Mechanism of sturgeon intestinal inflammation induced by Yersinia ruckeri and the effect of florfenicol intervention.

The mechanism by which Y. ruckeri infection induces enteritis in Chinese sturgeon remains unclear, and the efficacy of drug prevention and control measures is not only poor but also plagued with numerous issues. We conducted transcriptomic and 16 S rRNA sequencing analyses to examine the differences in the intestinal tract of hybrid sturgeon before and after Y. ruckeri infection and florfenicol intervention. Our findings revealed that Y. ruckeri induced the expression of multiple inflammatory factors, including il1ß, il6, and various chemokines, as well as casp3, casp8, and multiple tumor necrosis factor family members, resulting in pathological injury to the body. Additionally, at the phylum level, the relative abundance of Firmicutes and Bacteroidota increased, while the abundance of Plesiomonas and Cetobacterium decreased at the genus level, altering the composition of the intestinal flora. Following florfenicol intervention, the expression of multiple apoptosis and inflammation-related genes was down-regulated, promoting tissue repair. However, the flora became further dysregulated, increasing the risk of infection. In conclusion, our analysis of the transcriptome and intestinal microbial composition demonstrated that Y. ruckeri induces intestinal pathological damage by triggering apoptosis and altering the composition of the intestinal microbiota. Florfenicol intervention can repair pathological damage, but it also exacerbates flora imbalance, leading to a higher risk of infection. These findings help elucidate the molecular mechanism of Y. ruckeri-induced enteritis in sturgeon and evaluate the therapeutic effect of drugs on intestinal inflammation in sturgeon.

Recombinant expression of Yersinia ruckeri outer membrane proteins in Escherichia coli extracellular vesicles.

The secretion of extracellular vesicles (EVs) is a common process in Gram-negative bacteria and can be exploited for biotechnological applications. EVs pose a self-adjuvanting, non-replicative vaccine platform, where membrane and antigens are presented to the host immune system in a non-infectious fashion. The secreted quantity of EVs varies between Gram-negative bacterial species and is comparatively high in the model bacterium E. coli. The outer membrane proteins OmpA and OmpF of the fish pathogen Y. ruckeri have been proposed as vaccine candidates to prevent enteric redmouth disease in aquaculture. In this work, Y.ruckeri OmpA or OmpF were expressed in E. coli and recombinant EVs were isolated. To avoid competition between endogenous E. coli OmpA or OmpF, Y. ruckeri OmpA and OmpF were expressed in E. coli strains lacking ompA, ompF, and in a quadruple knockout strain where the four major outer membrane protein genes ompA, ompC, ompF and lamB were removed. Y.ruckeri OmpA and OmpF were successfully expressed in EVs derived from the E. coli mutants as verified by SDS-PAGE, heat modifiability and proteomic analysis using mass-spectrometry. Transmission electron microscopy revealed the presence of EVs in all E. coli strains, and increased EV concentrations were detected when expressing Y. ruckeri OmpA or OmpF in recombinant EVs compared to empty vector controls as verified by nanoparticle tracking analysis. These results show that E. coli can be utilized as a vector for production of EVs expressing outer membrane antigens from Y. ruckeri.

Effect of Dietary Origanum onites on Growth, Non Specific Immunity and Resistance against Yersinia ruckeri of Rainbow Trout ( Oncorhynchus mykiss).

Natural substances has been identified to maintain health and improve growth performance in the aquaculture. The effect of Origanum onites on growth and immune response of rainbow trout was investigated. Experimental groups (A and B) of 70 fish were separated into 10 different treatments. A groups were fed with dietary administration of O. onites essential oil (0.5 mL kg-1 and 3.0 mL kg-1) and crude powder (1.0 g kg-1 and 10.0 g kg-1) for a period of 8 weeks. Other groups (B) were vaccinated against Yersinia ruckeri at the beginning of experiment and then fed the same diets described above. Results showed that feed conversion ratio in fish fed a combination of O. onites and vaccine was statistically better than the control. NBT-positive cells, phagocytic activity, serum lysozyme activity and immunoglobulin M level were stimulated in both non vaccinated and vaccinated fish (p<0.05). Cumulative mortality in fish fed O. onites was lower than controls following challenge with Y. ruckeri. No mortality was observed in vaccinated fish fed with 0.5 mL kg-1 of O. onites. These results indicated that dietary administration of O. onites could act as an enhanced non specific immune response, growth performance and resistance to Y. ruckeri.

Exploring Yersinia ruckeri (O1 Biotype 2) infection in three early life-stages of rainbow trout.

Enteric redmouth disease (ERM) caused by the enterobacterium Yersinia ruckeri poses a significant threat to salmonid aquaculture globally. Despite decades of experimental infection studies, key knowledge gaps remain regarding the onset of disease susceptibility and mechanisms of immunity during early developmental stages, undermining disease management efforts in all susceptible life-stages. In this study, a series of immersion challenges were conducted, challenging and re-challenging rainbow trout Oncorhynchus mykiss (Walbaum) at 7, 14 and 51 d post-hatch (dph; mean weights = 0.085, 0.1 and 2.0 g respectively) to high concentrations (1.72 × 107-1.1 × 108 CFU) of Y. ruckeri at 15°C. This study indicates the hitherto unknown initial point of susceptibility to infection as the time of first ingestion of exogenous food (14 dph), and shows that individuals surviving primary challenge at 14 dph are significantly more likely to survive re-challenge at 51 dph compared with naive individuals (hazard ratio = 1.446, p = 0.032). Other key findings include large variation in mortality between different development-stages, from 21.1% at 14 dph to 81.2% at 51 dph, and novel age-dependent symptoms not reported previously. Results from this study enhance our understanding of ERM in juvenile rainbow trout and inform the development of improved aquatic animal health management strategies, thereby contributing to the productivity and sustainability of salmonid aquaculture into the future.

Yersinia ruckeri infection activates local skin and gill B cell responses in rainbow trout.

Teleost fish lack organized structures in mucosal tissues such as those of mammals, but instead contain dispersed B and T cells with the capacity to respond to external stimuli. Nonetheless, there is still a great lack of knowledge regarding how B cells differentiate to plasmablasts/plasma cells in these mucosal surfaces. To contribute to a further understanding of the mechanisms through which fish mucosal B cells are activated, in the current study, we have studied the B cell responses in the skin and gills of rainbow trout (Oncorhynchus mykiss) exposed to Yersinia ruckeri. We have first analyzed the transcription levels of genes related to B cell function in both mucosal surfaces, and in spleen and kidney for comparative purposes. In a second experiment, we have evaluated how the infection affects the presence and size of B cells in both skin and gills, as well as the presence of plasmablasts secreting total or specific IgMs. The results obtained in both experiments support the local differentiation of B cells to plasmablasts/plasma cells in the skin and gills of rainbow trout in response to Y. ruckeri. Interestingly, these plasmablasts/plasma cells were shown to secrete specific IgMs as soon as 5 days after the exposure. These findings contribute to a further understanding of how B cells in the periphery respond to immune stimulation in teleost fish.

Effects of diets rich in Agaricus bisporus polysaccharides on the growth, antioxidant, immunity, and resistance to Yersinia ruckeri in channel catfish.

To promote the application of Agaricus bisporus polysaccharides (ABPs) in channel catfish (Ictalurus punctatus) culture, we evaluated the effects of ABPs on the growth, immunity, antioxidant, and antibacterial activity of channel catfish. When the amount of ABPs was 250 mg/kg, channel catfish's weight gain and specific growth rates increased significantly while the feed coefficient decreased. We also found that adding ABPs in the feed effectively increased the activities of ACP, MDA, T-SOD, AKP, T-AOC, GSH, and CAT enzymes and immune-related genes such as IL-1ß, Hsp70, and IgM in the head kidney of channel catfish. Besides, long-term addition will not cause pathological damage to the head kidney. When the amount of ABPs was over 125 mg/kg, the protection rate of channel catfish was more than 60%. According to the intestinal transcriptome analysis, the addition of ABPs promoted the expression of intestinal immunity genes and growth metabolism-related genes and enriched multiple related KEEG pathways. When challenged by Yersinia ruckeri infection, the immune response of channel catfish fed with ABPs was intenser and quicker. Additionally, the 16S rRNA gene sequencing analysis showed that the composition of the intestinal microbial community of channel catfish treated with ABPs significantly changed, and the abundance of microorganisms beneficial to channel catfish growth, such as Firmicutes and Bacteroidota increased. In conclusion, feeding channel catfish with ABPs promoted growth, enhanced immunity and antioxidant, and improved resistance to bacterial infections. Our current results might promote the use of ABPs in channel catfish and even other aquacultured fish species.

Comprehensive Analysis of YTH Domain-Containing Genes, Encoding m6A Reader and Their Response to Temperature Stresses and Yersinia ruckeri Infection in Rainbow Trout (Oncorhynchus mykiss).

YTH domain-containing genes are important readers of N6-methyladenosine (m6A) modifications with ability to directly affect the fates of distinct RNAs in organisms. Despite their importance, little is known about YTH domain-containing genes in teleosts until now. In the present study, a total of 10 YTH domain-containing genes have been systematically identified and functionally characterized in rainbow trout (Oncorhynchus mykiss). According to the phylogenetic tree, gene structure and syntenic analysis, these YTH domain-containing genes could be classified into three evolutionary subclades, including YTHDF, YTHDC1 and YTHDC2. Of them, the copy number of OmDF1, OmDF2, OmDF3, and OmDC1 were duplicated or even triplicated in rainbow trout due to the salmonid-specific whole-genome duplication event. The three-dimensional protein structure analysis revealed that there were similar structures and the same amino acid residues that were associated with cage formation between humans and rainbow trout, implying their similar manners in binding to m6A modification. Additionally, the results of qPCR experiment indicated that the expression patterns of a few YTH domain-containing genes, especially OmDF1b, OmDF3a and OmDF3b, were significantly different in liver tissue of rainbow trout under four different temperatures (7 °C, 11 °C, 15 °C, and 19 °C). The expression levels of OmDF1a, OmDF1b and OmDC1a were obviously repressed in spleen tissue of rainbow trout at 24 h after Yersinia ruckeri infection, while increased expression was detected in OmDF3b. This study provides a systemic overview of YTH domain-containing genes in rainbow trout and reveals their biological roles in responses to temperature stress and bacterial infection.

Pan-genome survey of the fish pathogen Yersinia ruckeri links accessory- and amplified genes to virulence.

While both virulent and putatively avirulent Yersinia ruckeri strains exist in aquaculture environments, the relationship between the distribution of virulence-associated factors and de facto pathogenicity in fish remains poorly understood. Pan-genome analysis of 18 complete genomes, representing established virulent and putatively avirulent lineages of Y. ruckeri, revealed the presence of a number of accessory genetic determinants. Further investigation of 68 draft genome assemblies revealed that the distribution of certain putative virulence factors correlated well with virulence and host-specificity. The inverse-autotransporter invasin locus yrIlm was, however, the only gene present in all virulent strains, while absent in lineages regarded as avirulent. Strains known to be associated with significant mortalities in salmonid aquaculture display a combination of serotype O1-LPS and yrIlm, with the well-documented highly virulent lineages, represented by MLVA clonal complexes 1 and 2, displaying duplication of the yrIlm locus. Duplication of the yrIlm locus was further found to have evolved over time in clonal complex 1, where some modern, highly virulent isolates display up to three copies.

Screening and activity of potential gastrointestinal probiotic lactic acid bacteria against Yersinia ruckeri O1b.

Yersiniosis of cultured Atlantic salmon is a recurrent fish health management challenge in many continents. The causative organism, Yersinia ruckeri, can reside latently in the gut and lead to acute infection and disease during hatchery and sea-transfer stages. One potential prevention approach is the administration of probiotic bacteria to suppress gut colonization of Y. ruckeri. Our study aimed to isolate and identify anti-Yersinia activity among lactic acid bacteria (LAB) isolated from the gastrointestinal tract (GIT) of aquatic animals. Of the 186 aquatic GIT isolates examined, three strains showed diffusible antimicrobial activity towards Y. ruckeri O1b. Analysis of 16 s rRNA gene sequences indicated the three bacterial strains were Enterococci, related to Enterococcus sp. (99%), Enterococcus thailandicus (99%), and Enterococcus durans (99%). Anti-Yersinia activity was maintained at neutral pH (~6.5-7.0), and in-vitro environmental tolerance assays showed the three strains could withstand simulated salmonids gastrointestinal tract conditions of: low pH (3.4) and 3% bile salt content. All three Enterococci strains showed higher adhesion to the intestinal mucus of Atlantic salmon than Y. ruckeri O1b (E. durans 24%, E. enterococcus sp. 25% and E. thailandicus 98%, compared to Y. ruckeri O1b 5%). However, only Enterococcus sp. and E. thailandicus were able to grow in the salmon intestinal mucus broth while E. durans showed no growth. Anti-Yersinia activity was completely inactivated by proteinase-K treatment, suggesting that the active compound/s are proteinaceous and may be bacteriocin-like inhibitory substances (BLIS). Our data indicate that Enterococcus sp. MA176 and E. thailandicus MA122 are potential probionts for the prevention of yersiniosis in salmonids. Further in-vivo studies are required to determine whether these bacteria reduce the incidence of yersiniosis in Atlantic salmon.

Co-existence of two Yersinia ruckeri biotypes and serotype O1a retrieved from rainbow trout (Oncorhynchus mykiss) farmed in Puno, Peru.

Yersinia ruckeri causes important economic losses for rainbow trout (Oncorhynchus mykiss) farms worldwide. This bacterial disease is likely the most common among trout in Peru; however, no commercial vaccine is available nationally, which is, in part, due to a lack of information on the bacterium. The aim of the current study was to characterize 29 Y. ruckeri isolates sampled from seven cage-reared farms in the Puno Region, the focal point for aquaculture activities in Peru. For this, samples were taken from fish with clinical signs (i.e. haemorrhages, uni- or bilateral exophthalmia, hyphaemia and/or melanosis). Notable among our findings was the existence of both Y. ruckeri biotype 1 (9 isolates) and biotype 2 (20 isolates; negative for sorbitol and Tween 80). The isolates further differed in API profiles 5307100 (21 isolates), 1307100 (4 isolates), 1305100 (2 isolates), 1307120 (1 isolate) and 5305100 (1 isolate), with the main differences being in the tests for lysine decarboxylase, gelatine hydrolysis and D-saccharose fermentation. Despite these differences, all isolates shared identical ERIC-PCR and REP-PCR profiles and belonged to the O1a serotype. Fingerprints were identical to the reference strain CECT 955 (serotype O1a). The information obtained will be used for epidemiological purposes by health authorities and for the development of a vaccine against Y. ruckeri, a prominent request made by fish farmers in Peru.

qPCR screening for Yersinia ruckeri clonal complex 1 against a background of putatively avirulent strains in Norwegian aquaculture.

Although a number of genetically diverse Yersinia ruckeri strains are present in Norwegian aquaculture environments, most if not all outbreaks of yersiniosis in Atlantic salmon in Norway are associated with a single specific genetic lineage of serotype O1, termed clonal complex 1. To investigate the presence and spread of virulent and putatively avirulent strains in Norwegian salmon farms, PCR assays specific for Y. ruckeri (species level) and Y. ruckeri clonal complex 1 were developed. Following extensive screening of water and biofilm, the widespread prevalence of putatively avirulent Y. ruckeri strains was confirmed in freshwater salmon hatcheries, while Y. ruckeri clonal complex 1 was found in fewer farms. The formalin-killed bacterin yersiniosis vaccine was detected in environmental samples by both PCR assays for several weeks post-vaccination. It is thus important to interpret results from recently vaccinated fish with great care. Moreover, field studies and laboratory trials confirmed that stressful management procedures may result in increased shedding of Y. ruckeri by sub-clinically infected fish. Analysis of sea water sampled throughout thermal delousing procedures proved effective for detection of Y. ruckeri in sub-clinically infected populations.

Functional and Molecular Immune Response of Rainbow Trout (Oncorhynchus mykiss) Following Challenge with Yersinia ruckeri.

Currently, aquaculture production of rainbow trout (Oncorhynchus mykiss) is a multibillion dollar industry; nevertheless, the development of this sector has not been exempt from pitfalls related to the recurrent presence of pathogens of bacterial origin. This is the case of Yersinia ruckeri, the etiologic agent of the infectious pathology known as Enteric Red Mouth Disease (ERM), causing serious economic losses that can be as high as 30-70% of production. Although several studies have been performed regarding pathogen features and virulence factors, more information is needed about the host defense mechanism activation after infection. Given this perspective, this study aimed to evaluate rainbow trout's short-term innate immune response against infection with Y. ruckeri. A series of factors linked to the innate immune response were evaluated, including determination of hematological parameters, oxidative stress biomarkers, and analysis of the expression of immune-related genes. Results showed a significant decrease in several hematological parameters (white blood cell count, hematocrit, neutrophils, monocytes, lymphocytes, and thrombocytes) and oxidative stress indicators (SOD) between the control and infected groups. In addition, there were significant differences in the level of gene expression between infected individuals and the control group. Most of these genes (il-1ß, il-8, il-10, tnf-α1, tnf-α2, socs3, mmp-9, cath, hsp-70, saa, fer, pcb) were upregulated within the first 24 h following infection. Results from this study showed more insights into the short-term immune response of rainbow trout to infection with Y. ruckeri, which may be useful for the establishment of biomarkers that may be used for the early detection of ERM.

Biotyping reveals loss of motility in two distinct Yersinia ruckeri lineages exclusive to Norwegian aquaculture.

Non-motile strains of Yersinia ruckeri, known as Y. ruckeri biotype 2, now dominate amongst clinical isolates retrieved from rainbow trout internationally. Due to an acute increase in the number of yersiniosis cases in Norway in recent years, followed by introduction of widespread intraperitoneal vaccination against the disease, an investigation on the prevalence of Y. ruckeri biotype 2 in Norwegian aquaculture was conducted. We biotyped 263 Y. ruckeri isolates recovered from diseased salmonids in Norway between 1985 and 2020. A total of seven biotype 2 isolates were identified, four of which were collected between 1985 and 1987, and three of which belong to the current epizootic clone, isolated from two different sea-farms in 2017. Whole-genome sequencing revealed single non-synonymous nucleotide polymorphisms in the flagellar genes flhC in isolates from the 1980s, and in fliP in isolates from 2017. In both variants, motility was restored both by complementation with wild-type alleles in trans and via spontaneous mutation-driven reversion following prolonged incubation on motility agar. While biotype 2 strains do not yet seem to have become broadly established in Norwegian aquaculture, the seven isolates described here serve to document a further two independent cases of Y. ruckeri biotype 2 emergence in salmonid aquaculture.

Integrative analyses of probiotics, pathogenic infections and host immune response highlight the importance of gut microbiota in understanding disease recovery in rainbow trout (Oncorhynchus mykiss).

AIMS: Given the pivotal role played by the gut microbiota in regulating the host immune system, great interest has arisen in the possibility of controlling fish health by modulating the gut microbiota. Hence, there is a need to better understand of the host-microbiota interactions after disease responses to optimize the use of probiotics to strengthen disease resilience and recovery. METHODS AND RESULTS: We tested the effects of a probiotic feed additive in rainbow trout and challenged the fish with the causative agent for enteric red mouth disease, Yersinia ruckeri. We evaluated the survival, host immune gene expression and the gut microbiota composition. Results revealed that provision of probiotics and exposure to Y. ruckeri induced immune gene expression in the host, which were associated with changes in the gut microbiota. Subsequently, infection with Y. ruckeri had very little effect on microbiota composition when probiotics were applied, indicating that probiotics increased stabilisation of the microbiota. Our analysis revealed potential biomarkers for monitoring infection status and fish health. Finally, we used modelling approaches to decipher interactions between gut bacteria and the host immune gene responses, indicating removal of endogenous bacteria elicited by non-specific immune responses. CONCLUSIONS: We discuss the relevance of these results emphasizing the importance of host-microbiota interactions, including the protective potential of the gut microbiota in disease responses. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results highlight the functional consequences of probiotic-induced changes in the gut microbiota post infection and the resulting host immune response.

Long-term efficacy of nasal vaccination against enteric red mouth (ERM) disease and infectious hematopoietic necrosis (IHN) in juvenile rainbow trout (Oncorhynchus mykiss).

Previous research demonstrated that bacterial and viral vaccines delivered via the nasal route in rainbow trout (Oncorhynchus mykiss) at 7 and 28 days post-vaccination are highly protective (>95% protection). Long-term protection following nasal vaccination in teleosts has not been evaluated. The goal of this study was to assess efficacy and immune responses at 6 months (mo) post-vaccination (mpv), and long-lasting immune responses at 12 mpv of two different vaccines: an inactivated enteric red mouth disease (ERM) Yersinia ruckeri bacterin and a live attenuated infectious hematopoietic necrosis virus (IHNV) vaccine. Juvenile rainbow trout were vaccinated for Y. ruckeri via intraperitoneal (I.P.) and intranasal (I.N.) routes, and for IHNV by intramuscular (I.M.) and I.N. routes, then challenged at 6 mpv. Immune responses were determined at 6 and 12 mpv. ERM vaccine I.P. delivery elicited significantly higher serum IgM-specific titers that remained elevated compared to mock-vaccinated fish at 6 mpv. By 12 mpv, antibody titers to Y. ruckeri were not significantly different across all treatments. Following Y. ruckeri challenge at 6 mpv, a significant difference in cumulative percent mortality (CPM) was found for I.P.-vaccinated fish but not I.N.-vaccinated fish. I.M. and I.N. vaccination with live attenuated IHNV did not result in significant specific serum IgM titers at 6 or 12 mpv. Yet, I.N.-vaccinated fish showed the lowest CPM 6 mpv indicating long-term protection that does not correlate with systemic IgM responses. Repertoire analyses confirmed unique expansions of VH-JH rearrangements in the spleen of rainbow trout 12 mpv that varied with the type of vaccine and route of vaccination. Combined, these data demonstrate that I.N. vaccination with a live attenuated viral vaccine confers long lasting protection, but I.N. ERM vaccination does not and booster before 6 mpv is recommended.

Modulatory effect of dietary copper nanoparticles and vitamin C supplementations on growth performance, hematological and immune parameters, oxidative status, histology, and disease resistance against Yersinia ruckeri in rainbow trout (Oncorhynchus mykiss).

Copper and vitamin C are micronutrients needed for the living organism's functions. Vitamin C has a great effect on the immune system of fish. The present study aimed to evaluate the effects of dietary copper nanoparticles (Cu-NPs) and vitamin C (VC) supplementations on rainbow trout (Oncorhynchus mykiss) juveniles. So, 216 rainbow trout juveniles were randomly assigned to six groups with trial diets supplemented with Cu-NPs and VC including 0/0 (T1, control diet), 0/250 (T2), 0/500 (T3), 2/250 (T4), 2/500 (T5), and 2/0 (T6) mg Cu-NPs/VC per kg diet. After the feeding trial for 60 days, the fish were challenged with Yersinia ruckeri, and the survival rate was calculated for 15 days. Based on the data analysis, weight gain (WG), specific growth rate (SGR), protein efficiency ratio (PER), lysozyme, alternative complement activity (ACH50), hematocrit (Hct), hemoglobin (Hb), and mean corpuscular volume (MCV) were significantly (p < 0.05) increased in the fish fed on T4 and T5 diets compared with the control group. Catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPX) were significantly (p < 0.05) decreased in the fish fed with diets contain Cu-NPs and VC (T4 and T5). The expressions of TNF-α, IL-1ß, IL-10, SOD, CAT, and GPX genes were significantly (p < 0.05) decreased in the fish fed on T3, T4, and T5 diets versus the control. In addition, the dietary Cu-NPs and VC supplementations significantly enhanced resistance against pathogens and led to the control of infection in rainbow trout. In conclusion, Cu-NPs and VC administered as feed additives at 2/250-500 mg/kg elevated the growth performance, antioxidant capacity, and health of rainbow trout.

Ruckerbactin Produced by Yersinia ruckeri YRB Is a Diastereomer of the Siderophore Trivanchrobactin Produced by Vibrio campbellii DS40M4.

The Gram-negative bacterium Yersinia ruckeri is the causative agent for enteric red mouth disease in salmonids. The genome of Y. ruckeri YRB contains a biosynthetic gene cluster encoding the biosynthesis of catechol siderophores that are diastereomeric with the known vanchrobactin class of siderophores, (DHBDArgLSer)(1-3). Ruckerbactin (1), produced by Y. ruckeri YRB, was found to be the linear tris-l-serine ester composed of l-arginine and 2,3-dihydroxybenzoic acid, (DHBLArgLSer)3. The biscatechol, (DHBLArgLSer)2 (2), and monocatechol, DHBLArgLSer (3), compounds were also isolated and characterized. The macrolactone of ruckerbactin was not detected. The presence of LArg in ruckerbactin makes it the diastereomer of trivanchrobactin with DArg. The electronic circular dichroism spectra of Fe(III)-ruckerbactin and Fe(III)-trivanchrobactin reveal the opposite enantiomeric configurations at the Fe(III) sites. Fe(III)-ruckerbactin adopts the Δ configuration, and Fe(III)-trivanchrobactin adopts the Λ configuration. Y. ruckeri YRB was also found to produce the antimicrobial agent holomycin (4).

Effect of microplastics on Yersinia ruckeri infection in rainbow trout (Oncorhynchus mykiss).

Exposure to microorganisms such as Yersinia ruckeri can significantly affect bacterial infections in fish. Microplastics (MPs) may predispose fish to infection and act as carriers in pathogen transmission. Therefore, this study is designed to evaluate MPs' effect on damage caused by exposure to Y. ruckeri in rainbow trout. In this study, blood biochemical parameters and hepatic oxidative biomarkers as clinical signs were measured in the fish co-exposed to Y. ruckeri (5 and 10% the median lethal dose (LD50)) and MPs (500 and 1000 mg Kg-1) for 30 days. There were no significant changes in the creatinine, triglyceride, cholesterol levels, and glutamic-pyruvic transaminase activity in the blood of fish infected with Y. ruckeri. In contrast, exposure to MPs had a significant effect on most clinical parameters. The total protein, albumin, globulin, total immunoglobulins, high-density lipoprotein, low-density lipoprotein, cholesterol levels, and γ-glutamyltransferase activity decreased, whereas glucose, triglyceride, and creatinine levels, and glutamic-oxaloacetic transaminase, glutamic-pyruvic transaminase, alkaline phosphatase, and lactate dehydrogenase activities increased in the plasma of fish after co-exposure to MPs and Y. ruckeri. Dietary MPs combined with a Y. ruckeri challenge decreased catalase and glutathione peroxidase activities, and total antioxidant levels. However, superoxide dismutase activity and malondialdehyde contents increased in the hepatocyte of fish co-exposed to MPs and Y. ruckeri. This study suggests that fish exposure to MPs and simultaneous challenge with Y. ruckeri could synergistically affect clinical parameters.

Effect of autochthonous lactobacilli on immunologically important molecules of rainbow trout after bacterial infection studied on intestinal primoculture.

Nowadays, the aquaculture industry is one of the fastest growing industries. Intensive aquaculture has a negative impact on fish health. Probiotic bacteria are often used due to beneficial effect to health of host, e.i. decrease of diseases outbreaks, immunomodulatory effect or better utilization of feed. The aim of this work was to study the influence of probiotic bacteria on the immune response of trout intestinal cells in primoculture infected with pathogenic bacteria. In the experiment, we tested the effect of pre-treatment of intestinal cells with an autochthonous strain of Lactobacillus plantarum R2 Biocenol™ (CCM 8674) following infection with the most serious salmonid pathogens - Aeromonas salmonicida subsp. salmonicida (CCM 1307) and Yersinia ruckeri (CCM 6093). Tested probiotic strain reduced inflammation after A. salmonicida infection through decreased expression of pro-inflammatory cytokines and increased expression of anti-inflammatory cytokines. In contrast, after infection with Y. ruckeri, which causes immunosuppression, the probiotic strain stimulated immunity by up-regulation of expression of proinflammatory cytokines and suppressed the expression of anti-inflammatory cytokines. These results are a prerequisite for the immunomodulatory potential of the strain, but its action must be confirmed in subsequent in vivo experiments.

Elucidating bacterial adhesion to mucosal surface by an original AFM approach.

BACKGROUND: Fish skin represents an ancient vertebrate mucosal surface, sharing characteristics with other mucosal surfaces including those of the intestine. The skin mucosa is continuously exposed to microbes in the surrounding water and is therefore important in the first line defense against environmental pathogens by preventing bacteria from accessing the underlying surfaces. Understanding the microbe-host interactions at the fish skin mucosa is highly relevant in order to understand and control infection, commensalism, colonization, persistence, infection, and disease. Here we investigate the interactions between the pathogenic bacteria Aeromonas salmonicida (A. salmonicida) and Yersinia ruckeri (Y. ruckeri), respectively, and the skin mucosal surface of Atlantic salmon fry using AFM force spectroscopy. RESULTS: The results obtained revealed that when retracting probes functionalized with bacteria from surfaces coated with immobilized mucins, isolated from salmon mucosal surfaces, rupture events reflecting the disruption of adhesive interactions were observed, with rupture strengths centered around 200 pN. However, when retracting probes functionalized with bacteria from the intact mucosal surface of salmon fish fry no adhesive interactions could be detected. Furthermore, rheological measurements revealed a near fluid-like behavior for the fish fry skin mucus. Taken together, the experimental data indicate that the adhesion between the mucin molecules within the mucous layer may be significantly weaker than the interaction between the bacteria and the mucin molecules. The bacteria, immobilized on the AFM probe, do bind to individual mucins in the mucosal layer, but are released from the near fluid mucus with little resistance upon retraction of the AFM probe, to which they are immobilized. CONCLUSION: The data provided in the current paper reveal that A. salmonicida and Y. ruckeri do bind to the immobilized mucins. However, when retracting the bacteria from intact mucosal surfaces, no adhesive interactions are detected. These observations suggest a mechanism underlying the protective function of the mucosal surface based on the clearing of potential threats by adhering them to loosely attached mucus that is subsequently released from the fish skin.