Phenotypic properties, drug susceptibility and genetic relatedness of Stenotrophomonas maltophilia clinical strains from seven hospitals in Rio de Janeiro, Brazil.

AIMS: To investigate phenotypic aspects including biotyping, drug susceptibility and production of extracellular enzymes and genetic diversity of Stenotrophomonas maltophilia clinical strains obtained from seven hospitals in Rio de Janeiro, Brazil. METHODS AND RESULTS: Thirty-nine S. maltophilia strains were investigated by biotying, susceptibility testing, extracellular enzymes detection and by randomly amplified polymorphic DNA (RAPD)-PCR. Biotyping distinguished 13 biotypes among 39, and one of them was prevalent. The majority of the strains produced DNase, gelatinase and haemolysin. Protease, lipases and phospholipase C activities were observed in highly variable amounts. None of the strains was elastase producer. The percentage of full susceptibility, by agar dilution, was 100, 94.8, 81.6 and 26.3% for trimethoprim/sulphametoxazole, ticarcillin/clavulanate, ciprofloxacin and ceftazidime, respectively. Thirty-three RAPD-PCR profiles were obtained suggesting multiple sources of acquisition. CONCLUSIONS: The results pointed out the necessity of monitoring S. maltophilia especially in critical hospital wards, to assure effective control measures. SIGNIFICANCE AND IMPACT OF THE STUDY: Despite of the genetic diversity among the strains, in two situations it was observed indistinguishable RAPD-PCR profiles among strains isolated from different patients who had been hospitalized in the same hospital ward, suggesting the possibility of nosocomial transmission that until now has been rarely related.

Leuconostoc pseudomesenteroides as a cause of nosocomial urinary tract infections.

The phenotypic and genotypic characterization of five clinical isolates of Leuconostoc pseudomesenteroides associated with nosocomially acquired urinary tract infections is described. All the strains were susceptible to chloramphenicol, clindamycin, erythromycin, gentamicin, and tetracycline; all were resistant to nalidixic acid, norfloxacin, and vancomycin; and all were intermediately affected by ampicillin and penicillin. Analysis of chromosomal DNA by pulsed-field gel electrophoresis after treatment with SmaI indicated a clonal relationship of the isolates. The results provide evidence for the possibility of nosocomial transmission of this unusual opportunistic, vancomycin-resistant pathogen.

Phenotypic and genotypic characterization of Vagococcus fluvialis, including strains isolated from human sources.

This study presents phenotypic and genotypic data for seven isolates of Vagococcus fluvialis, including four strains recovered from human clinical sources, one strain isolated from an environmental source, and two strains isolated from pigs. On the basis of phenotypic characteristics, most isolates were initially classified as "unidentified enterococci," because they resembled atypical arginine-negative enterococcal species. All seven strains as well as the type strain of V. fluvialis reacted with the AccuProbe Enterococcus genetic probe. The seven isolates had virtually indistinguishable whole-cell protein profiles that were similar to that of the V. fluvialis type strain and distinct from those of Enterococcus and Lactococcus species. DNA-DNA reassociation experiments confirmed that the strains were V. fluvialis. They were 71% or more related to the V. fluvialis type strain under optimum and stringent conditions, with 2.5% or less divergence within related sequences. All strains were susceptible to ampicillin, cefotaxime, trimethoprim-sulfamethoxazole, and vancomycin and were resistant to clindamycin, lomefloxacin, and ofloxacin. Strain-to-strain variation was observed in relation to susceptibilities to 18 other antimicrobial agents. Chromosomal DNA was analyzed by pulsed-field gel electrophoresis (PFGE) after digestion with SmaI. Distinctive PFGE patterns were generated, suggesting the nonclonal nature of V. fluvialis strains. Although the number of strains was small, this report provides molecular characterization of V. fluvialis and the first evidence of a possible connection of this species with human infections.

Phenotypic and genotypic characterization of atypical Lactococcus garvieae strains isolated from water buffalos with subclinical mastitis and confirmation of L. garvieae as a senior subjective synonym of Enterococcus seriolicida.

During a survey of bacterial agents that cause subclinical mastitis in water buffalos, we isolated several strains of gram-positive cocci that appeared to be enterococci except that they grew very slowly at 45 degrees C and grew slowly in broth containing 6.5% NaCl. On the basis of the results of conventional physiologic tests, these strains were identified as Enterococcus durans. However, none of the strains reacted with the AccuProbe Enterococcus genetic probe. The whole-cell protein profiles of these organisms were compared with the profiles of Enterococcus and Lactococcus reference strains. apart from minor quantitative differences, the mastitis isolates had indistinguishable protein profiles that were similar to the profiles of the Lactococcus garvieae and Enterococcus seriolicida type strains. The results of DNA relatedness studies performed by using the hydroxyapatite method at 55 and 70 degrees C indicated that all of the mastitis isolates were related to the type strain of L. garvieae at the species level, despite the fact that they exhibited several uncommon phenotypic characteristics (growth at 45 degrees C, growth in broth containing 6.5% NaCl, and failure to produce acid from mannitol and sucrose). The high levels of DNA relatedness between strains of L. garvieae is a senior synonym of E. seriolicida, L. garvieae should be retained as the species name and strain ATCC 43921 should remain the type strain of this species.

Partial characterization of the cohemolytic factor produced by Streptococcus uberis and comparison with the CAMP-factor.

Exosubstances (cohemolysins) produced by Streptococcus agalactiae (CAMP-factor) and Streptococcus uberis (Uberis-factor) showing hemolytic synergism with beta-lysin produced by Staphylococcus aureus were compared. Cohemolytic activity was evaluated in the supernatants of bacterial cultures, before and after ammonium sulfate precipitation. Sheep erythrocytes sensitized with beta-lysin were used as substrate. The assays were performed in microtiter plates and results were expressed as cohemolytic units/ml. Maximum cohemolytic activity was detected, respectively, after 8 h and 14 h of growth in Columbia broth in S. uberis and S. agalactiae cultures. Cohemolytic activities of both microorganisms showed similarities when submitted to various physical and chemical treatments. They were significantly decreased by heating at 60 degrees C and 100 degrees C, or in presence of trypsin, and were abolished in the presence of Tween 20. Activities were found to be stable in crude supernatants and concentrated preparations maintained at -20 degrees C for 3 months. Differences were related to levels of activity and kinetics of detection during the growth cycle. The results indicate the proteic nature, at least in part, of the Uberis factor. Analysis by PAGE in the presence or absence of SDS allowed us to correlate Uberis activity with a protein band with apparent molecular mass of 42 kDa, while CAMP activity was associated with a protein band of 27 kDa.

Correlation between phenotypic characteristics and DNA relatedness within Enterococcus faecium strains.

We noted that a number of enterococcal strains isolated from human clinical specimens resembled Enterococcus faecium but were able to produce acid from glycerol, raffinose, and/or sorbitol, while others failed to form acid from mannitol. An additional concern was that many of these strains with atypical phenotypic characteristics also appeared to acquire vancomycin resistance. In order to determine if such atypical strains were variants of E. faecium or new Enterococcus species, 35 E. faecium or E. faecium-like strains (grouped into 10 phenotypes on the basis of the results of the following tests: capacity to form acid from glycerol, mannitol, raffinose, or sorbitol and susceptibility to vancomycin) and four strains of Enterococcus faecalis were taken from our culture collection, analyzed for their whole-cell protein profiles by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and identified to the species level by DNA-DNA reassociation experiments. All E. faecium-like strains, including four mannitol-negative variants, conformed to at least two of three DNA-DNA relatedness criteria: they were 70% or more related to the type strain of E. faecium at optimal conditions, they had less than 5% divergence within the related sequences, and they had a relatedness of 60% or greater under stringent conditions. The protein profiles of atypical strains were similar to those of typical strains and were easily distinguishable from those of E. faecalis and other enterococcal species. The five E. faecalis strains were 12 to 16% related to the E. faecium type strain. These results indicate that the phenotypic description of E. faecium should include all of these variable characteristics.