Molecular cloning and expression in Escherichia coli of the extracellular endoprotease of Aeromonas caviae T-64, a pro-aminopeptidase processing enzyme(1).
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PA protease (pro-aminopeptidase processing protease) activates the pro-aminopeptidases from Aeromonas caviae T-64 and Vibrio proteolytica by removal of their pro-regions. Cloning and sequencing of the PA protease gene revealed that PA protease was translated as a preproprotein consisting of four domains: a signal peptide; an N-terminal propeptide; a mature region; and a C-terminal propeptide. The deduced amino acid sequence of the PA protease precursor showed significant homology with several bacterial metalloproteases. Expression of the PA protease gene in Escherichia coli indicated that the N-terminal propeptide of the PA protease precursor is essential to obtain the active form of the protease. The N- and C-terminal propeptides of the expressed pro-PA protease were processed autocatalytically. (+info)
Diverse restriction fragment length polymorphism patterns of the PCR-amplified 16S rRNA genes in Aeromonas veronii strains and possible misidentification of Aeromonas species.
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Restriction fragment length polymorphism analysis after PCR amplification (RFLP-PCR) of the 16S rRNA gene has been previously proposed as a rapid method to identify Aeromonas species. In the present study, the precision of RFLP-PCR was evaluated with 62 Aeromonas reference strains. The analysis revealed that Aeromonas veronii biovar sobria strains produce various patterns, possibly leading to its misidentification as an environmental species. For most other Aeromonas species little variation was noted. This study supports the usefulness of RFLP-PCR analysis to separate three clinically important species but also reveals possible misidentifications that necessitate further biochemical tests to validate the preliminary identification. (+info)
Inducible oxacillin-hydrolyzing penicillinase in Aeromonas hydrophila isolated from fish.
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An inducible penicillinase was shown to be present in a strain of Aeromonas hydrophila subsp. hydrophila isolated from freshwater fish. Enzyme induction was observed with benzylpenicillin or 6-aminopenicillanic acid, and the enzyme was cell bound. The penicillinase was purified 50-fold from a crude cell extract. The molecular weight was estimated to be 23,000 by gel filtration. The pH and temperature optima for the enzyme activity were 8.0 and 35 degrees C, respectively. The penicillinase showed a unique substrate profile by hydrolyzing oxacillin about twice as rapidly as benzylpenicillin. The enzyme activity was weakly inhibited by sodium chloride but was not affected by p-chloromercuribenzoate. The property of penicillinase production by the A. hydrophila strain could not be transferred to Escherichia coli and also could not be eliminated from the bacteria by ethidium bromide treatment. (+info)
Functional characterization of type IV pili expressed on diarrhea-associated isolates of Aeromonas species.
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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. (+info)
XynX, a possible exo-xylanase of Aeromonas caviae ME-1 that produces exclusively xylobiose and xylotetraose from xylan.
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A gene, xynX, encoding a novel xylanase, was cloned from Aeromonas caviae ME-1. This gene encoded an enzyme that was constituted of 334 amino acid residues (38,580 Da) and was similar in sequence to Family 10 (Family F) beta-1,4 endo-xylanases. XynX produced only xylobiose and xylotetraose from birch wood xylan, and xylotriose, xylopentaose, and higher oligosaccharides were not detected in the TLC analysis. We designated it as X2/X4-forming xylanase. This enzyme does not have transglycosylation activity. These data suggested that this enzyme is a possible exo-xylanase. According to homology modeling, the enzyme has a ring-shaped (alpha/beta)8 barrel (TIM barrel) structure, typical of Family 10 endo-xylanases, with the extraordinary feature of a longer bottom-loop structure. (+info)
Flagellate predation on a bacterial model community: interplay of size-selective grazing, specific bacterial cell size, and bacterial community composition.
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The influence of grazing by the bacterivorous nanoflagellate Ochromonas sp. strain DS on the taxonomic and morphological structures of a complex bacterial community was studied in one-stage chemostat experiments. A bacterial community, consisting of at least 30 different strains, was fed with a complex carbon source under conditions of low growth rate (0.5 day(-1) when nongrazed) and low substrate concentration (9 mg liter(-1)). Before and after the introduction of the predator, the bacterial community composition was studied by in situ techniques (immunofluorescence microscopy and fluorescent in situ hybridization), as well as by cultivation on agar media. The cell sizes of nonspecifically stained and immunofluorescently labeled bacteria were measured by image analysis. Grazing by the flagellate caused a bidirectional change in the morphological structure of the community. Medium-size bacterial cells, which dominated the nongrazed community, were largely replaced by smaller cells, as well as by cells contained in large multicellular flocs. Cell morphological changes were combined with community taxonomic changes. After introduction of the flagellate, the dominating strains with medium-size cells were largely replaced by single-celled strains with smaller cells on the one hand and, on the other hand, by Pseudomonas sp. strain MWH1, which formed the large, floc-like forms. We assume that size-selective grazing was the major force controlling both the morphological and the taxonomic structures of the model community. (+info)
Phylogenetic positions of Aeromonas encheleia, Aeromonas popoffii, Aeromonas DNA hybridization group 11 and Aeromonas group 501.
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The 16S rDNA sequences of the recently described Aeromonas encheleia and Aeromonas popoffii, were determined and compared with data from all known Aeromonas sp. Diagnostic 16S rDNA regions were also sequenced for some strains previously considered as an extension of A. encheleia and a strain of Aeromonas Group 501 (formerly Enteric Group 501). Results indicated that A. encheleia and A. popoffii are phylogenetically separated species as originally described. A conclusion about HG11 taxonomic status is not recommended until previous discrepancies are clarified by further DNA-DNA hybridization and sequencing studies. (+info)
Carnobacterium inhibens sp. nov., isolated from the intestine of Atlantic salmon (Salmo salar).
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Strain K1T, isolated from the gastrointestinal tract of Atlantic salmon (Salmo salar), has the capacity to inhibit the growth of the fish pathogens Vibrio anguillarum and Aeromonas salmonicida. Strain K1T is a motile Gram-positive psychrophilic rod that lacks both catalase and oxidase, which does not grow on acetate containing media, but grows at pH 9 and in TSB with up to 6% sodium chloride content. Strain K1T is facultatively anaerobic and tryptone as a sole source of nutrient promotes growth. The most abundant cellular fatty acid of strain K1T is oleic acid (18:1cis9). Based on 16S rDNA sequence comparisons, it is suggested that strain K1T is phylogenetically closely related to C. alterfunditum. However, the unique phenotypic attributes of strain K1T suggest that it represents a new species. The name Carnobacterium inhibens is proposed, for which the type strain is K1T (= CCUG 31728T). (+info)