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PubMed1993

Adherence of haemagglutinating and non-haemagglutinating clinical and environmental isolates of Aeromonas.

Twelve haemagglutinating and non-haemagglutinating isolates of Aeromonas spp., comprising 6 each of clinical and environmental origin, were examined for their ability to adhere to rabbit intestinal epithelium, for inhibition of adhesion with sugars, and for delineation of the portion of intestine, jejunum, or ileum that is most susceptible to adhesion. Although the environmental isolates of Aeromonas haemagglutinated human erythrocytes that were inhibited by D-mannose and/or L-fucose, the majority of the clinical isolates of Aeromonas adhered to rabbit intestinal epithelium in almost equal proportions regardless of their haemagglutination (HA) properties, species designation, and source of isolation. Adhesion of both haemagglutinating and non-haemagglutinating isolates of Aeromonas was inhibited by sugars; however, the ability of sugar inhibition to adhere was similar to that observed with HA. This study suggests that adhesion is probably mediated by a variety of pilus or non-pilus colonisation factors which may or may not be a haemagglutinin. The jejunum was found to be more susceptible to adhesion than the ileum. However, no appreciable difference was observed in the number of adhered bacteria to adjacent loops.

Aeromonas↗

PubMed1992

Virulence characteristics of clinical and environmental isolates of Vibrio vulnificus.

Twenty-four randomly selected clinical and environmental Vibrio vulnificus isolates were tested for virulence in iron-overloaded mice (250 mg of iron dextran per kg of body weight). The log10 50% lethal doses of 17 isolates were lower by greater than or equal to 3.5 log10 units in iron-overloaded mice than in control mice. These isolates were classified as virulent. The 50% lethal doses of these virulent isolates were also lower in mice that were immunosuppressed by treatment with cyclophosphamide (150 mg/kg). Four of the seven isolates initially classified as avirulent were virulent in mice that were simultaneously iron overloaded and immunosuppressed. These isolates were classified as moderately virulent. The remaining three isolates were avirulent under all conditions. The incidence of virulent strains among clinical and environmental isolates did not differ. The virulent isolates produced high titers of hemolysin, were resistant to inactivation by serum complement, produced phenolate siderophore, and utilized transferrin-bound iron. The moderately virulent isolates differed from the virulent isolates only in their increased sensitivity to inactivation by serum complement. The avirulent isolates differed from those of the other two classes in their inability to either produce significant amounts of phenolate siderophore or utilize transferrin-bound iron. A modified agar plate diffusion method for transferrin-bound iron utilization was developed to differentiate the two classes of virulent isolates from the avirulent isolates in vitro.

Animals↗

PubMed2006

Antibiotic resistance and plasmid profile of environmental isolates of Vibrio species from Mai Po Nature Reserve, Hong Kong.

In this study, three environmental isolates of Vibrio species were isolated from water and sediment samples of Mai Po Nature Reserve, Hong Kong SAR. They were identified to be Vibrio cholerae MP-1, Vibrio aestuarianus MP-2 and Vibrio vulnificus MP-3 by both biochemical test and subsequent full-length 16S rDNA sequencing. Both disc diffusion susceptibility test and microplate dilution technique were used to investigate the antibiotic resistance profile of these three bacteria. Results from both assays were in generally good agreement. All three isolates were strongly resistant to beta-lactam family of antibiotics while susceptible to tetracycline, chloramphenicol, nalidixic acid and streptomycin. In addition, V. cholerae MP-1 was very tolerant to high concentration of vibriostatic agent O/129 to 40 microg/ml, at which the growth of V. aestuarianus MP-2 and V. vulnificus MP-2 was completely inhibited. A small naturally occurring plasmid was found in V. cholerae MP-1 while no plasmid was detected in the other two bacteria. This study suggests that Mai Po Nature Reserve harbors bacteria of unique characteristics that warrant further investigation.

Anti-Bacterial Agents↗

PubMed2005

Efficacy of species-specific recA PCR tests in the identification of Burkholderia cepacia complex environmental isolates.

In this study, we evaluated if recA species-specific PCR assays could be successfully applied to identify environmental isolates of the widespread Burkholderia cepacia complex (Bcc) species. A total of 729 Bcc rhizosphere isolates collected in different samplings were assigned to the species B. cepacia genomovar I (61), B. cenocepacia recA lineage IIIB (514), B. ambifaria (124) and B. pyrrocinia (30), by means of recA (RFLP) analysis, and PCR tests were performed to assess sensitivity and specificity of recA species-specific primers pairs. B. cepacia genomovar I specific primers produced the expected amplicon with all isolates of the corresponding species (sensitivity, 100%), and cross-reacted with all B. pyrrocinia isolates. On the contrary, B. cenocepacia IIIB primers did not give the expected amplicon in 164 B. cenocepacia IIIB isolates (sensitivity, 68.1%), and isolates of distinct populations showed different sensitivity. B. ambifaria primers failed to amplify a recA-specific fragment only in a few isolates of this species (sensitivity, 93.5%). The absence of specific amplification in a high number of B. cenocepacia rhizosphere isolates indicates that recA specific PCR assays can lead to an underestimation of environmental microorganisms belonging to this bacterial species.

Bacterial Proteins↗

PubMed2001

Clinical and environmental isolates of Vibrio cholerae serogroup O141 carry the CTX phage and the genes encoding the toxin-coregulated pili.

We report sporadic cases of a severe gastroenteritis associated with Vibrio cholerae serogroup O141. Like O1 and O139 serogroup strains of V. cholerae isolated from cholera cases, the O141 clinical isolates carry DNA sequences that hybridize to cholera toxin (CT) gene probes. The CT genes of O1 and O139 strains are carried by a filamentous bacteriophage (termed CTX phage) which is known to use toxin-coregulated pili (TCP) as its receptor. In an effort to understand the mechanism of emergence of toxigenic O141 V. cholerae, we probed a collection of O141 clinical and environmental isolates for genes involved in TCP production, toxigenicity, virulence regulation, and other phylogenetic markers. The collection included strains isolated between 1964 and 1995 from diverse geographical locations, including eight countries and five U.S. states. Information collected about the clinical and environmental sources of these isolates suggests that they had no epidemiological association. All clinical O141 isolates hybridized to probes specific for genes encoding CT (ctx), zonula occludens toxin (zot), repetitive sequence 1 (RS1), RTX toxin (rtxA), the major subunit of TCP (tcpA), and the essential regulatory gene that controls expression of both CT and TCP (toxR). In contrast, all but one of the nonclinical O141 isolates were negative for ctx, zot, RS1, and tcpA, although these strains were positive for rtxA and toxR. The one toxigenic environmental O141 isolate was also positive for tcpA. Ribotyping and CT typing showed that the O141 clinical isolates were indistinguishable or closely related, while a toxigenic water isolate from Louisiana showed a distantly related ribotype. Nonclinical O141 isolates displayed a variety of unrelated ribotypes. These data support a model for emergence of toxigenic O141 that involves acquisition of the CTX phage sometime after these strains had acquired the pathogenicity island encoding TCP. The clonal nature of toxigenic O141 strains isolated from diverse geographical locations suggests that the emergence is a rare event but that once it occurs, toxigenic O141 strains are capable of regional and perhaps even global dissemination. This study stresses the importance of monitoring V. cholerae non-O1, non-O139 serogroup strains for their virulence gene content as a means of assessing their epidemic potential.

Adult↗

PubMed1996

Diarrheal and environmental isolates of Aeromonas spp. produce a toxin similar to Shiga-like toxin 1.

Diarrheal and environmental isolates of 39 strains of Aeromonas spp. were studied for detection of virulence factors. Although these 39 strains did not produce either heat-labile or heat-stable enterotoxins, culture filtrates of 31 strains produced cytopathic effects on Vero cells. Among these, culture filtrates of three strains of Aeromonas hydrophila and one strain of Aeromonas caviae could be neutralized by Escherichia coli O157:H7 Shiga-like toxin 1 antiserum. A single band of plasmid DNA of 2.14 kbp was isolated from these strains of Aeromonas spp. and E. coli O157:H7, which could be amplified by the polymerase chain reaction (PCR), employing oligonucleotide primers from the Shiga-like toxin 1 (SLT1) gene of E. coli O157:H7. E. coli HB 101 cells when transformed with the same plasmid showed cytopathic effects on Vero cells, which indicates that the SLT 1 homolog gene(s) of Aeromonas spp. is plasmid encoded. These results suggest that Aeromonas spp. may also produce Shiga-like toxin 1, or at least a cytotoxin with some homology with the Shiga-like toxin 1 of E. coli O157:H7.

Aeromonas↗

PubMed2005

Consensus sequence-based scheme for epidemiological typing of clinical and environmental isolates of Legionella pneumophila.

A previously described sequence-based epidemiological typing method for clinical and environmental isolates of Legionella pneumophila serogroup 1 was extended by the investigation of three additional gene targets and modification of one of the previous targets. Excellent typeability, reproducibility, and epidemiological concordance were determined for isolates belonging to both serogroup 1 and the other serogroups investigated. Gene fragments were amplified from genomic DNA, and PCR amplicons were sequenced by using forward and reverse primers. Consensus sequences are entered into an online database, which allows the assignment of individual allele numbers. The resulting sequence-based type or allelic profile comprises a string of the individual allele numbers separated by commas, e.g., 1,4,3,1,1,1, in a predetermined order, i.e., flaA, pilE, asd, mip, mompS, and proA. The index of discrimination (D) obtained with these six loci was calculated following analysis of a panel of 79 unrelated clinical isolates. A D value of > 0.94 was obtained, and this value appears to be sufficient for use in the epidemiological investigation of outbreaks caused by L. pneumophila. The D value rose to 0.98 when the results of the analysis were combined with those of monoclonal antibody subgrouping. Sequence-based typing of L. pneumophila is epidemiologically concordant and discriminatory, and the data are easily transportable. This consensus method will assist in the epidemiological investigation of L. pneumophila infections, especially travel-associated cases, by which it will allow a rapid comparison of isolates obtained in more than one country.

Bacterial Typing Techniques↗

PubMed1999

Genotypic and phenotypic relationships between clinical and environmental isolates of Stenotrophomonas maltophilia.

While the gram-negative bacterium Stenotrophomonas maltophilia is used in biotechnology (e.g., for biological control of plant pathogens and for bioremediation), the number of S. maltophilia diseases in humans has dramatically increased in recent years. A total of 40 S. maltophilia isolates from clinical and environmental sources (plant associated and water) was investigated to determine the intraspecies diversity of the group and to determine whether or not the strains could be grouped based on the source of isolation. The isolates were investigated by phenotypic profiling (enzymatic and metabolic activity and antibiotic resistance patterns) and by molecular methods such as temperature-gradient gel electrophoresis of the 16S rRNA gene fragment, PCR fingerprinting with BOX primers, and pulsed-field gel electrophoresis (PFGE) after digestion with DraI. Results of the various methods revealed high intraspecies diversity. PFGE was the most discriminatory method for typing S. maltophilia when compared to the other molecular methods. The environmental strains of S. maltophilia were highly resistant to antibiotics, and the resistance profile pattern of the strains was not dependent on their source of isolation. Computer-assisted cluster analysis of the phenotypic and genotypic features did not reveal any clustering patterns for either clinical or environmental isolates.

Bacterial Typing Techniques↗

PubMed2002

Analysis of the Pseudomonas aeruginosa oprD gene from clinical and environmental isolates.

Genomes are constantly evolving. Our report highlights the wide mutational diversity of clinical as well as environmental isolates, compared with the laboratory strain(s), through the systematic genetic analysis of a chromosomal porin gene (oprD) in relation to a specific antibiotic resistance. Mutational inactivation of the oprD gene is associated with carbapenem resistance in Pseudomonas aeruginosa. The sequence of the oprD gene of 55 Pseudomonas aeruginosa natural isolates obtained from across the world--from sources as diverse as patients and rhizospheres--was analysed. A microscale mosaic structure for this gene--resulting from multiple intra- and possibly interspecies recombinational events--is reported. An array of independent and seemingly fast-occurring defective oprD mutations were found, none of which had been described before. A burn wound isolate demonstrated unusually high overall sequence variability typical of mutator strains. We also present evidence for the existence of OprD homologues in other fluorescent pseudomonads.

Amino Acid Sequence↗

PubMed2003

Gene function analysis in environmental isolates: the nif regulon of the strict iron oxidizing bacterium Leptospirillum ferrooxidans.

A random genomic library from an environmental isolate of the Gram-negative bacterium Leptospirillum ferrooxidans has been printed on a microarray. Gene expression analysis was carried out with total RNA extracted from L. ferrooxidans cultures in the presence or absence of ammonium as nitrogen source under aerobic conditions. Although practically nothing is known about the genome sequence of this bacterium, this approach allowed us the selection and sequencing of only those clones bearing genes that showed an altered expression pattern. By sequence comparison, we have identified most of the genes of nitrogen fixation regulon in L. ferrooxidans, like the nifHDKENX operon, encoding the structural components of Mo-Fe nitrogenase; nifSU-hesB-hscBA-fdx operon, for Fe-S cluster assembly; the amtB gene (ammonium transporter); modA (molybdenum ABC type transporter); some regulatory genes like ntrC, nifA (the specific activator of nif genes); or two glnB-like genes (encoding the PII regulatory protein). Our results show that shotgun DNA microarrays are very powerful tools to accomplish gene expression studies with environmental bacteria whose genome sequence is still unknown, avoiding the time and effort necessary for whole genome sequencing projects.

Bacteria↗

PubMed2004

Environmental isolation of Cryptococcus neoformans from an endemic region of HIV-associated cryptococcosis in Thailand.

We successfully isolated Cryptococcus neoformans from chicken faeces in suburban areas of Thailand. C. neoformans was isolated from 36/150 houses (24.0%) in the dry season and 6/150 (4.0%) in the rainy season. All environmental isolates were of serotype A. The high isolation rate of 24% from chicken faeces has never been reported previously. Our environmental study could probably explain the high incidence of cryptococcal meningitis in HIV patients in Thailand.

Animals↗

PubMed1998

Comparison of virulence between clinical and environmental isolates of Aspergillus fumigatus.

Using a mixed infection model of murine invasive pulmonary aspergillosis, the comparative virulence of three clinical and four environmental isolates of Aspergillus fumigatus was investigated after intranasal inoculation. Coloured conidiospore mutants were first derived from clinical strains by ultraviolet mutagenesis and then compared with the parental strains and environmental strains. When the slight reductions in virulence associated with the spore colour mutations were taken into account, some environmental strains were shown to be less virulent than their corresponding clinical strains. It has yet to be determined whether these differences can account for the observation that many patients with invasive pulmonary aspergillosis appear to be infected with a single strain of Aspergillus fumigatus.

Animals↗

PubMed1989

Homogeneity of a hemolysin(Vh-rTDH) produced by clinical and environmental isolates of Vibrio hollisae.

The characteristics of Vh-rTDH, a hemolysin similar to Vp-TDH of Vibrio parahaemolyticus produced by clinical and environmental isolates of Vibrio hollisae, were comparatively studied. All 7 strains of V. hollisae tested were found to produce indistinguishable Vh-rTDH when they were examined by heat-stability test, Western blotting analysis and conventional polyacrylamide slab gel electrophoresis.

Animals↗

PubMed1987

In vitro sensitivity of environmental isolates of pathogenic dematiaceous fungi to azole compounds and a phenylpropyl-morpholine derivative.

The in vitro sensitivity (minimum inhibitory concentrations; MICs) of 42 environmental isolates of pathogenic dematiaceous fungi to 7 azole compounds, viz. thiabendazole, ketoconazole, miconazole, econazole bifonazole, Bay n 7133, Bay 1 9139 and phenylpropyl-morpholine derivative, Ro14-4767/002 was studied by an agar dilution method using Emmon's Sabouraud dextrose agar (ESDA) as the culture medium. The isolates of Fonsecaea pedrosoi, Cladosporium carrionii, Exophiala jeanselmei and Ramichloridium subulatum were most sensitive to bifonazole with mean MICs of 0.06 microgram/ml or less; Phialophora verrucosa had an MIC of 0.05 microgram/ml to ketoconazole and Ro14-4767/002, respectively. Ochroconis sp had an MIC of 0.025 microgram/ml to Ro14-4767/002 and Cladosporium tennuisimum 0.39 microgram/ml to ketoconazole. Econazole and thiabendazole also showed good antifungal activity. The fungi were relatively resistant to the more recently developed azoles, viz. Bay n 7133 and Bay 1 9139, the later failing to inhibit C. tennuisimum at a concentration of 100 micrograms/ml. The minimum fungicidal concentrations (MFC) of the drugs were mostly within 2 to 8 fold of the MICs.

Animals↗

PubMed1998

Antigenic differences between clinical and environmental isolates of Burkholderia pseudomallei.

Burkholderia pseudomallei is a free-living organism that causes the potentially lethal tropical infection melioidosis. The disease is endemic in many parts of eastern Asia and northern Australia. The presence of two distinct biotypes in soil can be reliably distinguished by their ability to assimilate L-arabinose. Whereas some soil isolates could utilize this substrate (Ara+), the remaining soil isolates and all clinical isolates tested so far could not (Ara-). Only the Ara- isolates were virulent in animal models. We have raised a murine monoclonal antibody (MAb) that can readily distinguish Ara- from Ara+ biotypes. The MAb reacted with a high molecular weight component present only on the Ara- biotype. With this MAb, clinical and soil Ara- isolates gave identical positive reactions in agglutination, immunofluorescence, ELISA and immunoblot assays. Using these same assay systems, the soil Ara+ biotype did not react with the MAb. Similar but distinct immunoblot patterns were also noted when these two Ara biotypes were probed with sera from patients with melioidosis or with polyclonal immune rabbit sera. These data showed that the Ara- biotype from both clinical and environmental isolates is antigenically different from its Ara+ environmental counterpart. The SDS-PAGE protein and lectin-binding profiles of both groups of Ara- isolates were also found to be different from those of the Ara+ biotype.

Animals↗

PubMed1992

Identification and epidemiological typing of clinical and environmental isolates of the genus Rhodococcus with use of a digoxigenin-labeled rDNA gene probe.

Rhodococcus species are ubiquitous in the environment, and several species have been reported to have pathogenic potential for humans. Rhodococcus equi, in particular, has been reported to cause infections in patients with AIDS. However, the identification of Rhodococcus species with use of conventional biochemical tests is problematic, and no simple and reproducible method exists for their rapid identification and differentiation. We found that the type strains of the 20 recognized species in the genus Rhodococcus could be clearly distinguished with use of a combination of the Pvu II and Pst I rRNA gene restriction endonuclease patterns and a digoxigenin-labeled Escherichia coli rDNA probe. Analysis of four clinical or environmental isolates confirmed as Rhodococcus bronchialis showed no interstrain variation of rRNA gene bands. Analysis of 15 isolates confirmed as R. equi from 13 patients showed 11 different rRNA gene patterns. No discernible difference was observed in the ribotype patterns between R. equi isolates from patients for whom AIDS had been diagnosed and those from patients who did not have AIDS, and there was no evidence of geographic clustering of R. equi ribotype patterns. Three of five Rhodococcus species isolates that could not be differentiated with use of conventional biochemical methods were identified with use of ribotype analysis. Therefore, ribotype analysis may provide an important adjunct to current biochemical identification of environmental and clinical isolates of Rhodococcus species.

Acquired Immunodeficiency Syndrome↗

PubMed1999

Genotypic characterization of human and environmental isolates of Salmonella choleraesuis subspecies choleraesuis serovar infantis by pulsed-field gel electrophoresis.

To determine the extent of genetic diversity of Salmonella choleraesuis subspecies choleraesuis serovar Infantis and whether environmental isolates were similar or identical to human isolates, a total of 110 isolates from humans, broiler samples, egg production facilities, riverwater, sewage, and chicken meat were analyzed epidemiologically by pulsed-field gel electrophoresis. While the isolates showed 35 distinct pulsed-field profiles, none had the genotype of the human isolates. One pulsed-field profile was shared by 43 (39%) of the 110 isolates. These results indicate that relatively fewer clonal lines of S. serovar Infantis had spread widely while multiple clonal lines, including the strain involved in the outbreak, exist in Western Japan.

Animals↗

PubMed2004

Effects of several antifungal drug combinations against clinical and environmental isolates of Cryptococcus neoformans from China.

The in vitro interactions of caspofungin (CSP) with terbinafine (TRB) and ravuconazole (RVC) with 5-fluorocytosine (5-FC) were tested against 82 clinical and environmental isolates of Cryptococcus neoformans from China. The interaction of CSP with TRB proved synergistic against those isolates with a CSP MIC < or =2 microg ml-1 (5% of the isolates), additive against 42% of the isolates and indifferent against 53%. The effects of RVC with 5-FC were synergistic, additive or indifferent against 8%, 26% and 67% of the isolates, respectively. No antagonistic effects were found among any of the drugs. The combinations of CSP with TRB and RVC with 5-FC may display beneficial effects in a strain-dependent manner, while in no case showed antagonistic effects. These data might be of use to design safer and more efficient treatments for patients with cryptococcosis and warrant further evaluation.

Animals↗