between resistant and susceptible strains. Bacteriophage typing showed a higher occurrence of nontypeable strains among ciprofloxacin-resistant strains (54%). Review of our microbiology register showed a progressive increase in the rate of resistance to ciprofloxacin during the first year of use, with initial rates being about 10% and recent rates being higher than 80%. On the other hand, methicillin-susceptible S. aureus remained uniformly susceptible to ciprofloxacin (98.4%). We conclude that prior use of ciprofloxacin is an important factor for the selection of ciprofloxacin-resistant strains and that ciprofloxacin has limited usefulness against methicillin-resistant S. aureus.

Acquired Immunodeficiency Syndrome↗

A new methicillin- and gentamicin-resistant Staphylococcus aureus in Dublin: molecular genetic analysis.

In June 1985 two new strains of methicillin- and gentamicin-resistant Staphylococcus aureus (MGRSA) were isolated in a Dublin hospital. Of these, one strain spread rapidly, affecting a total of 65 patients during the following 18 months, and subsequently spread to a second Dublin hospital. Detailed laboratory studies, including plasmid screening, plasmid restriction enzyme digest pattern analysis, hybridisation analysis, location of resistance determinants, and bacteriophage typing with a set of experimental S. aureus typing phages, demonstrated that the "new" MGRSA organisms, termed Phenotype III Dublin isolates, were completely distinct from, and unrelated to, the MGRSA strains responsible for serious nosocomial infections in Dublin hospitals during the decade before June 1985. These Phenotype III isolates were very similar to MGRSA organisms isolated in a Baghdad hospital during 1984. Data from plasmid curing, plasmid transfer and hybridisation experiments indicated that 20% of the Phenotype-III isolates expressed chromosomally encoded, high level resistance to ethidium bromide (MIC 120 micrograms/ml), and that this was possibly due to chromosomal integration of a penicillinase-like plasmid.

Autoradiography↗

Genome fingerprinting of Pseudomonas aeruginosa indicates colonization of cystic fibrosis siblings with closely related strains.

The epidemiology of Pseudomonas aeruginosa infection at a cystic fibrosis (CF) center was monitored over a 3-year period. A total of 835 isolates from 72 unrelated patients and 22 siblings with CF were analyzed by genome fingerprinting and serotyping, bacteriophage typing, and pyocin typing. For genome fingerprinting, bacterial chromosomes were digested with one of the restriction endonucleases SpeI, DraI, XbaI, SspI, and NheI, which cut only rarely, and subsequently separated by field inversion gel electrophoresis. The physical genome analysis allowed us to classify P. aeruginosa strains in terms of DNA relatedness. Related strains differed by fewer than six DraI bands in the fingerprint, whereas unrelated strains differed by more than 20 DraI bands. All unrelated CF patients were colonized with different strains. The absence of a nosocomial spread of organisms at the CF center was attributed to the strict hygiene measures observed at the hospital. CF siblings were harboring either identical or closely related strains; transmission within the family is thought to be the most likely cause.

Bacteriophage Typing↗

Reemergence of epidemic methicillin-resistant Staphylococcus aureus in a general hospital associated with changing staphylococcal strains.

An epidemic of methicillin-resistant Staphylococcus aureus (MRSA) infections involving 323 patients occurred at the University of Cincinnati Hospital from 1977 to 1981. Subsequently, endemic MRSA persisted in the hospital for 6 years, until 1987, when a new epidemic began with 223 patients becoming infected over 3 years. Between the two epidemics, there was a major change in the MRSA recovered from infected patients, as demonstrated by three epidemiologic markers. Antibiograms showed that the tetracycline-resistant MRSA involved in the first epidemic was replaced by tetracycline-susceptible MRSA in the second epidemic; bacteriophage typing indicated that the original epidemic strain, D11/83A/85, had been replaced by new strains, many of which were susceptible to phage 54; and restriction endonuclease analysis of plasmid DNA confirmed that a single strain was involved in the first epidemic and that multiple strains were present in the second epidemic. The epidemiology of MRSA infections in the hospital changed with the change in staphylococcal strains. The first epidemic was hospital based with most infections occurring in surgical patients, and the burn unit was the major reservoir. In contrast, 28% of the patients in the second epidemic had community-acquired infections, and nursing home patients were an important source of these infections. Also, 29% of the hospital-acquired infections in this epidemic occurred in nonsurgical patients. This time the burn unit was not a reservoir of infection, but other intensive care units were. The increased diversity of strains of MRSA in the second epidemic might be related to increased transmission in the community and more widespread transmission in the hospital.

Bacteriophage Typing↗

Characterization of clinically significant isolates of Staphylococcus epidermidis from patients with endocarditis.

Biotyping, slime production, bacteriophage typing, serotyping, antibiograms, and plasmid profiles were used to characterize 19 Staphylococcus epidermidis strains isolated from 12 patients with prosthetic valve endocarditis and from 7 patients with native valve endocarditis. With the API Staph battery, 12 different biocodes with, at the most, three differences were obtained. Slime production was found for 10 strains (53%). Agglutinogens investigated by agglutination with two specific sera were found for 12 strains (63.1%). Three strains were phage typable (15.2%). Against a panel of nine antimicrobial agents, 15 different profiles were found. Multiply antibiotic-resistant strains were isolated from patients with prosthetic valve endocarditis when disease onset occurred less than 18 months after heart surgery and from patients with native valve endocarditis who received antibiotics immediately prior to their illness. All of the strains were available for plasmid analysis, and all the DNA profiles were distinct. On gels run in Tris-borate buffer, 73.7% of the strains had large plasmids of more than 30 megadaltons. A small plasmid of 2.8 megadaltons was found in multiply resistant strains and in strains resistant only to tetracyclines. None of the isolates appeared to be the same strain, and the bacteriological differences between the strains were confirmed mainly by the antibiotic susceptibility profile and the plasmid pattern analysis. These bacteriological results were in agreement with the clinical data.

Bacteriophage Typing↗

Investigation of a cluster of Staphylococcus aureus invasive infection in the top end of the Northern Territory.

INTRODUCTION: Staphylococcus aureus invasive infection remains a serious condition associated with considerable morbidity and mortality. Following notification of five cases at Royal Darwin Hospital (RDH), we searched for related cases, determined their epidemiological characteristics and attempted to identify the source of this apparent cluster. METHODS: We reviewed RDH microbiology records between June 1996 and April 1997 for S. aureus isolates with similar antibiograms to notified cases. We used antibiotic resistance patterns, bacteriophage typing and two molecular typing techniques to subtype implicated isolates. Hospital records were reviewed for admission details and associated costs were estimated. RESULTS: Fifty-four cluster-related isolates occurred in 47 separate presentations. The peak incidence was in the wet season. The most important risk factor for staphylococcal invasive infection was the presence of skin sores/scabies in 17/54 cases (31%), followed by intravascular line use in 14/54 (26%), open trauma in 11/54 (20%), underlying end stage renal failure and alcoholism each in ten of 54 (18%). The mean admission length was 30 days and antibiotics were given for an average of 23 days. Death due to S. aureus infection occurred in eight of 47 (17%) presentations. S. aureus pneumonia was community acquired in 12/13 patients (92%) and six of 13 (46%) died. Ten of 13 (80%) pneumonia patients had at least one other focus of S. aureus infection. The cost of antibiotics and hospital bed per presentation was approximately $16,000. Presentations with skin sores/scabies cost considerably more ($31,000). No common epidemiologic features were found for community or hospital acquired cases. CONCLUSION: Considerable mortality and cost was attributable to cases of S. aureus invasive infection during this cluster; particularly those with community acquired pneumonia or skin sores/scabies. Staphylococcal antibiotic cover should be considered early for unwell patients presenting to hospital with pneumonia and other signs of potential S. aureus infection. It is appropriate to target public health efforts to prevent skin sores and to provide adequate treatment when they occur.

Adolescent↗