LPS Profiles Research Articles

Changes in physiology and virulence during the selection of resistant Yersinia ruckeri mutants under subinhibitory cefotaxime concentrations.

Bacterial antibiotic resistance is one of the main healthcare problems currently. Apart from reducing antibiotic efficacy, it has awakened the interest of scientists due to its association with bacterial fitness and virulence. Interestingly, antibiotic resistance can be a source of both increased fitness and decreased fitness, even though the molecular basis of these relationships remains unknown. The aim of this work is to define the effects of sub-MIC concentrations of cefotaxime, an antibiotic extensively used in clinical practice, on the physiology and virulence of Yersinia ruckeri and to determine the importance of these sub-MIC concentrations for the selection of antibiotic-resistant mutants in the aquatic environment. Results indicated that exposure to sub-MIC concentrations of cefotaxime selected Y.ruckeri populations with irreversible alterations in the physiology, such as slow growth, aggregation in liquid cultures and modification of the colony morphology. These bacteria also displayed changes in the OMPs and LPS profiles and a full attenuation of virulence. An overexpression of the envelope stress regulator RpoE was also detected after exposure to the antibiotic. In conclusion, exposure to cefotaxime selected, at high frequency, Y.ruckeri strains that survive the antibiotic stress at the expense of a fitness cost and the loss of virulence.

Acid tolerance response of Bordetella bronchiseptica in avirulent phase

Bordetella bronchiseptica is a Gram-negative bacterium responsible for respiratory diseases in many mammalian hosts, including humans. This pathogen has been shown as able to persist inside the host cells, even in the phagosomes that are acidified to pH 4.5–5.0 after bacterial infection. Here we evaluated the resistance of B. bronchiseptica to survive under acidic conditions. In particular we analyzed the bacterial capacity to develop the mechanism known as acid tolerance response (ATR). Our studies were mainly focused on the avirulent phase of the bacteria since this phenotypic phase was reported to be more resistant to environmental stress conditions than the virulent phase. Results from B. bronchiseptica in virulent phase were also included for comparison purposes. In fact, for B. bronchiseptica 9.73 bacteria in virulent phase we observed that the viability of bacteria does not decrease significantly when grown at pH as low as 4.5, but it is affected when the pH of the medium was equal to or less than 4.0. After acid-adaptation at pH 5.5 for several hours, the survival rate of B. bronchiseptica 9.73 at lethal pH 4.0 for 6h was increased. Interestingly, the avirulent phase mediated by the two-component BvgAS system conferred further resistance to lethal acid challenge and a marked increase in the magnitude of the expressed ATR. The ATR for this avirulent phase seems to be associated with changes in LPS and surface protein profiles. 2D-gel electrophoresis revealed at least 25 polypeptides differentially expressed, 17 of which were only expressed or over-expressed under acid conditions. Using MALDI-TOF mass spectrometry, 10 of these differentially expressed polypeptides were identified.

Mutational analysis of the major periplasmic loops of Shigella flexneri Wzy: identification of the residues affecting O antigen modal chain length control, and Wzz-dependent polymerization activity.

The O antigen (Oag) component of LPS is a major Shigella flexneri virulence determinant. Oag is polymerized by WzySf, and its modal chain length is determined by WzzSf and WzzpHS2. Site-directed mutagenesis was performed on wzySf in pWaldo-wzySf-TEV-GFP to alter Arg residues in WzySf's two large periplasmic loops (PLs) (PL3 and PL5). Analysis of the LPS profiles conferred by mutated WzySf proteins in the wzySf deficient (Δwzy) strain identified residues that affect WzySf activity. The importance of the guanidium group of the Arg residues was investigated by altering the Arg residues to Lys and Glu, which generated WzySf mutants conferring altered LPS Oag modal chain lengths. The dependence of these WzySf mutants on WzzSf was investigated by expressing them in a wzySf and wzzSf deficient (Δwzy Δwzz) strain. Comparison of the LPS profiles identified a role for the Arg residues in the association of WzySf and WzzSf during Oag polymerization. Colicin E2 and bacteriophage Sf6c susceptibility supported this conclusion. Comparison of the expression levels of different mutant WzySf-GFPs with the wild-type WzySf-GFP showed that certain Arg residues affected production levels of WzySf in a WzzSf-dependent manner. To our knowledge, this is the first report of S. flexneri WzySf mutants having an effect on LPS Oag modal chain length, and identified functionally significant Arg residues in WzySf.

The Sinorhizobium fredii HH103 Lipopolysaccharide Is Not Only Relevant at Early Soybean Nodulation Stages but Also for Symbiosome Stability in Mature Nodules

In this work we have characterised the Sinorhizobium fredii HH103 greA lpsB lpsCDE genetic region and analysed for the first time the symbiotic performance of Sinorhizobium fredii lps mutants on soybean. The organization of the S. fredii HH103 greA, lpsB, and lpsCDE genes was equal to that of Sinorhizobium meliloti 1021. S. fredii HH103 greA, lpsB, and lpsE mutant derivatives produced altered LPS profiles that were characteristic of the gene mutated. In addition, S. fredii HH103 greA mutants showed a reduction in bacterial mobility and an increase of auto-agglutination in liquid cultures. RT-PCR and qPCR experiments demonstrated that the HH103 greA gene has a positive effect on the transcription of lpsB. Soybean plants inoculated with HH103 greA, lpsB or lpsE mutants formed numerous ineffective pseudonodules and showed severe symptoms of nitrogen starvation. However, HH103 greA and lps mutants were also able to induce the formation of a reduced number of soybean nodules of normal external morphology, allowing the possibility of studying the importance of bacterial LPS in later stages of the S. fredii HH103-soybean symbiosis. The infected cells of these nodules showed signs of early termination of symbiosis and lytical clearance of bacteroids. These cells also had very thick walls and accumulation of phenolic-like compounds, pointing to induced defense reactions. Our results show the importance of bacterial LPS in later stages of the S. fredii HH103-soybean symbiosis and their role in preventing host cell defense reactions. S. fredii HH103 lpsB mutants also showed reduced nodulation with Vigna unguiculata, although the symbiotic impairment was less pronounced than in soybean.

Colonization of gerbils withHelicobacter pyloriO-chain-deficient mutant SS1 HP0826::Kan results in gastritis and is associated withde novosynthesis of extended homopolymers

The O-chain polysaccharide of Helicobacter pylori is important for colonization and generation of chronic gastritis in mice. There are marked host differences in the development of H. pylori-induced gastric pathology in mice and gerbils. To investigate the role of the O-chain polysaccharide of H. pylori in colonization and gastritis in Mongolian gerbils, inoculation by oral gavage with H. pylori strain SS1 and its corresponding O-chain polysaccharide-deficient mutant SS1 HP0826::Kan was undertaken. Infection with both strains resulted in corpus atrophy, loss of parietal cells, and extensive mucous metaplasia at both 18 and 30 weeks postinfection. Contrary to previous results in splenocyte recipient severe combined immunodeficiency (SCID) mice, no difference was found in the grade of chronic gastritis, polymorphonuclear cell infiltration, atrophy, and mucous metaplasia in gerbils infected with the wild-type SS1 strain or SS1 HP0826::Kan strain. Examination of the effects of gerbil passage on LPS profiles of output SS1 HP0826::Kan isolates by SDS-PAGE, sugar, and methylation analyses revealed significant differences in LPS profiles of SS1 HP0826::Kan cells recovered from infected gerbils as compared to input bacteria. Specifically, the presence of a novel homopolymer of d-galactan, as well as an extended polymer of d-riban, was detected. These data provide evidence for the role of H. pylori LPS in bacterial adaption to promote colonization and pathology.

Helicobacter pylori isolates from Greek children express type 2 and type 1 Lewis and α1,6-glucan antigens in conjunction with a functional type IV secretion system

Helicobacter pylori infection is often acquired in childhood and can persist for life. Previous studies in adult patients have shown that H. pylori isolates from North American and European hosts express predominantly type 2 Lewis x (Le(x)) and Le(y) epitopes, while Asian strains have the capacity to express type 1 Le(a) and Le(b) structures. In order to understand the influence of environmental and host factors on the expression of Le antigens, we analysed 50 Greek H. pylori isolates from symptomatic children. Both CagA-positive and -negative strains were evaluated. The expression of Le antigens was determined by whole-cell indirect ELISA (WCE), and LPS profiles were assessed by gel electrophoresis and immunoblotting. Occurrence of Le(x) and/or Le(y) antigens was confirmed in 35 of the isolates (70 %) while 15 of the isolates were non-typable. It was found that 11 of the paediatric isolates had the propensity to express type 1 Le(b) blood-group antigen (22 %), a feature relatively uncommon in H. pylori isolates from adults. One strain expressed both Le(b) and Le(a) antigens. The majority of the isolates (49/50, 98 %) expressed α1,6-glucan, an antigenic non-Le determinant present in the outer core region of H. pylori LPS. All Le(x)- and Le(y)-expressing strains also carried a functional cag pathogenicity island-encoding a type IV secretion system, capable of translocating CagA protein, as well as the vacAs1 allele, suggesting that Le(x) and Le(y) epitopes may aid the persistence of more aggressive strains. No association between bacterial virulence characteristics and the histopathological observations was evident.

Growing Burkholderia pseudomallei in Biofilm Stimulating Conditions Significantly Induces Antimicrobial Resistance

Background Burkholderia pseudomallei, a Gram-negative bacterium that causes melioidosis, was reported to produce biofilm. As the disease causes high relapse rate when compared to other bacterial infections, it therefore might be due to the reactivation of the biofilm forming bacteria which also provided resistance to antimicrobial agents. However, the mechanism on how biofilm can provide tolerance to antimicrobials is still unclear.Methodology/Principal FindingsThe change in resistance of B. pseudomallei to doxycycline, ceftazidime, imipenem, and trimethoprim/sulfamethoxazole during biofilm formation were measured as minimum biofilm elimination concentration (MBEC) in 50 soil and clinical isolates and also in capsule, flagellin, LPS and biofilm mutants. Almost all planktonic isolates were susceptible to all agents studied. In contrast, when they were grown in the condition that induced biofilm formation, they were markedly resistant to all antimicrobial agents even though the amount of biofilm production was not the same. The capsule and O-side chains of LPS mutants had no effect on biofilm formation whereas the flagellin-defective mutant markedly reduced in biofilm production. No alteration of LPS profiles was observed when susceptible form was changed to resistance. The higher amount of N-acyl homoserine lactones (AHLs) was detected in the high biofilm-producing isolates. Interestingly, the biofilm mutant which produced a very low amount of biofilm and was sensitive to antimicrobial agents significantly resisted those agents when grown in biofilm inducing condition.Conclusions/SignificanceThe possible drug resistance mechanism of biofilm mutants and other isolates is not by having biofilm but rather from some factors that up-regulated when biofilm formation genes were stimulated. The understanding of genes related to this situation may lead us to prevent B. pseudomallei biofilms leading to the relapse of melioidosis.

Analysis of Helicobacter pylori isolates from Chile: occurrence of selective type 1 Lewis b antigen expression in lipopolysaccharide

Previous studies have shown that the LPS of Helicobacter pylori isolated from North American and European hosts predominantly expresses type 2 Lewis x (Le(x)) and Le(y) epitopes, whilst the LPS from Asian strains has the capacity to express type 1 Le(a) and Le(b) structures. The aim of this study was to evaluate the expression of Le antigens and the cytotoxin-associated antigen (CagA) by H. pylori isolates from Chile. A total of 38 isolates were screened. The expression of Le antigens and CagA was determined by whole-cell indirect ELISA, using commercially available monoclonal anti-Le and polyclonal anti-CagA antibodies. LPS profiles of H. pylori isolates were assessed by gel electrophoresis and Western blotting. Expression of Le(x) and/or Le(y) epitopes was confirmed in 32/38 isolates (84 %), whilst 9/38 isolates (24 %) expressed type 1 Le(b) blood group determinants, in addition to type 2 Le(x) and Le(y) structures. Six strains (16 %) were non-typeable. The majority of H. pylori strains examined were CagA-positive (83.3 %).

Diversity in a promiscuous group of rhizobia from three Sesbania spp. colonizing ecologically distinct habitats of the semi-arid Delhi region

Sesbania-rhizobia associations have immense significance in soil amelioration programs for diverse habitats. Diversity in symbiotic properties, LPS profiles, Sym plasmid and rhizobiophage sensitivity of 28 root- and stem-nodulating bacterial isolates of three Sesbania species ( S. sesban, S. aegyptica and S. rostrata) inhabiting six ecologically distinct sites of semi-arid Delhi region was analyzed. The isolates were highly promiscuous among the symbiotic partners ( Sesbania spp.). The root nodules formed by all the isolates were morphologically similar but they differed in their symbiotic efficiency and effectiveness. 16S rDNA sequence analyses revealed that root nodule isolates of sesbanias belong to diverse rhizobial taxa ( Sinorhizobium saheli, S. meliloti, Rhizobium huautlense) whereas stem-nodule isolates were strictly Azorhizobium caulinodans. Sinorhizobium spp. seem to dominate as microsymbiont partner of Sesbania in the Delhi region. The genetic diversity revealed by cluster analyses based on NPC-PCR reflects sorting of isolates across the ecological gradient. Parallel diversity was also observed in the grouping based on LPS profiles and sym plasmid (NPC-PCR). Segregation of different rhizobial taxa into distinct types/clusters based on LPS and NPC-PCR analyses suggest its significance in the circumscription of the taxa. However, subtypes and subclusters showed their sorting across the ecological gradients. Sesbania rhizobia showed extremely high specificity to rhizobiophages. Enormous diversity in LPS profiles and high specificity of rhizobiophages might be the result of environmental selection pressures operating in ecologically distinct habitats. The ability of sesbanias to enter into effective symbioses with different rhizobial taxa and colonize diverse habitats with various biotic and abiotic stresses appears to contribute to its wide ecological amplitude.

Morphological changes during dendritic cell maturation correlate with cofilin activation and translocation to the cell membrane.

Upon activation, tissue residing immature dendritic cells (DC) start to migrate towards the draining lymph node and mature into efficient antigen-presenting cells. During maturation DC loose their capacity to endocytose antigens, change their surface expression of adhesion molecules, chemokine receptors, and costimulatory molecules, and change morphology. We employed 2D-PAGE and mass spectrometry to identify additional differentially expressed proteins in immature and mature DC. Human monocyte-derived DC were matured with LPS and protein expression profiles were compared before and after maturation. One of the proteins differentially expressed between immature and mature DC was identified as the actin-binding protein cofilin. We show here that cofilin is dephosphorylated in response to several maturation stimuli (i.e. CD40 ligand, LPS or a combination of TNF-alpha and prostaglandin E2). Moreover, dephosphorylated cofilin translocated towards the plasma membrane during maturation. Importantly, this correlated with an increase in filamentous actin and the appearance of veils, suggesting a role for cofilin in cytoskeletal rearrangements during maturation.

Effects of plasmid curing on antibiotic susceptibility, phage type, lipopoly saccharide and outer membrane protein profiles in local Salmonella isolates

Five different Salmonella isolates (four Salmonella enteritidis and one Salmonella havana) of human, chicken or egg origin which all expressed smooth lipopolysaccharide were subjected to plasmid curing. Upon this treatment, four isolates lost at least one of their resident plasmids while all five isolates became rough in their LPS profiles. Two of the S. enteritidis isolates, CA13 and CE7, were found to convert phage types (PTs) as well. The phage type conversion in CA13 from PT6 to PT21 was found to be accompanied by the loss of a 19·0-MDa plasmid as well as the loss of sulbactam ampicillin and tetracycline resistance. The change in the PTs of the strain CE7 from PT20a to the PT20 variant was accompanied by the loss of two plasmids of 10·9 and 1·45 MDa. Outer membrane protein (OMP) type change occurred only in the isolate CA13, which lost an OMP of 27·2 kDa. The strains, CA13 and CE7, retained their high molecular weight plasmid (36 MDa), which has been very commonly found (77 of 82) among local S. enteritidis isolates.

Extended serotyping scheme for Vibrio anguillarum with the definition and characterization of seven provisional O-serogroups.

The present paper summarizes the serotyping scheme of the fish pathogenic bacterium Vibrio anguillarum and defines seven additional O-serogroups. Strains, collected in our laboratories that were nontypable with antisera against the previously defined 16 O-serotypes, were used for generating new antisera and were characterized further by means of LPS profiles, Western blots, and serological reactions. On the basis of the results, it is suggested that the seven new O-serogroups are to be included in the existing serotyping system as serotypes O17-O23. However, the existence of further V. anguillarum strains that were not typable with any of the 23 O-antisera suggested the existence of additional O-serotypes within this species. The relevance of the description of additional O-serotypes for the species V. anguillarum is discussed.

Acid-induced expression of an LPS-associated gene in Helicobacter pylori.

To investigate urease-independent mechanisms by which Helicobacter pylori resists acid stress, subtractive RNA hybridization was used to identify H. pylori genes whose expression is induced after exposure to acid pH. This approach led to the isolation of a gene that encoded a predicted 34.8kDa protein (WbcJ), which was homologous to known bacterial O-antigen biosynthesis proteins involved in the conversion of GDP-mannose to GDP-fucose. An isogenic wbcJ null mutant strain failed to express O-antigen and Lewis X or Lewis Y determinants and was more sensitive to acid stress than was the wild-type strain. Qualitative differences in LPS profiles were observed in H. pylori cells grown at pH 5 compared with pH 7, which suggests that H. pylori may alter its LPS structure in response to acidic pH. This may be an important adaptation facilitating H. pylori colonization of the acidic gastric environment.

Dual silver staining to characterise Helicobacterspp. outer membrane components

Helicobacter pylori is a bacterial pathogen, estimated to infect half the world's population. The bacterium is the aetiological cause of gastritis, the common precursor for peptic ulcer disease and gastric cancer. Immunisation of at-risk individuals is the most cost-effective means of dealing with such a widespread pathogen. Potential vaccine candidates need to be identified and characterised. Conventional silver staining is commonly used for the sensitive detection of bacterial protein components separated by SDS-PAGE. Modified silver stains employing periodate oxidation have also been developed for the analysis of purified bacterial lipopolysaccharide. By using these methods in parallel, as a dual silver stain, bacterial fractions can be characterised in terms of protein and LPS content. Strain differences can also be readily identified by comparing protein and LPS profiles. When combined with differential immunoblotting, the dual silver stain is a useful analytical tool for characterising potential vaccine candidate antigens.

A Comparison of Methods for the Typing of Fish-Pathogenic Vibrio spp.

Summary The validity and distinctiveness of Vibrio anguillarum (= Listonella anguillara), V. (= Photobacterium) damsela, V. ordalii and V. salmonicida was confirmed. However, strains received as V. cholerae and V. splendidus were heterogeneous. Ribotyping, phenotypic (BIOLOG-GN fingerprints and API 20E profiles), chemotaxonomic (lipopolysaccharide [LPS] and outer membrane proteins [OMP]), serogrouping and plasmid profiling data were not always congruent. V. anguillarum isolates were recovered in a single ribotype cluster, but many serogroups. There was little variation in OMP profiles, but not so for LPS profiles and plasmid composition. Heterogeneity was recorded in the phenotypic characters, particularly with the API 20E rapid identification system. V. damsela displayed heterogeneity by ribotyping, but homogeneity by BIOLOG-GN fingerprints and API 20E profiles. Four serogroups were defined, but only one LPS profile was recognised. V. ordalii was homogeneous by ribotyping, serogrouping and plasmid profiling, was accommodated in two LPS groups, but was more heterogeneous by BIOLOG-GN and API 20E. Despite its more exacting cultural requirements, V. salmonicida autoagglutinated and could not be serogrouped, but was accommodated in a single LPS group showing a profile associated with rough strains, and contained plasmids of 4.7 and 42 kb. Heterogeneity was recorded with the API 20E rapid identification system.

Role of LPS length in clearance rate of bacteria from the bloodstream in mice.

Strains of Pseudomonas aeruginosa isolated from patients with cystic fibrosis (CF) never spread systemically. This may be due to serum sensitivity since these strains are very sensitive to complement-mediated bactericidal activity. A serum-resistant mutant, P. aeruginosa TUM3 HSR, was obtained from serum-sensitive strain TUM3 from a CF patient in order to clarify the mechanism of failure of systemic spread. LPS profiles on silver-stained gels and immunological analysis revealed that a long O-polysaccharide side chain was overproduced on the LPS molecules of TUM3 HSR as compared with the LPS of TUM3. The clearance rate from the bloodstream in mice was compared in the two strains. The number of TUM3 bacteria in 1 ml of blood, 10 min after injection into the tail vein, significantly decreased from 1.7 x 10(8) to 3.7 x 10(5) c.f.u. ml-1. In contrast, TUM3 HSR was not eliminated during the same period (decrease from 1.9 x 10(8) to 3.4 x 10(7) c.f.u. ml-1). Interestingly, these isogenic strains were not killed by 40% murine serum, probably reflecting immaturity of the complement-mediated killing system in mice. These results pointed to a correlation between LPS structure and blood clearance rate in mice. This was confirmed by examining blood clearance kinetics using the smooth-LPS strain Salmonella typhimurium LT2 and LPS-deficient mutants derived from it. S. typhimurium LT2 resisted blood clearance while the LPS-deficient mutants were cleared rapidly. None of the S. typhimurium strains were killed by murine serum. The number of P. aeruginosa TUM3 and S. typhimurium LPS-deficient mutants trapped in the liver following injection into the peripheral circulation was greater than that of their counterparts.(ABSTRACT TRUNCATED AT 250 WORDS)

Characterization of Vibrio anguillarum Strains Isolated from Diseased Fish in Finland

Characterization of V. anguillarum strains (n = 109) isolated from diseased salmonids was performed. Eight O serovars were found among the strains. Serovar O1 was predominant (90%), while serovars O2, O3, O5, O8, O9, and a new serovar Va NT2, were represented by 1 or 2 strains. Two strains remained non-typeable. One of these was cross-reactive with several antisera, but had a LPS profile identical to that of serovar O8. All serovars showed specific LPS profiles. All but 1 of the O1 strains had a plasmid comparable in size to the pJM1 virulence plasmid, while plasmids of different sizes were found in O2, Va NT2 and the non-typeable strains. Apart from a single strain resistant to tetracycline, all the strains were sensitive to oxolinic acid, tetracycline, and trimethoprim-sulfonamides. By their biochemical and antigenic properties strains causing vibriosis among salmonids in Finland closely resemble Scandinavian strains. Predominance of the serovars O1 and O2 suggests that commercial vaccines containing these serovars should afford sufficient protection against vibriosis in Finland.

O-serogrouping and surface components of Aeromonas hydrophila and Aeromonas jandaei pathogenic for eels.

The relationship between virulence, O-serogroup, and some cell-surface features (self-pelleting [SP] and precipitation after boiling [PAB], profile of lipopolysaccharides [LPSs] and outer membrane proteins [OMPs]) was investigated in strains of the pathogenic species Aeromonas hydrophila and A. jandaei isolated from eels. Virulent strains of A. hydrophila reacted mostly with O:19 antiserum, and those of A. jandaei reacted with O:4, O:11, O:15 and O:29 antisera (Guinée and Jansen system). Regarding the PAB and LPS profiles two groups could be distinguished; (i) five PAB+ strains of serotype O:19 that possessed a homogeneous O polysaccharide side chain and (ii) thirteen PAB- strains antigenically diverse that either exhibited a heterogeneous side chain or were side chain deficient. A major 50 kDa protein was only found in the PAB+ strains, whereas major OMPs detected in PAB- strains ranged from 33 to 45 kDa irrespective of the species. Epizootic eel isolates of A. hydrophila belong to serotype O:19 and share cell-surface features with the Aeromonas highly virulent for other hosts. In contrast, epizootic A. jandaei isolates were antigenically diverse. These findings reinforce the importance of an O-serotype as an epidemiological marker in motile Aeromonas strains pathogenic for eels.

Antigenic and Molecular Characterization of Yersinia ruckeri Proposal for a New Intraspecies Classification

The serological and molecular properties of a group of 17 Yersinia ruckeri strains isolated from cultured rainbow trout in Spain were compared with a total of 36 collection strains isolated from fish in other geographic areas. The serological relationship among isolates, as well as the study of their antigenic determinants (LPS and membrane proteins) allow to propose a new serotyping scheme with four O-serotypes (from O1 to O4). Moreover, serotypes O1 and O2 were divided in two and three groups respectively based mainly on minor differences observed in LPS and membrane protein profiles. The patterns obtained in the analysis of the LPS and proteins present in the extracellular products also support the serological scheme proposed. The strains showed a great genetic homogeneity, displaying similar DNA fingerprint patterns regardless of their serotype. In contrast, only the serotype O1 isolates harbour a plasmid of about 50 MDa, while the strains of the other serotypes presented no extrachromosomical DNA or only small plasmids (less than 10 MDa). © 1993, Gustav Fischer Verlag, Stuttgart · New York. All rights reserved.

Vibrio mimicus and Vibrio cholerae non-01 Isolated from Wild and Hatchery-Reared Fish.

In this work, we conducted a phenotypic and serological characterization of a group of Vibrio mimicus and Vibrio cholerae non-01 strains isolated from different fish species in the Chesapeake Bay area (Maryland, USA) in comparison with reference strains. Pathogenicity for fish and mice as well as the biological activities present in their extracellular products (ECP) were also evaluated. The taxonomic analysis revealed that all the strains possessed similar physiological and biochemical profiles with the exception of sucrose fermentation, which is the main differential trait between these two Vibrio species.The isolates showed drug resistance patterns very similar, being all of them sensitive to the majority of the chemotherapeutic agents employed. The serological analysis revealed an antigenic diversity among the strains which was confirmed by the LPS profiles. However, although differences were observed in the profiles of total and outer membrane proteins, the western blot analysis showed that most of these Vibrio isolates shared several immunoreactive bands in the region of 60-70 kDa. Virulence assays demonstrated that although the V. cholerae non-01 and V. mimicus isolates were not pathogenic for the fish tested, all of them proved to be virulent for mice. The extra cellular products (ECP) of all V. cholerae and V.mimicus strains showed high proteolytic and phospholipase activities. Moreover, they exhibited a positive cytotoxic response for all the fish and homoiothermic cell lines tested and produced exotoxins with lethaleffects for fish and mice. The virulence properties for mammals exhibited by all these Vibrio strains is a question of special concern mainly in countries where raw fish or fish products are consumed.