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PubMed2002

Type II topoisomerase quinolone resistance-determining regions of Aeromonas caviae, A. hydrophila, and A. sobria complexes and mutations associated with quinolone resistance.

Most Aeromonas strains isolated from two European rivers were previously found to be resistant to nalidixic acid. In order to elucidate the mechanism of this resistance, 20 strains of Aeromonas caviae (n = 10), A. hydrophila (n = 5), and A. sobria (n = 5) complexes, including 3 reference strains and 17 environmental isolates, were investigated. Fragments of the gyrA, gyrB, parC, and parE genes encompassing the quinolone resistance-determining regions (QRDRs) were amplified by PCR and sequenced. Results obtained for the six sensitive strains showed that the GyrA, GyrB, ParC, and ParE QRDR fragments of Aeromonas spp. were highly conserved (> or =96.1% identity), despite some genetic polymorphism; they were most closely related to those of Vibrio spp., Pseudomonas spp., and members of the family Enterobacteriaceae (72.4 to 97.1% homology). All 14 environmental resistant strains carried a point mutation in the GyrA QRDR at codon 83, leading to the substitution Ser-83-->Ile (10 strains) or Ser-83-->Arg. In addition, seven strains harbored a mutation in the ParC QRDR either at position 80 (five strains), generating a Ser-80-->Ile (three strains) or Ser-80-->Arg change, or at position 84, yielding a Glu-84-->Lys modification. No amino acid alterations were discovered in the GyrB and ParE QRDRs. Double gyrA-parC missense mutations were associated with higher levels of quinolone resistance compared with the levels associated with single gyrA mutations. The most resistant strains probably had an additional mechanism(s) of resistance, such as decreased accumulation of the drugs. Our data suggest that, in mesophilic Aeromonas spp., as in other gram-negative bacteria, gyrase and topoisomerase IV are the primary and secondary targets for quinolones, respectively.

4-Quinolones↗
PubMed1988

Virulence properties and enterotoxin production of Aeromonas strains isolated from fish.

The biological activities in vivo and in vitro of 59 motile Aeromonas spp. isolated from fish and water tanks were simultaneously analyzed in poikilothermic and homoiothermic systems. A total of 64.3% of the isolates tested were pathogenic for fish, and 62% of Aeromonas hydrophila and A. sobria isolates either virulent or nonvirulent for fish were enterotoxigenic. Although the majority of the strains were proteolytic and amylolytic and produced DNase, other activities, such as elastase and staphylolysis, were only present in A. hydrophila. Most of the strains (96%) produced hemolysins, and 68% had agglutinating capacity, but neither isolates pathogenic for fish nor enterotoxigenic isolates showed specificity for trout or human erythrocytes, respectively. The production of siderophores, agglutination in acriflavine, and precipitation after boiling were found not to be useful tests for screening virulent strains. Although statistical analysis revealed a significant relationship between virulence for fish and positive results for arabinose and sucrose fermentations, elastase, and hemolysis of human erythrocytes, only lysine decarboxylase showed a significant positive relationship with enterotoxigenicity. Using extracellular products from representative Aeromonas strains with different virulence markers and belonging to distinct O serogroups, we demonstrated a lack of correlation between cytotoxicity for fish and homoiothermic cell lines and pathogenicity. The extracellular products from selected pathogenic A. hydrophila strains were lethal for rainbow trout and displayed proteolytic, hemolytic, and cytotoxic activities which were simultaneously lost after heat treatment. The findings reported here indicate that it is not possible to establish a common and single mechanism involved in the invasion of Aeromonas spp. in poikilothermic and homoiothermic hosts.

Aeromonas↗
PubMed1999

Symbiosis of Aeromonas veronii biovar sobria and Hirudo medicinalis, the medicinal leech: a novel model for digestive tract associations.

Hirudo medicinalis, the medicinal leech, is applied postoperatively in modern medicine. Infections by Aeromonas occur in up to 20% of patients unless a preemptive antibiotic treatment is administered. The associated infections demonstrate the need for a better understanding of the digestive tract flora of H. medicinalis. Early studies reported the presence of a single bacterial species in the digestive tract and suggested that these bacteria were endosymbionts contributing to the digestion of blood. In this study, we cultivated bacteria from the digestive tract and characterized them biochemically. The biochemical test results identified the isolates as Aeromonas veronii biovar sobria. This species identification was supported by sequence comparison of a variable region of the genes coding for 16S rRNA. In a colonization assay, a rifampin-resistant derivative of a symbiotic isolate was fed in a blood meal to H. medicinalis. The strain colonized the digestive tract rapidly and reached a concentration similar to that of the native bacterial flora. For the first 12 h, the in vivo doubling time was 1.2 h at 23 degreesC. After 12 h, at a density of 5 x 10(7) CFU/ml, the increase in viable counts ceased, suggesting a dramatic reduction in the bacterial growth rate. Two human fecal isolates, identified as Aeromonas hydrophila and A. veronii biovar sobria, were also able to colonize the digestive tract. These data demonstrate that the main culturable bacterium in the crop of H. medicinalis is A. veronii biovar sobria and that the medicinal leech can be used as a model for digestive tract association of Aeromonas species.

Aeromonas↗
PubMed1999

Functional characterization of type IV pili expressed on diarrhea-associated isolates of Aeromonas species.

Our past work has shown that long, flexible type IV pili (single or in bundles) are the predominant pili expressed on fecal isolates of diarrhea-associated species of Aeromonas (Aeromonas veronii biovar sobria and A. caviae). They represent a family of type IV pili which we have designated Bfp (for bundle-forming pili). Reports from Japan suggest that Bfp are intestinal colonization factors. This study presents compelling evidence to support this conclusion. Aeromonas bacteria and/or Bfp purified from a strain of A. veronii biovar sobria were shown to adhere to epithelial and intestinal cell lines, freshly isolated human enterocytes, and fresh and fixed human and rabbit intestinal tissues, as determined by light and electron microscopy and immunohistochemical detection. Removal of Bfp by mechanical means decreased adhesion to cell lines by up to 80%. Purified Bfp blocked adhesion of the test strain to intestinal cells in a dose-dependent manner. Adhesion was also blocked by the Fab fraction of anti-Bfp immunoglobulin G. Moreover, ultrastructural studies (ruthenium red staining and transmission and scanning electron microscopy) demonstrated for the first time that Aeromonas adhesion to human enterocytes is pilus mediated and suggested that Bfp may also promote colonization by forming bacterium-to-bacterium linkages. Bfp-positive isolates examined for type IV pilus-mediated twitching motility in agar and slide culture assays developed for Pseudomonas aeruginosa did not, however, exhibit this function.

Aeromonas↗
PubMed2004

Aeromonas flagella (polar and lateral) are enterocyte adhesins that contribute to biofilm formation on surfaces.

Aeromonas spp. (gram-negative, aquatic bacteria which include enteropathogenic strains) have two distinct flagellar systems, namely a polar flagellum for swimming in liquid and multiple lateral flagella for swarming over surfaces. Only approximately 60% of mesophilic strains can produce lateral flagella. To evaluate flagellar contributions to Aeromonas intestinal colonization, we compared polar and lateral flagellar mutant strains of a diarrheal isolate of Aeromonas caviae for the ability to adhere to the intestinal cell lines Henle 407 and Caco-2, which have the characteristic features of human intestinal enterocytes. Strains lacking polar flagella were virtually nonadherent to these cell lines, while loss of the lateral flagellum decreased adherence by approximately 60% in comparison to the wild-type level. Motility mutants (unable to swim or swarm in agar assays) had adhesion levels of approximately 50% of wild-type values, irrespective of their flagellar expression. Flagellar mutant strains were also evaluated for the ability to form biofilms in a borosilicate glass tube model which was optimized for Aeromonas spp. (broth inoculum, with a 16- to 20-h incubation at 37 degrees C). All flagellar mutants showed a decreased ability to form biofilms (at least 30% lower than the wild type). For the chemotactic motility mutant cheA, biofilm formation decreased >80% from the wild-type level. The complementation of flagellar phenotypes (polar flagellar mutants) restored biofilms to wild-type levels. We concluded that both flagellar types are enterocyte adhesins and need to be fully functional for optimal biofilm formation.

Adhesins, Bacterial↗
PubMed2006

Analysis of the lateral flagellar gene system of Aeromonas hydrophila AH-3.

Mesophilic Aeromonas strains express a polar flagellum in all culture conditions, and certain strains produce lateral flagella on semisolid media or on surfaces. Although Aeromonas lateral flagella have been described as a colonization factor, little is known about their organization and expression. Here we characterized the complete lateral flagellar gene cluster of Aeromonas hydrophila AH-3 containing 38 genes, 9 of which (lafA-U) have been reported previously. Among the flgLL and lafA structural genes we found a modification accessory factor gene (maf-5) that is involved in formation of lateral flagella; this is the first time that such a gene has been described for lateral flagellar gene systems. All Aeromonas lateral flagellar genes were located in a unique chromosomal region, in contrast to Vibrio parahaemolyticus, in which the analogous genes are distributed in two different chromosomal regions. In A. hydrophila mutations in flhAL, lafK, fliJL, flgNL, flgEL, and maf-5 resulted in a loss of lateral flagella and reductions in adherence and biofilm formation, but they did not affect polar flagellum synthesis. Furthermore, we also cloned and sequenced the A. hydrophila AH-3 alternative sigma factor sigma54 (rpoN); mutation of this factor suggested that it is involved in expression of both types of flagella.

Aeromonas hydrophila↗
PubMed1981

Association of Aeromonas sobria with human infection.

Fifteen Aeromonas isolates from various human infections and nine isolates from polluted water were identified as either Aeromonas hydrophila or Aeromonas sobria and examined for cytotoxigenicity, enterotoxigenicity, adherence to epithelial cells, and other virulence-associated factors, including proteases, lipases, elastases, and hemolysins. Two groups of organisms (I and II) were distinguishable based on differences in median lethal doses in mice and cytotoxicity for Y-1 adrenal cells. Group I clinical and environmental strains had median lethal doses of less than 10(7) colony-forming units, were cytotoxic, frequently possessed several virulence-associated factors, and had lysine decarboxylase-positive or Voges-Proskauer-positive phenotypes or both. Piliation of Aeromonas was associated strongly with ability to adhere to human buccal cells, and these characteristics were associated with group I strains. Group II clinical and environmental strains had median lethal doses of greater than or equal to 10(7) colony-forming units, were not cytotoxic, and usually were lysine decarboxylase negative or Voges-Proskauer negative or both. Clinical strains in group II exhibited enterotoxigenicity, which was not detected in group II environmental strains. A sobria was more frequently associated with human infections; 13 of the 15 clinical strains were A. sobria, and 2 were A. hydrophila. On the other hand, the majority of the environmental strains (seven of nine) were A. hydrophila.

Aeromonas↗
PubMed1984

Biotyping of Aeromonas isolates as a correlate to delineating a species-associated disease spectrum.

A group of 147 Aeromonas isolates from diverse clinical and environmental sources was subjected to the biotyping scheme of Popoff and Veron. Of the 147 isolates biotyped, 137 (93%) could be identified, with Aeromonas hydrophila predominating (48%) and equal percentages (25 to 27%) of the other two species (Aeromonas sobria and Aeromonas caviae). A number of additional biochemical properties were found to be significantly associated with one or more of these three species. These included lysine decarboxylase activity, hemolysis of sheep erythrocytes, lecithinase production, staphylolytic activity, arbutin hydrolysis, and acid production from utilization of various carbohydrates. By incorporating these phenotypic properties into an extended biotyping system, 98% of the isolates were identified. Selective distribution of individual species with respect to certain body sites was noted.

Aeromonas↗
PubMed1987

Comparison of selective media for primary isolation of Aeromonas species from human and animal feces.

Five selective media were evaluated for their effectiveness in the primary isolation of Aeromonas spp. from human patients with acute diarrhea and from healthy domestic animals. Sheep blood agar with 30 mg of ampicillin per liter (ASBA30) yielded a significantly higher percentage of positive specimens as compared with the four other media. The effective combination of two selective media with which 98% of all isolates were detected and with which all of the three human-associated Aeromonas spp. could be isolated was ASBA30-DNase-toluidine blue agar. ASBA30 was the most sensitive medium since it permitted more growth of Aeromonas colonies and effectively suppressed competing microflora. We recommend the use of ASBA30-DNase-toluidine blue agar for investigations in which an attempt is made to assess the significance of Aeromonas spp. in the etiology of human diarrhea.

Acute Disease↗
PubMed1987

Clinical significance of Aeromonas species isolated from patients with diarrhea.

A total of 248 strains of Aeromonas spp. were isolated from 3,334 human fecal specimens submitted to a state public health laboratory over a 2-year period to be cultured for enteric pathogens. Cary-Blair transport medium, blood ampicillin agar, and alkaline peptone water enrichment provided optimal recovery of Aeromonas spp. A questionnaire requesting clinical and epidemiological information was sent to physicians, who submitted stool samples for testing, with each laboratory report for 107 consecutive stool isolates of Aeromonas spp. The 56 questionnaires which were completed and returned were analyzed to determine the seasonal distribution of illness and the age and sex distribution of patients; characteristic symptoms; and predisposing factors for gastrointestinal disease caused by Aeromonas spp. It was concluded that some A. hydrophila, A. sobria, and A. caviae strains are capable of causing diarrhea and that antibiotic therapy and the drinking of untreated water are significant risk factors for susceptible hosts.

Adolescent↗
PubMed1987

Aeromonas veronii, a new ornithine decarboxylase-positive species that may cause diarrhea.

In 1983, the vernacular name Enteric Group 77 was coined for a group of strains that had been referred to our laboratory as "possible Vibrio cholerae except for gas production." By DNA-DNA hybridization (hydroxyapatite, 32P), 8 of 10 strains of Enteric Group 77 were very highly related to the labeled strain 1169-83 (74 to 100% at 60 degrees C and 75 to 100% at 75 degrees C; percent divergence, 0.0 to 2.5). Type strains of six other Aeromonas species were 45 to 66% related (60 degrees C) to strain 1169-83, but type strains of 27 Vibrio species were only 2 to 6% related. The name Aeromonas veronii is proposed for the highly related group of nine strains formerly known as Enteric Group 77. The type strain is designated as ATCC 35604 (CDC 1169-83). Strains of A. veronii grew well at 36 degrees C and had positive reactions at this temperature for indole, methyl red, Voges-Proskauer, citrate, lysine and ornithine decarboxylases, DNase, lipase, and motility; the strains had negative reactions for arginine decarboxylase, H2S, urea, and malonate. The following sugars were fermented: D-glucose (acid and gas), cellobiose (seven of nine strains), D-galactose, maltose, D-mannitol, D-mannose, alpha-methyl-D-glucoside (eight of nine strains), salicin, sucrose, and trehalose. The following sugars were not fermented: adonitol, L-arabinose, D-arabitol, dulcitol, erythritol, myo-inositol, lactose, raffinose, L-rhamnose, D-sorbitol, and D-xylose. The positive ornithine decarboxylase reaction differentiates A. veronii from other Aeromonas species. The antibiogram of A. veronii is typical of other Aeromonas strains (resistance to ampicillin and carbenicillin and susceptibility to most other agents). A. veronii strains were isolated from three clinical sources: respiratory secretions of four victims of drowning or near drowning in fresh water (probably not clinically significant); infected wounds of two patients previously exposed to fresh water (unknown clinical significance); and stools from three patients with diarrhea (probably clinically significant).

Adult↗
PubMed1987

Clinical and biochemical significance of toxin production by Aeromonas hydrophila.

Production of cytotoxin and enterotoxin by Aeromonas strains obtained from stools of 50 children in Mexico and Texas and from blood of 9 children with sepsis was determined. Results were correlated with clinical features of infected children as well as with biochemical traits of Aeromonas strains. Cytotoxin was produced by 40 of 42 Aeromonas strains (95%) isolated from stools of children with diarrhea, by all 8 isolates from stools of well children, and by all 9 isolates from children with sepsis. There was no difference in the quantities (amount of cytotoxin per milligram of protein required to kill 50% of the cells) of cytotoxin produced and in clinical manifestations among the groups. None of the isolates produced a toxin that could be neutralized by antiserum raised against Shiga toxin produced by Shigella dysenteriae 1 60R. Heat-labile-like enterotoxin (LT) was produced by 26 of 42 stool isolates (62%), while only 1 of the 42 isolates (2%) produced enterotoxinlike activity in suckling mice; 65% of the cytotoxin-producing strains also produced an LT-like material. All strains from blood produced LT-like material, and 2 of 6 (33%) produced activity in suckling mice. All strains produced hemolysin; 37 of 57 (65%) were Voges-Proskauer positive; 27 of 57 (47%) were lysine decarboxylase positive by API 20E strips, none were positive for lysine decarboxylose production by lysin-iron agar slants at 24 h, but 17 of 54 (31%) were positive at 48 h. There was no correlation between biochemical reactions and enterotoxin or cytotoxin production. There appears to be no correlation between toxin production by Aeromonas spp. and gastroenteritis.

Acute Disease↗
PubMed1989

Species identification of Aeromonas strains based on carbon substrate oxidation profiles.

Twenty clinical strains each of Aeromonas hydrophila, Aeromonas caviae, and Aeromonas sobria were evaluated for their abilities to oxidize one or more of 95 carbon sources on a GN Microplate (BIOLOG, Hayward, Calif.). Nine substrates yielded good, discriminatory values for the three species tested. The panel appears to be useful for the species identification of Aeromonas isolates originating from human material.

Aeromonas↗
PubMed1990

Pathogenicity factors and virulence for rainbow trout (Salmo gairdneri) of motile Aeromonas spp. isolated from a river.

Ninety-seven motile Aeromonas strains were isolated over a period of a year from samples of water and sediment collected at different sites along a river. Strains were regularly recovered from all samples, regardless of the source of isolation or seasonal conditions. Isolates were biochemically characterized by the API 20NE system (Analytab Products, Plainview, N.Y.) and classified as Aeromonas hydrophila (74 strains), Aeromonas sobria (11 strains), and Aeromonas caviae (12 strains). Despite the high level of homogeneity observed in their biochemical patterns, they displayed different degrees of virulence for fish; 72.02% of A. hydrophila isolates and 63% of A. sobria isolates were virulent for fish by intramuscular challenge, but lower frequencies of virulence were observed when intraperitoneal injections were used. All A. caviae strains proved to be avirulent. Caseinases, hemolysins, and Vero cytotoxins were produced by 100, 91, and 94.59%, respectively, of A. hydrophila strains and with lower frequencies and lower caseinase activities by A. sobria isolates. No correlation was found between these activities and the degree of virulence of the strains for fish. Most hydrophobic strains seem to be concentrated in A. caviae, A. sobria, and avirulent A. hydrophila groups. Known virulence markers commonly associated with virulent strains (acriflavine negative and self-pelleting negative and precipitation after boiling positive phenotypes) had a low representation in the total of strains studied and were not associated with virulence.

Aeromonas↗
PubMed1991

Aeromonas jandaei (formerly genospecies DNA group 9 A. sobria), a new sucrose-negative species isolated from clinical specimens.

A large numerical taxonomy study conducted in 1988 of 165 mostly clinical Aeromonas strains from diverse geographic sources produced a cluster (S = 84%, SSM) of four sucrose-negative strains that included the DNA definition strain for DNA group 9 A. sobria (CDC 0787-80). These four strains, together with five additional strains received in 1989, were subjected to DNA-DNA hybridization (hydroxyapatite, 32P, 60 and 75 degrees C), and all eight strains were closely related to the ninth labeled DNA group 9 definition strain CDC 0787-80 (73 to 86% relatedness at 60 degrees C and 68 to 80% relatedness at 75 degrees C; percent divergence, 2.0 to 3.5). Type strains and DNA definition strains for all other established Aeromonas species were only 35 to 72% related (60 degrees C) to CDC 0787-80. We propose the name Aeromonas jandaei for this highly related group of nine strains, formerly known as DNA group 9 A. sobria. The type strain was designated ATCC 49568 (CDC 0787-80). The nine strains were examined at 36 degrees C and were found to be resistant to 0/129 (vibriostatic agent) and uniformly positive for oxidase, gas production from glucose, indole, lysine decarboxylase, arginine dihydrolase, o-nitrophenyl-beta-D-galactopyranoside, motility (25 degrees C), nitrate reduction, citrate utilization, hemolysis on sheep blood agar, and growth in Trypticase soy broth with no added NaCl. They all fermented D-glucose, D-mannitol, and mannose but did not ferment sucrose, cellobiose, L-arabinose, inositol, salicin, or D-sorbitol. They were uniformly negative for esculin and urea hydrolysis, elastase production, ornithine decarboxylation, and the string test. The antibiogram of A. jandaei resembled that of other aeromonads (resistance to ampicillin and cephalothin), but it differed from most other aeromonads because of resistance to single dilution of colistin and differed from clinical A. veronii biogroup sorbria (formerly A. sobria) by its nearly uniform resistance to cephalothin. The esculin-, sucrose-, and cellobiose-negative and colistin-resistant profile distinguished A. jandaei from other Aeromonas species. These A. jandaei strains were isolated from blood (two strains), wounds (two strains), diarrheal stools (four strains), and a prawn (one strain). The blood and wound isolates, in particular, suggest that there is a possible clinical significance for this species and justify identification of and further research on this group of motile aeromonads.

Aeromonas↗
PubMed1998

Misidentification of unusual Aeromonas species as members of the genus Vibrio: a continuing problem.

Two unusual cases of Aeromonas infection are described, one associated with bacteremia (Aeromonas schubertii) and another in which the organism was recovered from an infected gall bladder (Aeromonas veronii biotype veronii). These strains were initially identified as Vibrio damsela and Vibrio cholerae by the Vitek and API 20E systems, respectively. Use of appropriate screening tests and familiarity with the newer Aeromonas species could prevent initial misidentifications and potential public health consequences.

Aeromonas↗
PubMed2004

Type III secretion system genes in clinical Aeromonas isolates.

We have identified the genes ascF and ascG, which encode components of a putative type III secretion system (TTSS) in AEROMONAS: We investigated the distribution of these and other TTSS genes in 84 clinical isolates and found hybridizing sequences in 50% of the strains, with a higher prevalence in Aeromonas hydrophila and Aeromonas veronii than in Aeromonas caviae.

Aeromonas↗