Distribution of Aeromonas hydrophila in natural and man-made thermal effluents.

Densities of Aeromonas hydrophila showed distinct thermal optima (25 to 35 degrees C) and thermal maxima (45 degrees C) when measured along thermal gradients created by geothermal and nuclear reactor effluents. Survival of A. hydrophila never exceeded 48 h at temperatures of greater than 45 degrees C. Thermophilic strains could not be isolated at any site.     

Phospholipids and lipopolysaccharide of Aeromonas hydrophila.

The phospholipids and lipopolysaccharide of Aeromonas hydrophila were characterized. Phosphatidylethanolamine and phosphatidylglycerol were the major phospholipid components. The outer membrane contained more phosphatidylethanolamine and less phosphatidylglycerol than the inner membrane, and the phospholipids of the outer membrane contained a higher proportion of saturated fatty acids. Only four fatty acids (C14:0, C16:0, C16:1, and C18:1) were found in the phospholipids. The lipopolysaccharide of A. hydrophila did not contain the eight-carbon sugar 3-deoxyoctulosonic acid nor did it contain C16:0, both of which are typical constituents of the lipopolysaccharide of many other species.     

Clinical involvement of Aeromonas hydrophila.

Aeromonas hydrophila has for some time been regarded as an opportunistic pathogen in hosts with impaired local or general defence mechanisms. Infections in such individuals are generally severe. The organism is also being isolated with increasing frequency throughout the world from a variety of focal and systemic infections of varying severity in persons that are apparently immunologically normal. Most commonly it causes acute diarrheal disease by producing an enterotoxin. Thus the organism appears to have greater clinical significance that was hitherto suspected. The organism has been infrequently reported from humans in Canada, but its correct laboratory identification, together with increased awareness that it can contribute to illness, will undoubtedly lead to more reports of its isolation in Canada.     

Survival of Aeromonas hydrophila, Aeromonas caviae and Aeromonas sobria in soil

The survival of mesophilic Aeromonas spp. in soil in the presence or absence of indigenous microflora was evaluated in a laboratory study. Two cytotoxic ( Aer. hydrophila and Aer. caviae ) and one invasive ( Aer. sobria ) clinical isolate strains were selected for this study. After contamination of sterile or unsterilized soil with the three strains of Aeromonas , the number of living cells was determined over at least 5 months. For all strains the survival curves were characterized by an initial re-growth followed by a slow inactivation of bacteria, with significant differences due to the presence of indigenous microflora. The times necessary to achieve a 95% reduction of the initial population were > 140, 113 and 62 d in sterilized soil respectively for Aer. caviae, Aer. hydrophila and Aer. sobria , while the corresponding times in unsterilized soil were 42, 38 and 11 d. All strains preserved the virulence factors for the entire period of the study. These results suggest that the soil may be an important reservoir for Aeromonas spp. and, thus, may play an important role in the epidemiology of Aeromonas -associated human infections.     

RAPD analysis of Aeromonas salmonicida and Aeromonas hydrophila

The randomly amplified polymorphic DNA (RAPD) technique was used to analyse the genetic differentiation of 13 strains of Aeromonas salmonicida subsp. salmonicida , and seven strains of Aer. hydrophila. Reproducible profiles of genomic DNA fingerprints were generated by polymerase chain reaction (PCR) using a single randomly designed primer. The RAPD profiles of all the non-motile aeromonads, Aer. salmonicida subsp. salmonicida were identical. However, profiles of the motile aeromonads, Aer. hydrophila differed between isolates. These findings reveal genomic homogeneity in Aer. salmonicida subsp. salmonicida and genetic variety in Aer. hydrophila strains.     

Dynamics of Aeromonas hydrophila, Aeromonas sobria, and Aeromonas caviae in a sewage treatment pond.

The spatiotemporal dynamics of Aeromonas spp. and fecal coliforms in the sewage treatment ponds of an urban wastewater center were studied after 20 months of sampling from five stations in these ponds. Isolation and identification of 247 Aeromonas strains were undertaken over four seasons at the inflow and outflow of this pond system. The hemolytic activity of these strains was determined. The Aeromonas spp. and the fecal coliform distributions showed seasonal cycles, the amplitude of which increased at distances further from the wastewater source, so that in the last pond there was an inversion of the Aeromonas spp. cycle in comparison with that of fecal coliforms. The main patterns in these cycles occurred simultaneously at all stations, indicating control of these bacterial populations by seasonal factors (temperature, solar radiation, phytoplankton), the effects of which were different on each bacterial group. The analysis of the Aeromonas spp. population structure showed that, regardless of the season, Aeromonas caviae was the dominant species at the pond system inflow. However at the outflow the Aeromonas spp. population was dominated by A. caviae in winter, whereas Aeromonas sobria was the dominant species in the treated effluent from spring to fall. Among the Aeromonas hydrophila and A. sobria strains, 100% produced hemolysin; whereas among the A. caviae strains, 96% were nonhemolytic.     

Characterization of lactoferrin binding by Aeromonas hydrophila.

Various lactoferrin preparations (iron-saturated and iron-depleted human milk lactoferrins and bovine milk and colostrum lactoferrins) were bound by Aeromonas hydrophila. Binding was (i) reversible (65% of bound lactoferrin was displaced by unlabeled lactoferrin), (ii) specific (lactoferrin but not other iron-containing glycoproteins such as ferritin, transferrin, hemoglobin, and myoglobin inhibited binding), and (iii) significantly reduced by pepsin and neuraminidase treatment of the bacteria. The glycosidic domains of the lactoferrin molecule seem to be involved in binding since precursor monosaccharides of the lactoferrin oligosaccharides (mannose, fucose, and galactose) and glycoproteins which have homologous glycosidic moieties similar to those of the lactoferrin oligosaccharides (asialofetuin or fetuin) strongly inhibited lactoferrin binding. A. hydrophila also binds transferrin, ferritin, cytochrome c, hemin, and Congo red. However, binding of these iron-containing compounds seems to involve bacterial surface components different from those required for lactoferrin binding. Expression of lactoferrin binding by A. hydrophila was influenced by culture conditions. In addition, there was an inverse relationship between lactoferrin binding and siderophore production by the bacterium.     

Characterization of lactoferrin binding by Aeromonas hydrophila.

Various lactoferrin preparations (iron-saturated and iron-depleted human milk lactoferrins and bovine milk and colostrum lactoferrins) were bound by Aeromonas hydrophila. Binding was (i) reversible (65% of bound lactoferrin was displaced by unlabeled lactoferrin), (ii) specific (lactoferrin but not other iron-containing glycoproteins such as ferritin, transferrin, hemoglobin, and myoglobin inhibited binding), and (iii) significantly reduced by pepsin and neuraminidase treatment of the bacteria. The glycosidic domains of the lactoferrin molecule seem to be involved in binding since precursor monosaccharides of the lactoferrin oligosaccharides (mannose, fucose, and galactose) and glycoproteins which have homologous glycosidic moieties similar to those of the lactoferrin oligosaccharides (asialofetuin or fetuin) strongly inhibited lactoferrin binding. A. hydrophila also binds transferrin, ferritin, cytochrome c, hemin, and Congo red. However, binding of these iron-containing compounds seems to involve bacterial surface components different from those required for lactoferrin binding. Expression of lactoferrin binding by A. hydrophila was influenced by culture conditions. In addition, there was an inverse relationship between lactoferrin binding and siderophore production by the bacterium.     

Aeromonas hydrophila: analysis of 11 cases.

A retrospective analysis of 11 cases in which Aeromonas hydrophila was isolated indicated that the organsim caused local infection in 7 cases and asymptomatic colonization in 4. There were no cases of septicemia and none of the patients were known to have a malignant disease or immunosuppression. There were no deaths, although three of the patients required amputation of limbs because of myonecrosis. Chloramphenicol and aminoglycosides appeared to be appropriate therapeutic agents.     

Specific binding of lactoferrin to Aeromonas hydrophila.

The interaction of lactoferrin (Lf) with Aeromonas hydrophila (n = 28) was tested in a 125I-labeled protein-binding assay. The mean per cent binding values for human Lf (HLf) and bovine Lf (BLf) were 13.4 +/- 2.0 (SEM), and 17.5 +/- 2.7 (SEM), respectively. The Lf binding was characterized in type strain A. hydrophila subsp. hydrophila CCUG 14551. The HLf and BLf binding reached a complete saturation within 2 h. Unlabeled HLf and BLf displaced 125I-HLf binding in a dose-dependent manner, and more effectively by the heterologous (1 microgram for 50% inhibition) than the homologous (10 micrograms for 50% inhibition) ligand. Apo- and holo-forms of HLf and BLf both inhibited more than 80%, while mucin caused approx. 50% inhibition of the HLf binding. Various other proteins (including transferrin) or carbohydrates did not block the binding. Two HLf-binding proteins with an estimated molecular masses of 40 kDa and 30 kDa were identified in a boiled-cell-envelope preparation, while the unboiled cell envelope demonstrated a short-ladder pattern at the top of the separating gel and a second band at approx. 60 kDa position. These data establish a specific interaction of Lf and the Lf-binding proteins seem to be porins in A. hydrophila.     

Electroporation-mediated transformation of Aeromonas hydrophila

A strain of Aeromonas hydrophila producing copolyesters of 3-hydroxybutyrate and 3-hydroxyhexanoate, abbreviated as PHBHHx, was successfully transformed by electroporation. The plasmid used was a broad host range plasmid pBBR1MCS. Electroporation conditions were varied systemically to develop an electroporation protocol. The optimal yield of transformant was approximately 4x10(2) CFU/microg DNA at 12.5 kV/cm and 1000 Omega, resulting in a time constant of approximately 5 ms. The A. hydrophila transformants expressed plasmid-encoded resistance to chloromphenicol. Plasmid DNA in the A. hydrophila transformant was stably maintained. This is the first report of transformation of bacteria A. hydrophila.     

Prevalence and distribution of Aeromonas hydrophila in the United States.

The abundance of Aeromonas hydrophila was measured in 147 natural aquatic habitats in 30 states and Puerto Rico. Viable cell counts were used to estimate density at all sites by using Rimler-Shotts medium, a differential presumptive medium for A. hydrophila. Temperature, pH, conductivity, salinity, and turbidity were measured simultaneously with water sample collection. The density of A. hydrophila was higher in lotic than in lentic systems. Saline systems had higher densities of A. hydrophila than did freshwater systems. A. hydrophila could not be isolated from extremely saline, thermal, or polluted waters, even though it was found over wide ranges of salinity, conductivity, temperature, pH, and turbidity. Of the water quality parameters measured, only conductivity was significantly regressed with density of A. hydrophila.     

Polar flagellum biogenesis in Aeromonas hydrophila

Mesophilic Aeromonas spp. constitutively express a single polar flagellum that helps the bacteria move to more favorable environments and is an important virulence and colonization factor. Certain strains can also produce multiple lateral flagella in semisolid media or over surfaces. We have previously reported 16 genes (flgN to flgL) that constitute region 1 of the Aeromonas hydrophila AH-3 polar flagellum biogenesis gene clusters. We identified 39 new polar flagellum genes distributed in four noncontiguous chromosome regions (regions 2 to 5). Region 2 contained six genes (flaA to maf-1), including a modification accessory factor gene (maf-1) that has not been previously reported and is thought to be involved in glycosylation of polar flagellum filament. Region 3 contained 29 genes (fliE to orf29), most of which are involved in flagellum basal body formation and chemotaxis. Region 4 contained a single gene involved in the motor stator formation (motX), and region 5 contained the three master regulatory genes for the A. hydrophila polar flagella (flrA to flrC). Mutations in the flaH, maf-1, fliM, flhA, fliA, and flrC genes, as well as the double mutant flaA flaB, all caused loss of polar flagella and reduction in adherence and biofilm formation. A defined mutation in the pomB stator gene did not affect polar flagellum motility, in contrast to the motX mutant, which was unable to swim even though it expressed a polar flagellum. Mutations in all of these genes did not affect lateral flagellum synthesis or swarming motility, showing that both A. hydrophila flagellum systems are entirely distinct.     

Prevalence and distribution of Aeromonas hydrophila in the United States.

The abundance of Aeromonas hydrophila was measured in 147 natural aquatic habitats in 30 states and Puerto Rico. Viable cell counts were used to estimate density at all sites by using Rimler-Shotts medium, a differential presumptive medium for A. hydrophila. Temperature, pH, conductivity, salinity, and turbidity were measured simultaneously with water sample collection. The density of A. hydrophila was higher in lotic than in lentic systems. Saline systems had higher densities of A. hydrophila than did freshwater systems. A. hydrophila could not be isolated from extremely saline, thermal, or polluted waters, even though it was found over wide ranges of salinity, conductivity, temperature, pH, and turbidity. Of the water quality parameters measured, only conductivity was significantly regressed with density of A. hydrophila.     

Enumeration and characterization of Aeromonas hydrophila and Aeromonas caviae isolated from grocery store produce.

Starch-ampicillin agar was used to quantitatively isolate Aeromonas sp. from retail grocery store produce. All produce sampled, including parsley, spinach, celery, alfalfa sprouts, broccoli, and lettuce, contained Aeromonas sp. In most instances, the count of Aeromonas sp. increased 10- to 1,000-fold during 2 weeks of storage at 5 degrees C. Eleven (92%) of 12 kinds of produce yielded cytotoxic Aeromonas sp. Identification as Aeromonas hydrophila was the strongest indicator of cytotoxicity, and all 29 (100%) A. hydrophila isolates and 1 (6%) of 16 A. caviae isolates were cytotoxic. Twenty-seven (90%) of 30 cytotoxic Aeromonas sp. strains produced hemolysins. Strong correlations were also noted between ability to produce cytotoxin and positive Voges-Proskauer, lysine decarboxylase, and sorbitol fermentation reactions. It appears that grocery store produce is a potentially significant source of cytotoxic Aeromonas sp. and should be considered in the epidemiology of A. hydrophila gastroenteritis.     

Inactivation of Aeromonas hydrophila metallo-beta-lactamase by cephamycins and moxalactam.

Incubation of moxalactam and cefoxitin with the Aeromonas hydrophila metallo-beta-lactamase CphA leads to enzyme-catalyzed hydrolysis of both compounds and to irreversible inactivation of the enzyme by the reaction products. As shown by electrospray mass spectrometry, the inactivation of CphA by cefoxitin and moxalactam is accompanied by the formation of stable adducts with mass increases of 445 and 111 Da, respectively. The single thiol group of the inactivated enzyme is no longer titrable, and dithiothreitol treatment of the complexes partially restores the catalytic activity. The mechanism of inactivation by moxalactam was studied in detail. Hydrolysis of moxalactam is followed by elimination of the 3' leaving group (5-mercapto-1-methyltetrazole), which forms a disulfide bond with the cysteine residue of CphA located in the active site. Interestingly, this reaction is catalyzed by cacodylate.     

Characterization of an O-antigen bacteriophage from Aeromonas hydrophila.

A unique bacteriophage of Aeromonas hydrophila serotype O:34 was isolated, purified, and characterized. The bacterial surface receptor was shown to be the O-antigen polysaccharide component of lipopolysaccharide specific to serotype O:34, which was chemically characterized. The high molecular weight lipopolysaccharide fraction (a fraction enriched in O antigen) was fully able to inactivate bacteriophage PM1. Phage-resistant mutants of A. hydrophila O:34 were isolated and found to be specifically devoid of lipopolysaccharide O antigen. No other cell-surface molecules were involved in phage binding. The host range of bacteriophage PM1 was found to be very narrow, producing plaques only on A. hydrophila strains from serotype O:34.     

Survival of genetically-marked Aeromonas hydrophila in water

Survival of a genetically-marked Aeromonas hydrophila was monitored in water microcosms. There was no apparent loss of a marker plasmid which encoded the xylE reporter gene during prolonged incubation in lake water. Survival was best in sterile lake water but in sea water, cells died rapidly during the first 9 d, recovered up to day 12 and thereafter numbers fell up to 28 d accompanied by loss of the plasmid in a proportion of the cells.     

Dynamics of Aeromonas hydrophila, Aeromonas sobria, and Aeromonas caviae in a sewage treatment pond

The spatiotemporal dynamics of Aeromonas spp. and fecal coliforms in the sewage treatment ponds of an urban wastewater center were studied after 20 months of sampling from five stations in these ponds. Isolation and identification of 247 Aeromonas strains were undertaken over four seasons at the inflow and outflow of this pond system. The hemolytic activity of these strains was determined. The Aeromonas spp. and the fecal coliform distributions showed seasonal cycles, the amplitude of which increased at distances further from the wastewater source, so that in the last pond there was an inversion of the Aeromonas spp. cycle in comparison with that of fecal coliforms. The main patterns in these cycles occurred simultaneously at all stations, indicating control of these bacterial populations by seasonal factors (temperature, solar radiation, phytoplankton), the effects of which were different on each bacterial group. The analysis of the Aeromonas spp. population structure showed that, regardless of the season, Aeromonas caviae was the dominant species at the pond system inflow. However at the outflow the Aeromonas spp. population was dominated by A. caviae in winter, whereas Aeromonas sobria was the dominant species in the treated effluent from spring to fall. Among the Aeromonas hydrophila and A. sobria strains, 100% produced hemolysin; whereas among the A. caviae strains, 96% were nonhemolytic.