Microbe profile: Aeromonas salmonicida: an opportunistic pathogen with multiple personalities.

The bacterial species Aeromonas salmonicida is a fish pathogen. Feared by fish farmers everywhere on Earth over the past century, this species has turned out to be more diverse than initially suspected. While some psychrophilic subspecies cannot grow at temperatures above 25 °C or 30 °C, other mesophilic strains growing up to 37 °C and above are now characterized. Adding to the surprising diversity of this species, some of the mesophilic strains infect mammals and birds. The remarkable diversity is explained in part by the presence of numerous mobile genetic elements, which sculpt and modify the genome of the various strains of this species.

Antibiotics modulate biofilm formation in fish pathogenic isolates of atypical Aeromonas salmonicida.

Atypical Aeromonas salmonicida causes furunculosis infections of non-salmonid fish, which requires antibiotic therapy. However, antibiotics may induce biofilm in some bacteria, which protects them against hostile conditions while allowing them to persist on surfaces, thus forming a reservoir for infection. The aim of this study was to determine whether atypical isolates of A. salmonicida increased biofilm in the presence of two antibiotics, florfenicol and oxytetracycline. A microtitre plate assay was used to quantify biofilm in the presence and absence of each antibiotic. Fifteen of 28 isolates formed biofilms under control conditions, while 23 of 28 isolates increased biofilm formation in the presence of at least one concentration of at least one antibiotic. For oxytetracycline, the most effective concentration causing biofilm to increase was one-quarter of that preventing visible bacterial growth, whereas for florfenicol it was one-half of this value. This is the first study to demonstrate that a bacterial pathogen of fish increases biofilm in response to antibiotics. Biofilm formation may increase the risk of re-infection in culture systems and this lifestyle favours the transmission of genetic material, which has implications for the dissemination of antibiotic-resistance genes and demonstrates the need for enhanced disease prevention measures against atypical A. salmonicida.

Growth and spoilage metabolites production of a mesophilic Aeromonas salmonicida strain in Atlantic salmon (Salmo salar L.) during cold storage in modified atmosphere.

AIMS: The aim of the study was to quantify the growth kinetic parameters and spoilage-associated metabolites of an inoculated strain of Aeromonas salmonicida in pre-rigor filleted Atlantic salmon (Salmo salar L.) stored in vacuum (VP) or modified atmosphere (MAP 60/40% CO2 /N2 ) at 4 and 8°C. METHODS AND RESULTS: The maximum growth rate of A. salmonicida in VP salmon stored at 4°C was 0·56 ± 0·04 day-1 with no detectable lag-phase and the concentration of Aeromonas reached 8·33 log CFU per g after 10 days. The growth rates and maximum population density of Aeromonas in MAP salmon were lower but the applied atmosphere did not inhibit the growth. A selection of metabolites associated with fish spoilage were quantified using 1 H nuclear magnetic resonance (NMR) spectroscopy. The concentration of trimethylamine (TMA) was significantly affected by storage time and temperature, packaging atmosphere and inoculation with A. salmonicida (General Linear Model (GLM), P < 0·001 for all factors). CONCLUSION: The study presents preliminary results on A. salmonicida as a potential spoilage organism in vacuum-packaged salmon during cold storage. The combination of refrigeration and a packaging atmosphere consisting of 60/40 % CO2 /N2 did not completely inhibit the growth but prevented the formation of TMA. SIGNIFICANCE AND IMPACT OF THE STUDY: Little information is available on the spoilage potential of Aeromonas spp. in minimally processed salmon products under different packaging conditions. The study clearly demonstrates the importance of hurdle technology and provides data to further elucidate the significance of Aeromonas spp. as a spoilage organism.

Growth of mesophilic Aeromonas salmonicida in an experimental model of nigiri sushi during cold storage.

The genus Aeromonas includes human pathogenic bacteria frequently isolated from seafood, and the increased consumption of ready-to-eat seafood poses new food safety issues regarding the presence of potentially pathogenic Aeromonas spp. in stored products for raw consumption, such as retail sushi with a shelf life of up to three days. This study assessed 1) the growth kinetics of a mesophilic A. salmonicida strain during storage at 4 °C and 8 °C in a nigiri sushi model, and 2) the strain variability in growth at pH ranging from 3.5 to 10 for a subset of mesophilic Aeromonas strains previously isolated from sushi. Inoculated slices of raw salmon were compared with and without rice. A predictive model for A. hydrophila (ComBase Predictor) did not sufficiently predict growth of the tested strain under the intrinsic conditions of nigiri sushi or salmon at both temperatures. Refrigeration alone (4 °C) did not inhibit growth of A. salmonicida on salmon. Within the first 72 h, representing the typical shelf life of retail sushi products, we observed a 10-fold increase in the concentration of the inoculated strain (including a lag-phase of approximately 34 h). Contact with acidified rice, resulting in a pH drop in the salmon, was the reason for the decreased bacterial viability in the nigiri sushi samples. However, the effect of acidification decreased at 8 °C, resulting in a 2-fold increase in the growth rate and a reduced lag-phase compared to refrigeration. Variability in the ability to grow in different pH levels was observed between strains. The highest color formation rates, representing cellular respiration analyzed in a phenotypic microarray system, were observed between pH 5 and 8. A few strains, including the A. salmonicida strain applied in the nigiri sushi model, were able to grow at pH 4.5 (at optimal temperature). The results demonstrated that mesophilic Aeromonas spp. can represent a microbiological hazard in retail sushi products during cold storage. Rice acidification in combination with low storage temperature (≤4 °C) are prerequisites to prevent growth of potentially pathogenic Aeromonas species during the relatively short shelf life.

Silver nanoparticles: Their role as antibacterial agent against Aeromonas salmonicida subsp. salmonicida in rainbow trout (Oncorhynchus mykiss).

The rise of bacterial resistance to antibiotics is one of the great challenges of our age. One of the strategies to limit the development of antibiotics resistance is the investigation of alternative antimicrobials. As silver nanoparticles demonstrated a potent bactericidal activity in vitro, the aim of this study was to evaluate the in vivo antibacterial activity of silver nanoparticles against Aeromonas salmonicida subsp. salmonicida. Rainbow trout (n = 120) were divided into four groups of 30 fish each. First group was challenged with A. salmonicida (Positive control), the second group was challenged with A. salmonicida and exposed to silver nanoparticles by immersion for three hours (100 µg/L), the third group was challenged with A. salmonicida and intraperitoneally injected with silver nanoparticles (17 µg/mL) and the fourth group was sham-treated and served as a negative control group. At the 7th day post challenge, histopathology of the positive control group revealed the presence of bacterial aggregates in tissues with degenerative and necrotic changes, while at the 35th day post challenge, only liver necrosis persisted. Silver nanoparticles-treated and negative control groups did not show any clinical signs, mortalities or histopathological alterations and they were tested negative for A. salmonicida. The immersion in silver nanoparticles did not result in detectable residues of silver in the muscles 35 days after treatment. These findings demonstrate the antibacterial properties of silver nanoparticles against A. salmonicida infection. Therefore, they could be used for development of antibacterial agents in aquaculture.

Aeromonas salmonicida Growth in Response to Atlantic Salmon Mucins Differs between Epithelial Sites, Is Governed by Sialylated and N-Acetylhexosamine-Containing O-Glycans, and Is Affected by Ca2.

Aeromonas salmonicida causes furunculosis in salmonids and is a threat to Atlantic salmon aquaculture. The epithelial surfaces that the pathogen colonizes are covered by a mucus layer predominantly comprised of secreted mucins. By using mass spectrometry to identify mucin glycan structures with and without enzymatic removal of glycan residues, coupled to measurements of bacterial growth, we show here that the complex Atlantic salmon intestinal mucin glycans enhance A. salmonicida growth, whereas the more simple skin mucin glycans do not. Of the glycan residues present terminally on the salmon mucins, only N-acetylglucosamine (GlcNAc) enhances growth. Sialic acids, which have an abundance of 75% among terminal glycans from skin and of <50% among intestinal glycans, cannot be removed or used by A. salmonicida for growth-enhancing purposes, and they shield internal GlcNAc from utilization. A Ca2+ concentration above 0.1 mM is needed for A. salmonicida to be able to utilize mucins for growth-promoting purposes, and 10 mM further enhances both A. salmonicida growth in response to mucins and binding of the bacterium to mucins. In conclusion, GlcNAc and sialic acids are important determinants of the A. salmonicida interaction with its host at the mucosal surface. Furthermore, since the mucin glycan repertoire affects pathogen growth, the glycan repertoire may be a factor to take into account during breeding and selection of strains for aquaculture.

The protective effect of humic-rich substances on atypical Aeromonas salmonicida subsp. salmonicida infection in common carp (Cyprinus carpio L.).

When challenged with atypical Aeromonas salmonicida subsp. salmonicida, exposure of the common carp (Cyprinus carpio L.) to different humic-rich compounds resulted in a significant reduction in infection rates. Specifically, in fish exposed to (i) humic-rich water and sludge from a recirculating system, (ii) a synthetic humic acid, and (iii) a Leonardite-derived humic-rich extract, infection rates were reduced to 14.9%, 17.0% and 18.8%, respectively, as compared to a 46.8% infection rate in the control treatment. An additional set of experiments was performed to examine the effect of humic-rich components on the growth of the bacterial pathogen. Liquid culture medium supplemented with either humic-rich water from the recirculating system, the synthetic humic acid or the Leonardite humic-rich extract resulted in a growth reduction of 41.1%, 45.2% and 61.6%, respectively, as compared to the growth of the Aeromonas strain in medium devoid of humic substances. Finally, in a third set of experiments it was found that while the innate immune system of the carps was not affected by their exposure to humic-rich substances, their acquired immune system was affected. Fish, immunized against bovine serum albumin, displayed elevated antibody titres as compared to immunized carps which were not exposed to the various sources of humic substances.

Evaluation of the spoilage potential of bacteria isolated from chilled chicken in vitro and in situ.

Microorganisms play an important role in the spoilage of chilled chicken. In this study, a total of 53 isolates, belonging to 7 species of 3 genera, were isolated using a selective medium based on the capacity to spoil chicken juice. Four isolates, namely Aeromonas salmonicida 35, Pseudomonas fluorescens H5, Pseudomonas fragi H8 and Serratia liquefaciens 17, were further characterized to assess their proteolytic activities in vitro using meat protein extracts and to evaluate their spoilage potential in situ. The in vitro studies showed that A. salmonicida 35 displayed the strongest proteolytic activity against both sarcoplasmic and myofibrillar proteins. However, the major spoilage isolate in situ was P. fragi H8, which exhibited a fast growth rate, slime formation and increased pH and total volatile basic nitrogen (TVBN) on chicken breast fillets. The relative amounts of volatile organic compounds (VOCs) originating from the microorganisms, including alcohols, aldehydes, ketones and several sulfur compounds, increased during storage. In sum, this study demonstrated the characteristics of 4 potential spoilage bacteria on chilled yellow-feather chicken and provides a simple and convenient method to assess spoilage bacteria during quality management.

Characterization and mechanism of anti-Aeromonas salmonicida activity of a marine probiotic strain, Bacillus velezensis V4.

The bacterium Aeromonas salmonicida is the causative agent of furunculosis, a systemic, ubiquitous disease of fish in the salmon family, characterized by high mortality and morbidity. Probiotics are a promising approach for prevention of furunculosis in aquaculture. A bacterial strain with anti-A. salmonicida properties, Bacillus velezensis V4, was isolated and the mechanisms underlying these properties were investigated. Anti-A. salmonicida compounds present in cell-free supernatant of V4 were purified and structurally identified as members of the iturin, macrolactin, and difficidin groups. The compounds contributed jointly to inhibition of A. salmonicida, and the diversity of the compounds was related to the versatility of their mode of action. Addition of the compounds to A. salmonicida cell suspensions reduced cell density. Analyses by confocal microscopy and scanning electron microscopy revealed cell membrane disruption, deletion of cellular content, and cell lysis of A. salmonicida. The V4 genome was sequenced, and gene clusters involved in synthesis of anti-Aeromonas compounds were detected and identified. A possible probiotic effect on growth performance of Oncorhynchus mykiss (rainbow trout) was investigated by addition of 0, 1, and 3 % (v/w) V4. Relative to control, mortality was reduced 27.25 % in the 1 % addition group and 81.86 % in the 3 % addition group. Feed coefficient ratio was reduced 19.49 % and weight gain ratio was increased 71.22 % in the 1 % addition group. Our findings demonstrate that V4 is an effective probiotic strain in O. mykiss and has clear potential for both control of furunculosis and growth promotion of aquaculture animals.

Identification of the Ferric-Acinetobactin Outer Membrane Receptor in Aeromonas salmonicida subsp. salmonicida and Structure-Activity Relationships of Synthetic Acinetobactin Analogues.

Aeromonas salmonicida subsp. salmonicida, the causative agent of furunculosis in several fish species, produces acinetobactin and amonabactin as siderophores. In a previous study, we chemically characterized these siderophores and proposed a biosynthetic pathway based on genetic analysis. However, the internalization mechanisms of ferric-acinetobactin and ferric-amonabactin remain largely unknown. In the present study, we demonstrate that the outer membrane protein FstB is the ferric-acinetobactin receptor in A. salmonicida since an fstB defective mutant is unable to grow under iron limitation and does not use acinetobactin as an iron source. In order to study the effect that structural changes in acinetobactin have on its siderophore activity, a collection of acinetobactin-based analogues was synthesized, including its enantiomer and four demethylated derivatives. The biological activity of these analogues on an fstB(+) strain compared to an fstB(-) strain allowed structure-activity relationships to be elucidated. We found a lack of enantiomer preference on the siderophore activity of acinetobactin over A. salmonicida or on the molecular recognition by FstB protein receptor. In addition, it was observed that A. salmonicida could not use acinetobactin analogues when imidazole or a similar heterocyclic ring was absent from the structure. Surprisingly, removal of the methyl group at the isoxazolidinone ring induced a higher biological activity, thus suggesting alternative route(s) of entry into the cell that must be further investigated. It is proposed that some of the synthetic acinetobactin analogues described here could be used as starting points in the development of novel drugs against A. salmonicida and probably against other acinetobactin producers like the human pathogen Acinetobacter baumannii.

High Concentration of Red Clay as an Alternative for Antibiotics in Aquaculture.

The use of antibiotics in aquaculture raises environmental and food safety concerns because chronic exposure of an aquatic ecosystem to antibiotics can result in the spread of antibiotic resistance, bioaccumulation of antibiotics in the organisms, and transfer of antibiotics to humans. In an attempt to overcome these problems, high-concentration red clay was applied as an alternative antibiotic against the following common fish pathogens: Aeromonas salmonicida, Vibrio alginolyticus, and Streptococcus equinus. The growth of A. salmonicida and V. alginolyticus was retarded by red clay, whereas that of S. equinus was promoted. Phase contrast and scanning electron microscopy analyses confirmed the attachment of red clay on cell surfaces, resulting in rapid gravitational removal and cell surface damage in both A. salmonicida and V. alginolyticus, but not in S. equinus. Different cell wall properties of grampositive species may explain the unharmed cell surface of S. equinus. Significant levels of oxidative stress were generated in only the former two species, whereas significant changes in membrane permeability were found only in S. equinus, probably because of its physiological adaptation. The bacterial communities in water samples from Oncorhynchus mykiss aquacultures supplemented with red clay showed similar structure and diversity as those from oxytetracycline-treated water. Taken together, the antibiotic effects of high concentrations of red clay in aquaculture can be attributed to gravitational removal, cell surface damage, and oxidative stress production, and suggest that red clay may be used as an alternative for antibiotics in aquaculture.

AsaGEI2b: a new variant of a genomic island identified in the Aeromonas salmonicida subsp. salmonicida JF3224 strain isolated from a wild fish in Switzerland.

Aeromonas salmonicida subsp. salmonicida is the causal agent of furunculosis in salmonids. We recently identified a group of genomic islands (AsaGEI) in this bacterium. AsaGEI2a, one of these genomic islands, has almost exclusively been identified in isolates from North America. To date, Aeromonas salmonicida subsp. salmonicida JF3224, a strain isolated from a wild brown trout (Salmo trutta) caught in Switzerland, was the only European isolate that appeared to bear AsaGEI2a. We analyzed the genome of JF3224 and showed that the genomic island in JF3224 is a new variant of AsaGEI, which we have called AsaGEI2b. While AsaGEI2b shares the same integrase gene and insertion site as AsaGEI2a, it is very different in terms of many other features. Additional genomic investigations combined with PCR genotyping revealed that JF3224 is sensitive to growth at 25°C, leading to insertion sequence-dependent rearrangement of the locus on the pAsa5 plasmid that encodes a type three secretion system, which is essential for the virulence of the bacterium. The analysis of the JF3224 genome confirmed that AsaGEIs are accurate indicators of the geographic origins of A. salmonicida subsp. salmonicida isolates and is another example of the susceptibility of the pAsa5 plasmid to DNA rearrangements.

In vivo study of spoilage bacteria on polyphenoloxidase activity and melanosis of modified atmosphere packaged Pacific white shrimp.

This study investigated the effect of the three spoilage bacteria (Carnobacterium maltaromaticum, Shewanella putrefaciens and Aeromonas salmonicida) on the development of melanosis by inoculating the bacteria on modified atmosphere packaged shrimp. The three bacteria, which inoculated at about 5 log cfu/g, proliferated to a maximum level of 7.49, 6.86 and 6.89 log cfu/g, respectively at the end of storage. In regards to the effect of bacteria on melanosis, it was found that C. maltaromaticum did not display a significant effect on PPO activity and melanosis, and A. salmonicida showed an inhibitory effect on PPO activity with an activity value of less than 1.5 Units/ml. However, the PPO activity of shrimp inoculated S. putrefaciens was about one time higher than other samples and greater melanosis was displayed in the first 48 h. As melanosis can cause sensory quality loss, the growth of S. putrefaciens should be limited.

[Studies of prevalence rate of furunculosis caused by infection by Aeromonas salmonicida in salmonids of the southern part of Sakhalin Island].

Ichthyological studies of spawners of salmonids in the south of Sakhalin Island were studied. Cases of furunculosis disease were revealed. The agent of the disease Aeromonas salmonicida was isolated. Its morphological, physiological-biochemical, and antagonistic properties were studied, and the virulence of the isolated strains was determined. For supporting the species status of the studied strains of A. salmonicida, a molecular-genetic analysis was performed.

An insertion sequence-dependent plasmid rearrangement in Aeromonas salmonicida causes the loss of the type three secretion system.

Aeromonas salmonicida, a bacterial fish pathogen, possesses a functional Type Three Secretion System (TTSS), which is essential for its virulence. The genes for this system are mainly located in a single region of the large pAsa5 plasmid. Bacteria lose the TTSS region from this plasmid through rearrangements when grown in stressful growth conditions. The A. salmonicida genome is rich in insertion sequences (ISs), which are mobile DNA elements that can cause DNA rearrangements in other bacterial species. pAsa5 possesses numerous ISs. Three IS11s from the IS256 family encircle the rearranged regions. To confirm that these IS11s are involved in pAsa5 rearrangements, 26 strains derived from strain A449 and two Canadian isolates (01-B526 and 01-B516) with a pAsa5 rearrangement were tested using a PCR approach to determine whether the rearrangements were the result of an IS11-dependent process. Nine out of the 26 strains had a positive PCR result, suggesting that the rearrangement in these strains were IS-dependent. The PCR analysis showed that all the rearrangements in the A449-derived strains were IS11-dependent process while the rearrangements in 01-B526 and 01-B516 could only be partially coupled to the action of IS11. Unidentified elements that affect IS-dependent rearrangements may be present in 01-B526 and 01-B516. Our results suggested that pAsa5 rearrangements involve IS11. This is the first study showing that ISs are involved in plasmid instability in A. salmonicida.

Alteration of virulence factors and rearrangement of pAsa5 plasmid caused by the growth of Aeromonas salmonicida in stressful conditions.

Aeromonas salmonicida, a fish pathogen, is the causative agent of furunculosis. It was already shown that growing this bacterium in stressful conditions such as temperature above 22°C might lead to virulence attenuation. Unfortunately, many veterinary microbiology services and reference centers still routinely cultivate A. salmonicida at 25°C. Here we tested the presence of virulence factors by growth on specific medium as well as the integrity of the pAsa5 plasmid, which bears an important virulence factor, the type III secretion system (TTSS), by PCR analysis in twenty strains, most of which were grown at 25°C in their laboratory of origin. The analysis revealed that strains, which encountered the more stressful growth conditions displayed the most frequent absence of A-layer protein and secreted proteolytic activity. Moreover, many strains had lost parts of the pAsa5 plasmid in which the TTSS region was almost always affected. To confirm the effect of stressful growth conditions on the plasmid, three strains with an intact pAsa5 were cultured at 25°C for two weeks. A low but significant fraction of the tested colonies displayed pAsa5 rearrangements. The rearrangement always affected the TTSS region and led to a loss of virulence in the Dictyostelium discoideum co-culture assay. These results demonstrate that the instability of pAsa5 did not lead to its complete loss as previously proposed but to a more complex rearrangement phenomenon and emphasizes the necessity to grow A. salmonicida in appropriate conditions to preserve the complete virulence of the bacterium.

Responses of Atlantic cod Gadus morhua head kidney leukocytes to phytase produced by gastrointestinal-derived bacteria.

This study identified phytase-producing bacteria that were previously isolated from the gastrointestinal tract of Atlantic cod, Gadus morhua and determined its effect on head kidney leukocytes. Out of the 216 bacterial strains tested, the two phytase producers were identified as Pseudomonas sp. and Psychrobacter sp. based on their 16S rDNA sequence. Crude phytase from these two bacterial strains was produced employing the shake flask method. Even though the total protein of the crude phytase was not significantly different for the two bacteria, the phytase activity of the crude enzyme produced by Pseudomonas sp. (97.1±16.7 U) was significantly higher than that of the enzyme from Psychrobacter sp. (75.9±2.4 U). When cod head kidney leukocytes were incubated with the crude phytase (50 µg ml(-1)), it resulted in enhanced cell proliferation, higher myeloperoxidase, and acid phosphatase activities. Extracellular responses-respiratory burst activity and hydrogen peroxide production were not enhanced by the crude enzyme. As a consequence, the growth of two pathogenic bacteria Aeromonas salmonicida and Vibrio anguillarum was not suppressed by the supernatants obtained from head kidney leukocytes incubated with the crude bacterial phytase. Thus, the enzyme from phytase-producing intestinal bacteria of Atlantic cod can stimulate intracellular head kidney leukocyte activities but not the production of extracellular substances that are involved in antibacterial response. These have implications on the potential use of bacterial phytase as feed supplement to boost cellular immune response of the fish and could be employed as a health management strategy in culture systems.

Interleukin-17D in Atlantic salmon (Salmo salar): molecular characterization, 3D modelling and promoter analysis.

IL-17 is a proinflammatory cytokine that plays an important role in the clearance of extracellular bacteria and contributes to the pathology of many autoimmune and allergic conditions. Much work on IL-17 has been done in humans and higher vertebrates while little work has been conducted in lower vertebrates including fish. In this study, we have cloned and characterized the full-length cDNA and genomic sequence of IL-17D from Atlantic salmon. The Atlantic salmon IL-17D (AsIL-17D) cDNA possessed an open reading frame of 621 bp encoding a putative protein of 206 aa with a predicted molecular weight of 23 kDa. The AsIL-17D gene has two exons and one intron showing the same (genome) organisation compared to zebrafish IL-17D. The encoded protein showed 97.6-48.8% identities to other IL-17D homologues, eight conserved cysteine residues were found within this group. Conserved residues believed to be important in receptor binding were also confirmed in salmon IL-17D by homology modelling. Phylogenetic analysis also confirmed the close relationship with other IL-17D homologues. Functional characterization of the 5' flanking region indicated that the region between -1552 and -150 contained sufficient elements for promoter activity. Tissue expression studies by real-time PCR showed a predominant expression of IL-17D transcript in gonads, skin, intestine, thymus of Atlantic salmon. The involvement of IL-17D during proinflammatory responses was demonstrated by investigating the time-dependent expression profile of IL-17D in head kidney and spleen following intraperitoneal injection of live Aeromonas salmonicida, LPS, and beta-glucan. This study provides further evidence for the existence of distinct homologue of IL-17D isoform in fish showing early expression induced by immunostimulants and bacterial infection that supports the fact that IL-17D is regulated by inflammatory processes in fish.

Effect of Lactococcus lactis CLFP 100 and Leuconostoc mesenteroides CLFP 196 on Aeromonas salmonicida Infection in brown trout (Salmo trutta).

Aeromonas salmonicida is the etiological agent of furunculosis in salmonid fish. This pathogen is important from an epizootic perspective because fish surviving an outbreak can remain lifelong asymptomatic carriers, serving as reservoirs of infection. As a result, the early detection and the control of infection are essential to prevent the spread of new furunculosis outbreaks. We have thus analyzed the effect of probiotic administration on the incidence of A. salmonicida in brown trout (Salmo trutta), that were subjected to temperature stress. Treatment with probiotic strains (Lactococcus lactis CLFP 100 and Leuconostoc mesenteroides CLFP 196) resulted in a higher survival rate after challenge, activation of phagocytic cells in the head kidney, and a lower rate of pathogen proliferation in the intestine as determined by real-time PCR.

Application of an immunoaffinity-based preconcentration method for mass spectrometric analysis of the O-chain polysaccharide of Aeromonas salmonicida from in vitro- and in vivo-grown cells.

In this study, application of magnetic beads (Dynabeads) coated with Aeromonas salmonicida lipopolysaccharide-specific polyclonal antisera to MS-based characterization of bacterial lipopolysaccharides has been evaluated. The results showed that the affinity-based preconcentration strategy resulted in at least a 100-fold increase in the detection of sensitivity, affording direct capillary electrophoresis (CE)-MS analysis of A. salmonicida lipopolysaccharide O-chain polysaccharide from in vitro-cultured cells. Subsequent CE-MS analysis of in vivo-grown cells of A. salmonicida confirmed significant changes in the structure of the lipopolysaccharide O-chain polysaccharide as a result of in vivo cultivation.