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A series of 6 novel ester derivatives 2–7 of natural product geodin 1 were designed and semi-synthesized through one mild step reaction with high yield. Compounds 2–7 showed strong inhibitory activities against Staphylococcus aureus in the range of 2.35–9.41 μM. Compounds 4 and 7 showed very strong inhibitory activities against antifouling bacteria Aeromonas salmonicida with MICs of 2.42 μM and 4.56 μM respectively. Most notably compounds 3–7 showed potent antifungal activities against Candida albicans in the range of 0.59–2.44 μM. Particularly, compound 3 showed the highest antifungal activity against C. albicans with a MIC value of 0.59 μM. The preliminary structure activity relationship of these derivatives showed that replacement of 4-OH group with benzoyl substituents could enhance the antibacterial and antifungal activities of geodin 1. [ABSTRACT FROM AUTHOR]

Studying inflammatory responses induced by vaccination can contribute to a more detailed understanding of underlying immune mechanisms in lumpfish (Cyclopterus lumpus). Tissue samples from lumpfish intraperitoneally immunized with a divalent oil‐adjuvanted vaccine (Aeromonas salmonicida and Vibrio salmonicida) at water temperatures of 5, 10, and 15°C were collected at 630 day degrees and 18 weeks post injection. The relative amount of secretory and membrane‐bound immunoglobulin M (IgM) gene transcripts in the head kidney was determined by qPCR. Vaccine‐induced inflammatory lesions were assessed on histological sections of abdominal pancreatic/intestinal tissue from vaccinated fish in all three temperature groups. Inflammatory cells forming dense aggregations in lesions showed proliferative activity, many of which were identified as eosinophilic‐granulocyte‐like cells. IgM+ cells were scattered in inflammatory tissue dominated by connective tissue, showing no difference in numbers between lesions from fish vaccinated at 5, 10, and 15°C. Relative gene expression analysis of secretory and membrane‐bound IgM revealed low overall expression in the head kidney of vaccinated fish at both 630 day‐degrees and 18 weeks post injection. The results of this study indicate that the vaccine stimulated prolonged local inflammatory responses at the injection site, which were not influenced by temperature. [ABSTRACT FROM AUTHOR]

Featured Application: Although this is a fundamental microbiological study, its results indicate the necessity of changing the approach to fish ponds maintenance and focusing all hygiene measures on the eradication of biofilms and not on a simple disinfection. Also, the practical laboratory approach to testing biofilms of A. salmonicida subspecies salmonicida should be changed. Biofilm formation of clinical isolates of Aeromonas salmonicida subspecies salmonicida was compared using scarce (minimal M9 and ABTG w/o amino acids) and enriched nutrient media (Tryptone Soya broth) at 8 °C, 16 °C, and 25 °C using direct enumeration of viable cells in biofilm (log CFU), crystal violet staining (ODc) of the formed biofilm biomass, and liquid–air border biofilm formation (pellicle test). Whole-genome sequencing (WGS) was performed with the usage of an Oxford nanopore system by Genomics and Transcriptomics Labor Düsseldorf (Heinrich-Heine-Universität Düsseldorf, Germany). A bioinformatic analysis was performed with the usage of Geneious Prime® 2023.0.4 (Biomatters, Inc., Boston, MA, USA). All data were analyzed using Statistica software version 13.0, and changes in biofilm production in correlation to changes in the type of nutritional medium and temperature were compared between groups using a one-way ANOVA analysis and Tukey's test. All isolates formed biofilms in minimal M9 at 8 °C and 16 °C, and nine isolates failed to form biofilms in minimal M9 at 25 °C. In an ABTG medium, all isolates produced biofilms at 8 °C; however, three isolates at 16 °C and seven isolates at 25 °C failed to form any biofilms. Significant biofilm formation was observed in TSB at all temperatures. Some strains that formed a good biofilm in solid–liquid interface did not have the ability to form a pellicle (liquid–air border biofilm), and vice versa. In all cases of nutritional medium and temperature changes, there were statistically significant differences in the intensity of biofilm production, especially in the detected number of viable cells inside biofilms (log CFU, p < 0.005). Multiple biofilm-production genes, including polar flagella (filM) LuxR family (transcriptional regulators) and VapA family of histidine-kinase-associated genes, were sequenced from all studied isolates. Genetic differences based on geographical origin were not observed among the isolates. Significant variations in isolates' ability to form biofilms were observed, possibly due to epigenetic factors. The optimal temperature for biofilm formation of A. salmonicida subspecies salmonicida in scarce media was 8 °C, and the majority of isolates were not capable of biofilm formation at 25 °C without enriched nutrient media. [ABSTRACT FROM AUTHOR]

Introduction: Piscirickettsia salmonis, the causative agent of Piscirickettsiosis, poses a significant threat to the Chilean aquaculture industry, resulting in substantial economic losses annually. The pathogen, first identified as specie in 1992, this pathogen was divided into two genogroups: LF-89 and EM-90, associated with different phenotypic mortality and pathogenicity. Traditional genotyping methods, such as multiplex PCR, are effective but limited by their cost, equipment requirements, and the need for specialized expertise. Methods: This study validates Loop-mediated Isothermal Amplification (LAMP) as a rapid and specific alternative for diagnosing P. salmonis infections. We developed the first qPCR and LAMP assay targeting the species-conserved tonB receptor gene (tonB-r, WP_016210144.1) for the specific species-level identification of P. salmonis. Additionally, we designed two genotyping LAMP assays to differentiate between the LF-89 and EM-90 genogroups, utilizing the unique coding sequences Nitronate monooxygenase (WP_144420689.1) for LF-89 and Acid phosphatase (WP_016210154.1) for EM-90. Results: The LAMP assays demonstrated sensitivity and specificity comparable to real-time PCR, with additional benefits including rapid results, lower costs, and simplified operation, making them particularly suitable for field use. Specificity was confirmed by testing against other salmonid pathogens, such as Renibacterium salmoninarum, Vibrio ordalii, Flavobacterium psychrophilum, Tenacibaculum maritimum, and Aeromonas salmonicida, with no cross-reactivity observed. Discussion: The visual detection method and precise differentiation between genogroups underscore LAMP's potential as a robust diagnostic tool for aquaculture. This advancement in the specie detection (qPCR and LAMP) and genotyping of P. salmonis represents a significant step forward in disease management within the aquaculture industry. The implementation of LAMP promises enhanced disease surveillance, early detection, and improved management strategies, ultimately benefiting the salmonid aquaculture sector. [ABSTRACT FROM AUTHOR]

Background/Objectives: Antibiotic resistance gene (ARG) spread is driven by horizontal gene transfer (HGT). Ciliated protozoa may contribute to this process, as their predation has been shown to facilitate HGT in certain bacteria. Here, this phenomenon was further investigated using A. salmonicida subsp. salmonicida. This fish pathogen bears an extensive and dynamic plasmidome, suggesting a high potential for HGT. Methods: A. salmonicida strains carrying one of three conjugative plasmids bearing ARGs (pSN254b, pRAS1b or pAsa4b) were cocultured with a recipient, either A. salmonicida, E. coli or A. hydrophila. Conjugation rates were assessed in the presence and absence of the ciliate Tetrahymena borealis. PCR genotyping confirmed the acquisition of the conjugative plasmids and was used to verify the mobilization of other plasmids. Results: The basal rate of conjugation observed was high. Under the conditions studied, ciliate predation did not appear to influence the conjugation rate, except at higher proportions of ciliates, which typically hampered conjugation. Microscopy revealed that most bacteria were digested in these conditions. PCR screening demonstrated that small mobilizable plasmids from A. salmonicida (pAsa1, pAsa2, pAsa3, and pAsal1) were acquired by the recipients along with the conjugative plasmids, with a slight effect of the ciliates in some donor/recipient cell combination. Conclusions: These results highlight how A. salmonicida can conjugate efficiently with different species and how complex its relationship with ciliates is. [ABSTRACT FROM AUTHOR]

Outbreaks of furunculosis cause significant losses in salmonid aquaculture worldwide. With a recent rise in antimicrobial resistance, regulatory measures to minimize the use of antibiotics in animal husbandry, including aquaculture, have increased scrutiny and availability of veterinary medical products to control this disease in production facilities. In such a regulatory environment, the utility of autogenous vaccines to assist with disease prevention and control as a veterinary-guided prophylactic measure is of high interest to the producers and veterinary services alike. However, evolving concepts of epidemiological units and epidemiological links need to be considered during approval and acceptance procedures for the application of autogenous vaccines in multiple aquaculture facilities. Here, we present the results of solid-state nanopore sequencing (Oxford Nanopore Technologies, ONT) performed on 54 isolates of Aeromonas salmonicida ssp. salmonicida sampled during clinical outbreaks of furunculosis in different aquaculture facilities from Bavaria, Germany, from 2017 to 2020. All of the performed analyses (phylogeny, single nucleotide polymorphism and 3D protein modeling for major immunogenic proteins) support a high probability that all studied isolates belong to the same epidemiological unit. Simultaneously, we describe a cost/effective method of whole genome analysis with the usage of ONT as a viable strategy to study outbreaks of other pathogens in the field of aquatic veterinary medicine for the purpose of developing the best autogenous vaccine candidates applicable to multiple aquaculture establishments. [ABSTRACT FROM AUTHOR]

Aeromonas salmonicida is a major pathogenic bacterium that poses a significant threat to salmonid fish. Yadong County, located in the Xizang Autonomous Region, is renowned for its characteristic industry of Salmo trutta aquaculture. In recent years, the outbreak of Bacterial Gill Disease (BGD) has led to substantial economic losses for S. trutta farmers. Our prior research identified A. salmonicida as one of the primary culprits behind BGD. To mitigate the impact of A. salmonicida on S. trutta, we conducted a comprehensive study aimed at identifying genes associated with resistance to A. salmonicida. This involved transcriptome sequencing and 16S rRNA sequencing of intestinal flora, providing valuable insights for the study of disease resistance in S. trutta. In this study, we identified 324 genera with 5171 ASVs in the susceptible group and 293 genera with 5669 ASVs in the resistant group. Notably, Methylobacterium and Sphingomonas were common bacteria present in the salmon's gut, and their proportions remained relatively stable before and after infection. Shewanella, with its antagonistic relationship with Aeromonas, may play a crucial role in the salmon's defense against A. salmonicida. Several related genes were identified, including angptl4, cipcb, grasp, ccr9a, sulf1, mtmr11, B3GNT3, mt2, PLXDC1, and ank1b. [ABSTRACT FROM AUTHOR]

Background: Accurate identification of infectious diseases using molecular techniques, such as PCR and NGS, is well-established. This study aims to assess the utility of Bactfast and Fungifast in diagnosing bloodstream infections in ICU settings, comparing them against traditional culture methods. The objectives include evaluating sensitivity and specificity and identifying a wide range of pathogens, including non-culturable species. Methods: We collected 500 non-duplicate blood samples from ICU patients between January 2023 and December 2023. Specimens underwent traditional culture, MALDI-TOF, VITEK®2 compact system, and NGS-based Bactfast and Fungifast analyses. Results: Out of the 500 samples, 26.8% (n=134) showed bacterial growth via traditional culture methods, while 4.8% (n=24) were positive for fungal growth. MALDI-TOF and VITEK®2 compact system yielded comparable results, identifying 26.4% (n=132) of specimens with bacterial growth. NGS-based Bactfast detected bacterial presence in 38.2% (n=191) of samples, including non-culturable bacteria missed by traditional methods. However, NGS-based Fungifast showed concordant fungal detection rates with culture methods. Among identified pathogens by culture method included Klebsiella pneumoniae 20.89% (n=28), Enterococcus faecalis 18.65% (n=25), Escherichia coli 15.67% (n=21), Pseudomonas aeruginosa 12.68% (n=17), Acinetobacter baumannii 10.44% (n=14), various Streptococcus species 7.46% (n=10), Mycobacterium tuberculosis 6.71% (n=9), Mycobacterium abscessus 4.47% (n=6), and Salmonella spp 2.98% (n=4). Non-culture-based NGS identified additional (n=33) pathogens, including Klebsiella pneumoniae 27.27% (n=9), Bacteroides fragilis 21.21% (n=7), Aerococcus viridans 15.15% (n=5), Elizabethkingia anopheles 12.12% (n=4), Aeromonas salmonicida 9% (n=3), Clostridium 9% (n=3), and Bacteroides vulgatus 6% (n=2). Candida albicans was reported in 5% (n=24) of samples by both methods. Conclusion: NGS-based Bactfast and Fungifast demonstrate high sensitivity in identifying a wide array of bacterial and fungal pathogens in ICU patients, outperforming traditional culture methods in detecting non-culturable organisms. These molecular assays offer rapid and comprehensive diagnostic capabilities, potentially improving clinical outcomes through timely and accurate pathogen identification. [ABSTRACT FROM AUTHOR]

Infections pose a challenge for the fast growing aquaculture sector. Glycosphingolipids are cell membrane components that pathogens utilize for attachment to the host to initiate infection. Here, we characterized rainbow trout glycosphingolipids from five mucosal tissues using mass spectrometry and nuclear magnetic resonance and investigated binding of radiolabeled Aeromonas salmonicida to the glycosphingolipids on thin-layer chromatograms. 12 neutral and 14 acidic glycosphingolipids were identified. The glycosphingolipids isolated from the stomach and intestine were mainly neutral, whereas glycosphingolipids isolated from the skin, gills and pyloric caeca were largely acidic. Many of the acidic structures were poly-sialylated with shorter glycan structures in the skin compared to the other tissues. The sialic acids found were Neu5Ac and Neu5Gc. Most of the glycosphingolipids had isoglobo and ganglio core chains, or a combination of these. The epitopes on the rainbow trout glycosphingolipid glycans differed between epithelial sites leading to differences in pathogen binding. A major terminal epitope was fucose, that occurred attached to GalNAc in a α1-3 linkage but also in the form of HexNAc-(Fuc-)HexNAc-R. A. salmonicida were shown to bind to neutral glycosphingolipids from the gill and intestine. This study is the first to do a comprehensive investigation of the rainbow trout glycosphingolipids and analyze binding of A. salmonicida to glycosphingolipids. The structural information paves the way for identification of ways of interfering in pathogen colonization processes to protect against infections in aquaculture and contributes towards understanding A. salmonicida infection mechanisms. [ABSTRACT FROM AUTHOR]

This research aimed to investigate the effects of polysaccharide from Atractylodes macrocephala koidze (PAMK) on growth performance, antioxidant capacity, immune response, and resistance of rainbow trout (Oncorhynchus mykiss) to Aeromonas salmonicida infection. Fish (average weight 58.40 ± 1.10 g) were fed four concentrations (0, 400, 800, and 1600 mg/kg diet) of PAMK in 4 treatments with 3 replicates for 8 weeks. At the end of the feeding trial, the fish were infected with A. salmonicida for 7 days. Results revealed that fish fed diets supplemented with 800 mg/kg PAMK exhibited enhancing growth performance and feed utilization compared to those in the other groups (p < 0.05), whereas differences were not found in body composition among the groups (p > 0.05). The serum superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), lysozyme (LZM), alkaline phosphatase (AKP), acid phosphatase (ACP), total protein (TP), and albumin (ALB) contents were significantly greater in the 800 mg/kg PAMK group than in the control group (p < 0.05). Before and after infection, dietary PAMK upregulated the mRNA expression of anti-inflammatory cytokines (TGF-β, IL-4/13A and IL-10), complement components (C3 and C4) and immunoglobulin M (IgM) but downregulated the expression of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6); these parameters significantly changed in the 800 mg/kg PAMK group (p < 0.05). The highest survival rate was observed in the 800 mg/kg PAMK group. These results collectively suggest that 800 mg/kg PAMK can be used to enhance the growth performance, antioxidant activity, immune response, and disease resistance of rainbow trout. [ABSTRACT FROM AUTHOR]

Aquaculture is expected to play a vital role in solving the challenge of sustainably providing the growing world population with healthy and nutritious food. Pathogen outbreaks are a major risk for the sector, so early detection and a timely response are crucial. This can be enabled by monitoring the pathogen levels in aquaculture facilities. This paper describes a photonic biosensing platform based on silicon nitride waveguide technology with integrated active components, which could be used for such applications. Compared to the state of the art, the current system presents improvements in terms of miniaturization of the Photonic Integrated Circuit (PIC) and the development of wafer-level processes for hybrid integration of active components and for material-selective chemical and biological surface modification. Furthermore, scalable processes for integrating the PIC in a microfluidic cartridge were developed, as well as a prototype desktop readout instrument. Three bacterial aquaculture pathogens (Aeromonas salmonicida, Vagococcus salmoninarum, and Yersinia ruckeri) were selected for assay development. DNA biomarkers were identified, corresponding primer-probe sets designed, and qPCR assays developed. The biomarker for Aeromonas was also detected using the hybrid PIC platform. This is the first successful demonstration of biosensing on the hybrid PIC platform. [ABSTRACT FROM AUTHOR]

The diversity of O-polysaccharides (O-antigens) among 28 Aeromonas salmonicida strains isolated from ill fish has been determined by using high-resolution magic angle spinning (HR MAS) NMR spectroscopy. The new O-polysaccharide has been identified in two isolates. This new structure was investigated by 1H and 13C NMR spectroscopy and matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The following structure of the linear hexasaccharide repeating unit of A. salmonicida O-antigen has been established: →3)-α-L-Rhap-(1→3)-α-D-ManpNAc-(1→2)-β-D-Glcp-(1→3)-α-L-Rhap2OAc4OAc-(1→3)-β-D-ManpNAc-(1→3)-α-D-Glcp-(1→. This new A. salmonicida O-polysaccharide was detected among two isolates collected from trout and turbot fish in 2010 and 2011, respectively. Further investigations should be conducted to evaluate the distribution of this new O-polysaccharide among a larger collection of isolates, depending on their geographic origin, the species of fish, and the health status of the fish. [ABSTRACT FROM AUTHOR]

Hsp40–Hsp70 typically function in concert as molecular chaperones, and their roles in post-infection immune responses are increasingly recognized. However, in the economically important fish species Scophthalmus maximus (turbot), there is still a lack in the systematic identification, interaction models, and binding site analysis of these proteins. Herein, 62 Hsp40 genes and 16 Hsp70 genes were identified in the turbot at a genome-wide level and were unevenly distributed on 22 chromosomes through chromosomal distribution analysis. Phylogenetic and syntenic analysis provided strong evidence in supporting the orthologies and paralogies of these HSPs. Protein–protein interaction and expression analysis was conducted to predict the expression profile after challenging with Aeromonas salmonicida. dnajb1b and hspa1a were found to have a co-expression trend under infection stresses. Molecular docking was performed using Auto-Dock Tool and PyMOL for this pair of chaperone proteins. It was discovered that in addition to the interaction sites in the J domain, the carboxyl-terminal domain of Hsp40 also plays a crucial role in its interaction with Hsp70. This is important for the mechanistic understanding of the Hsp40–Hsp70 chaperone system, providing a theoretical basis for turbot disease resistance breeding, and effective value for the prevention of certain diseases in turbot. [ABSTRACT FROM AUTHOR]

Infectious disease is a major challenge in fish and shrimp farming systems caused by microorganisms including bacteria, viruses and parasites. Physical detection has limitations in detecting diseases in cultivated animals earlier, because of the varied clinical symptoms. The aims of this study was to identify the physical and molecular presence of infectious pathogens in fish and shrimp cultivated in several ponds in West Java, Banten and Jakarta. Monitoring was carried out in two stages, namely the period March–April 2021 and September 2021 by taking samples from several ponds in Bogor, Tangerang, Depok and Jakarta. The sample criteria used were fish and shrimp showing normal and clinical symptoms of being attacked by a disease which were collected by purposive sampling. The collected samples were examined in two stages, namely physical examination of clinical symptoms and molecular examination using PCR. The results showed that of the 24 species samples collected, 17 species showed normal clinical symptoms and seven species showed clinical symptoms. Of the 24 fish and shrimp samples, four samples were infected (2 samples from normal species and 2 samples with clinical symptoms). The infected normal samples were Osphronemus goramy and Litopenaeus vannamei. The infected samples with clinical symptoms were Carassius auratus by Aeromonas salmonicida and red zebra cichlid (Metriaclima estherae) by Red Sea Bream Iridoviral Disease (RSBIV). Physical testing supported by molecular detection of aquatic animals can be an effort to manage aquaculture systems in Indonesia. [ABSTRACT FROM AUTHOR]

Aeromonas salmonicida is a common pathogen of salmonid fishes that poses a significant threat to the fresh water and marine culture industry, potentially resulting in huge economic losses. To prevent and control fish diseases caused by A. salmonicida, rapid and effective diagnostic approaches must be developed, and which are important for routine monitoring and clinical care. By combining recombinase polymerase amplification (RPA) technology with a visible lateral flow strip (RPA-LF), we have enhanced both the precision of RPA detection and the convenience of real-time monitoring. In this study, we introduce a robust method for detecting A. salmonicida using RPA-LF. This assay specifically targets the ASA_1441 gene of A. salmonicida, ensuring high specificity, without cross-reactivity with other prevalent fresh water or marine pathogens. The optimal amplification temperature of the RPA assay was 39 °C. Its sensitivity extends to as low as 100 fg of purified DNA, representing more than 1000-fold higher sensitivity than conventional PCR methods. Furthermore, to enhance the usability of the RPA-LF assay, we developed a rapid sample preparation method using cellulose dipsticks for nucleic acid extraction. This method achieves a limit of detection (LOD) as low as 1.67 CFU/µL and completes the entire process within 20 min. In conclusion, our findings present a rapid and precise tool for monitoring A. salmonicida infection in aquaculture and marine culture. This advancement offers valuable insights for effective disease prevention and control strategies. [ABSTRACT FROM AUTHOR]

Aeromonas salmonicida, a widely distributed aquatic pathogen causing furunculosis in fish, exhibits varied virulence, posing challenges in infectious disease and immunity studies, notably in vaccine efficacy assessment. Lumpfish (Cyclopterus lumpus) has become a valuable model for marine pathogenesis studies. This study evaluated several antigen preparations against A. salmonicida J223, a hypervirulent strain of teleost fish, including lumpfish. The potential immune protective effect of A. salmonicida bacterins in the presence and absence of the A‐layer and extracellular products was tested in lumpfish. Also, we evaluated the impact of A. salmonicida outer membrane proteins (OMPs) and iron‐regulated outer membrane proteins (IROMPs) on lumpfish immunity. The immunized lumpfish were intraperitoneally (i.p.) challenged with 104A. salmonicida cells/dose at 8 weeks‐post immunization (wpi). Immunized and non‐immunized fish died within 2 weeks post‐challenge. Our analyses showed that immunization with A. salmonicida J223 bacterins and antigen preparations did not increase IgM titres. In addition, adaptive immunity biomarker genes (e.g., igm, mhc‐ii and cd4) were down‐regulated. These findings suggest that A. salmonicida J223 antigen preparations hinder lumpfish immunity. Notably, many fish vaccines are bacterin‐based, often lacking efficacy evaluation. This study offers crucial insights for finfish vaccine approval and regulations. [ABSTRACT FROM AUTHOR]

The misuse of antibiotics and the emergence of antimicrobial resistance (AMR) is a concern in the aquaculture industry because it contributes to global health risks and impacts the environment. This study analyzed the AMR of sentinel bacteria associated with striped catfish (Pangasisanodon hypophthalmus) and giant snakehead (Channa micropeltes), the two main fish species reared in the pond culture in Cambodia. Phenotypic and genotypic characterization of the recovered isolates from fish, water, and sediment samples revealed the presence of bacteria, such as 22 species belonging to families Aeromonadaceae, Enterobacteriaceae, and Pseudomonadaceae. Among 48 isolates, Aeromonas caviae (n = 2), Aeromonas hydrophila (n = 2), Aeromonas ichthiosmia (n = 1), Aeromonas salmonicida (n = 4) were detected. A. salmonicida and A. hydrophilla are known as fish pathogens that occur worldwide in both fresh and marine water aquaculture. Antibiotic susceptibility testing revealed antibiotic resistance patterns of 24 (50 %) isolates among 48 isolates with higher multiple antibiotic resistance index (> 0.2). All the isolates of Enterobacteriaceae were susceptible to ciprofloxacin. Ciprofloxacin is a frontline antibiotic that is not recommended to use in aquaculture. Therefore, its use has to be strictly controlled. This study expands our knowledge of the AMR status in aquaculture farms which is very limited in Cambodia. [ABSTRACT FROM AUTHOR]

This study evaluates the effects of dietary Clostridium butyricum on growth performance and intestinal health in triploid rainbow trout (Oncorhynchus mykiss). Administered in a 12-week trial, five isonitrogenous and isolipidic feeds contained different C. butyricum levels [G1 (0), G2 (1.6 × 106 CFU/g), G3 (1.2 × 107 CFU/g), G4 (1.1 × 108 CFU/g), and G5 (1.3 × 109 CFU/g)]. Significant enhancements in growth performance, including improved feed conversion ratios and specific growth rates in the G4 group, were indicative of enhanced nutrient utilization, corroborated by optimal digestive enzyme activity levels. Antioxidant capabilities were also enhanced in the G4 group, indicated by increased serum superoxide dismutase and catalase activities, along with a significant decrease in malondialdehyde levels. Gut microbiota analysis indicated a probiotic concentration-dependent modulation of microbial communities, with a marked enrichment in beneficial bacterial phyla like Firmicutes in the G4 group. This microbial shift correlated with significant downregulations in immune-related gene expressions, including interleukins (IL-1β, IL-8), NF-κB, MyD88, and TNF-α, highlighting an activated immune response. Correspondingly, serum lysozyme and immunoglobulin M contents were significantly elevated in the G4 group. Challenge tests with Aeromonas salmonicida exhibited the higher disease resistance of fish fed the G4 diet. In conclusion, the study shows the potential of dietary C. butyricum, especially at 1.1 × 108 CFU/g, in enhancing the growth, health, and disease resistance of triploid rainbow trout through modulating gut microflora and stimulating immune responses. [ABSTRACT FROM AUTHOR]

Lactic acid bacteria Ligilactobacillus salivarius has been shown to be a substitute for antibiotics in the treatment of bacterial disease in high animals. However, its beneficial mechanism in fish farming is still unclear. This study evaluated the antagonistic effects of the Ligilactobacillus salivarius GX118 strain on Aeromonas salmonicida and its regulation of rainbow trout growth in vivo and in vitro. The results found that GX118 produces an antibacterial substance that can directly destroy the cell wall of A. salmonicida. Whole-genome sequencing of GX118 revealed that Enterolysin A is a type III bacteriocin with antibacterial properties. An in vivo experiment showed that the supplementation of GX118 in diet competitively inhibited the colonization of A. salmonicida in the intestine. In addition, it was able to improve the growth performance of rainbow trout within a 21-day feeding experiment. The supplementation of GX118 increased the diversity of gut microbiota, in which the abundance of Bacteroidota, Blautia, and Rhodobacteraceae increased. In addition, the use of GX118 activated the expression of IFN-γ and NF-κB genes and reduced the expression level of IL-6 and IL-8, thus exhibiting a certain effect on activating the immunity of rainbow trout. This study provides a scientific basis for the development of antibacterial probiotics in the healthy farming of rainbow trout. [ABSTRACT FROM AUTHOR]

This study evaluated the effects of oregano essential oil (OEO)-meditated intestinal microbiota and immunity on growth, amino acid composition, and Aeromonas salmonicida resistance in rainbow trout. A total of 480 fish were allocated to four feeding treatments: dietary supplementation with 2.0 mL OEO/kg (group L), 4.0 mL OEO/kg (group M), 8.0 mL OEO/kg (group H), and no supplementation as control group (CK group). The results indicated that dietary OEO had a positive impact on the growth performance and pathogen resistance of rainbow trout, with higher amino acid deposition in the muscle and, in particular, a significant enrichment in amino acid pathways was observed. The use of OEO led to an increase in abundance of Proteobacteria and decrease in abundance of Firmicutes in intestinal microbiota. Moreover, the abundance of taxa containing beneficial bacteria such as Streptococcus lutetiensis, Oscillospirales, and Methylobacterium-Methylorubrum increased, while the abundance of taxa containing potential pathogens such as Mycoplasma, Morganellaceae, and Paenalcaligenes decreased in OEO treatment groups. In addition, transcriptome analysis showed that a number of immune-related genes such as TLR3, MyD88, IRF7, and TRAF2 were up-regulated, while il-6, il-21, and il-34 were down-regulated. Thus, OEO intervention contributed to the amino acid deposition and resistance to A. salmonicida infection in rainbow trout by mediating the link between fish immunity and intestinal microbiota function. [ABSTRACT FROM AUTHOR]

The Aeromonas salmonicida is responsible for causing furunculosis in various fish species. Furunculosis is a ubiquitous disease that affects the aquaculture industry and causes the mass mortality of turbot. Vibrio vulnificus is a pathogen that causes skin ulcers and hemorrhagic septicemia in fish, resulting in significant mortality in aquaculture. In this study, we have established a bivalent inactivated vaccine against A. salmonicida and V. vulnificus with Montanide™ ISA 763 AVG as an adjuvant. This bivalent inactivated vaccine was used to immunize turbot by intraperitoneal injection, and the relevant immune indexes were detected. The results demonstrate that the bivalent inactivated vaccine exhibited a relative percent survival (RPS) of 77% following A. salmonicida and V. vulnificus intraperitoneal challenge. The vaccinated group exhibited higher levels of acid phosphatase activity and lysozyme activity compared to the control group. ELISA results showed a significant increase in serum antibody levels in immunized turbot, which was positively correlated with immunity. In the kidney tissue, related immune genes (TLR5, CD4, MHCI and MHCII) were up-regulated significantly, showing that the vaccine can induce cellular and humoral immune responses in turbot. In conclusion, the bivalent inactivated vaccine against A. salmonicida and V. vulnificus was immunogenic, efficiently preventing turbot from infection, which has the potential to be applied in aquaculture. [ABSTRACT FROM AUTHOR]

Aeromonas salmonicida is an important pathogen that causes furunculosis in trout and salmon with high morbidity and mortality, resulting in significant economic losses in aquaculture. Overuse of antibiotics has led to the continuous emergence of drug-resistant strains. Hence, there is an urgent need to find an alternative environmentally friendly antimicrobial agent. In this study, we isolated a virulent phage of A. salmonicida, named ASG01, which belongs to the Myoviridae family and maintains lytic activity at a pH value range from 4 to 12 and in the temperature range from 30 °C to 60 °C. The whole genomic sequence of ASG01 showed 82% similarity to Aeromonas phage pAh6-C. The cell wall hydrolase (Cwh)-encoding gene from the genome of ASG01 was predicted and heterologously expressed. Notably, in the absence of additional phage genes, endogenous expression of Cwh could lyse E. coli cells and greatly inhibit the growth of tested fish pathogenic bacteria. The lytic activity of Cwh was eliminated when the predicted active site was mutated. These results indicate that Cwh of ASG01 possessed excellent lytic activity and a wide antibacterial spectrum, suggesting its potential as an effective enzybiotic. [ABSTRACT FROM AUTHOR]

Representatives of the bacterial genus Aeromonas are some of the most notorious aquaculture pathogens associated with a range of diseases in different fish species. As the world forges toward the post-antibiotic era, alternative options for combating bacterial pathogens are needed. One such alternative option is phage biocontrol. In this study, a novel podophage—JELG-KS1—infecting Aeromonas salmonicida was retrieved from wastewater along with its host strain. The genome of the JELG-KS1 phage is a 40,505 bp dsDNA molecule with a GC% of 53.42% and 185 bp direct terminal repeats and encodes 53 predicted proteins. Genomic analysis indicates that JELG-KS1 might represent a novel genus within the subfamily Studiervirinae. Podophage JELG-KS1 is a strictly lytic phage without any identifiable virulence or AMR genes that quickly adsorbs onto the surface of host cells to initiate a 48 min long infectious cycle, resulting in the release of 71 ± 12 JELG-KS1 progeny virions per infected cell. JELG-KS1 effectively lyses its host population in vitro, even at very low multiplicities of infection. However, when challenged against a panel of Aeromonas spp. strains associated with diseases in aquaculture, JELG-KS1 shows host-specificity that is confined only to its isolation strain, immediately compromising its potential for Aeromonas spp. biocontrol in aquaculture. [ABSTRACT FROM AUTHOR]

17α-ethinylestradiol (EE2), a synthetically derived analogue of endogenous estrogen, is widely employed as a hormonal contraceptive and is recognized as a highly hazardous emerging pollutant, causing acute and chronic toxic effects on both aquatic and terrestrial organisms. It has been included in the initial Water Watch List. The aim of this study was to isolate bacteria from consortia recovered from mine sediments and acid mine drainage samples, both considered extreme environments, with the ability to degrade EE2. From the most promising consortia, isolates affiliated with the Aeromonas, Rhizobium, and Paraburkholderia genera were obtained, demonstrating the capability of growing at 50 mg/L EE2. Subsequently, these isolates were tested with 9 mg/L of EE2 as the sole carbon source. Among the isolated strains, Aeromonas salmonicida MLN-TP7 exhibited the best performance, efficiently degrading EE2 (95 ± 8%) and reaching concentrations of this compound below the limits of detection within 7 and 9 days. The final metabolites obtained are in accordance with those of the TCA cycle; this may indicate EE2 mineralization. As far as is known, Aeromonas salmonicida was isolated for the first time and identified in acid mine drainage, demonstrating its capacity to degrade EE2, making it a promising candidate for bioaugmentation and suggesting its possible applicability in low pH environments. [ABSTRACT FROM AUTHOR]

Given the dual roles in immune function and metabolism, the liver emerge as a compelling candidate to bridge host defense and metabolic adjustments during pathogen infections in teleosts. In order to dissect the roles of the liver in the immune response of Sebastes schlegelii, we conducted an investigation into the activities of SOD, CAT and GPX4, performed a systematic analysis of circRNA, miRNA and mRNA expression profiles, as well as constructed circRNA-miRNA-mRNA regulatory networks in the liver of S. schlegelii following Aeromonas salmonicida infection were performed in the present study. The present results demonstrated the content of SOD, CAT and GPX4 increased significantly at early infection stage to protect the liver tissue from excessive damage. Additionally, 622 circRNA-miRNAs pairs, 78 miRNA-mRNA pairs and 327 circRNA-miRNA-mRNA pairs were identified in our study. These differently expressed circRNAs and miRNAs were found to be associated with the regulation of LMNB1, DMBT1, NAMPT, IFIT1, CELSRs, PYGL. GO and KEGG enrichment analysis revealed that these differently expressed genes were involved in TLR signal pathway, RLRs signal pathway, PPAR signal pathway, shedding light on antibacterial ceRNA networks within the liver of S. schlegelii post A. hydrophila post infection, which provides new insights into the immune mechanisms of teleost.

Background/Objectives: Antibiotic resistance gene (ARG) spread is driven by horizontal gene transfer (HGT). Ciliated protozoa may contribute to this process, as their predation has been shown to facilitate HGT in certain bacteria. Here, this phenomenon was further investigated using A. salmonicida subsp. salmonicida. This fish pathogen bears an extensive and dynamic plasmidome, suggesting a high potential for HGT. Methods: A. salmonicida strains carrying one of three conjugative plasmids bearing ARGs (pSN254b, pRAS1b or pAsa4b) were cocultured with a recipient, either A. salmonicida, E. coli or A. hydrophila. Conjugation rates were assessed in the presence and absence of the ciliate Tetrahymena borealis. PCR genotyping confirmed the acquisition of the conjugative plasmids and was used to verify the mobilization of other plasmids. Results: The basal rate of conjugation observed was high. Under the conditions studied, ciliate predation did not appear to influence the conjugation rate, except at higher proportions of ciliates, which typically hampered conjugation. Microscopy revealed that most bacteria were digested in these conditions. PCR screening demonstrated that small mobilizable plasmids from A. salmonicida (pAsa1, pAsa2, pAsa3, and pAsal1) were acquired by the recipients along with the conjugative plasmids, with a slight effect of the ciliates in some donor/recipient cell combination. Conclusions: These results highlight how A. salmonicida can conjugate efficiently with different species and how complex its relationship with ciliates is.

Aeromonas salmonicida is a major pathogenic bacterium that poses a significant threat to salmonid fish. Yadong County, located in the Xizang Autonomous Region, is renowned for its characteristic industry of Salmo trutta aquaculture. In recent years, the outbreak of Bacterial Gill Disease (BGD) has led to substantial economic losses for S. trutta farmers. Our prior research identified A. salmonicida as one of the primary culprits behind BGD. To mitigate the impact of A. salmonicida on S. trutta, we conducted a comprehensive study aimed at identifying genes associated with resistance to A. salmonicida. This involved transcriptome sequencing and 16S rRNA sequencing of intestinal flora, providing valuable insights for the study of disease resistance in S. trutta. In this study, we identified 324 genera with 5171 ASVs in the susceptible group and 293 genera with 5669 ASVs in the resistant group. Notably, Methylobacterium and Sphingomonas were common bacteria present in the salmon’s gut, and their proportions remained relatively stable before and after infection. Shewanella, with its antagonistic relationship with Aeromonas, may play a crucial role in the salmon’s defense against A. salmonicida. Several related genes were identified, including angptl4, cipcb, grasp, ccr9a, sulf1, mtmr11, B3GNT3, mt2, PLXDC1, and ank1b.

Here, we provide evidence that the freshwater parasitic copepod, Salmincola californiensis, acts as a vector for Aeromonas salmonicida. While investigating the effects of S. californiensis on Chinoook salmon (Oncorhynchus tshawytscha), we tangentially observed that fish infected with the copepod developed furunculosis, caused by A. salmonicida. This occurred despite being reared in pathogen‐free well water in a research facility with no prior history of spontaneous infection. We further investigated the possibility of S. californiensis to serve as a vector for the bacterium via detection of fluorescently labelled A. salmonicida inside the egg sacs from copepods in which the fish hosts were experimentally infected with GFP‐A449 A. salmonicida. We then evaluated copepod egg sacs that were collected from adult Chinook salmon from a freshwater hatchery with A. salmonicida infections confirmed by either culture or PCR. The bacterium was cultured on tryptic soy agar plates from 75% of the egg sacs, and 61% were positive by PCR. These three separate experiments indicate an alternative tactic of transmission in addition to direct transmission of A. salmonicida in captivity. The copepod may play an important role in transmission of the bacterium when fish are more dispersed, such as in the wild. [ABSTRACT FROM AUTHOR]

Furunculosis, caused by Aeromonas salmonicida, poses a significant threat to both salmonid and non-salmonid fish in diverse aquatic environments. This study explores the genomic intricacies of re-emergent A. salmonicida outbreaks in Atlantic salmon (Salmo salar). Previous clinical cases have exhibited pathological characteristics, such as periorbital hemorrhages and gastrointestinal abnormalities. Genomic sequencing of three Chilean isolates (ASA04, ASA05, and CIBA_5017) and 25 previously described genomes determined the pan-genome, phylogenomics, insertion sequences, and restriction-modification systems. Unique gene families have contributed to an improved understanding of the psychrophilic and mesophilic clades, while phylogenomic analysis has been used to identify mesophilic and psychrophilic strains, thereby further differentiating between typical and atypical psychrophilic isolates. Diverse insertion sequences and restriction-modification patterns have highlighted genomic structural differences, and virulence factor predictions can emphasize exotoxin disparities, especially between psychrophilic and mesophilic strains. Thus, a novel plasmid was characterized which emphasized the role of plasmids in virulence and antibiotic resistance. The analysis of antibiotic resistance factors revealed resistance against various drug classes in Chilean strains. Overall, this study elucidates the genomic dynamics of re-emergent A. salmonicida and provides novel insights into their virulence, antibiotic resistance, and population structure. [ABSTRACT FROM AUTHOR]

Sustainable ecosystem is of essence to the river for its survival and maintenance of ecological balance. Gomti river, a groundwater-fed tributary of the Holy Ganges, is the lifeline of the middle Indo-Gangetic Plains. Screening of river pollutants for river cleanup is a challenging task. To this end, 20 km Lucknow stretch of Gomti was sampled at 5 locations for physicochemical–bacteriological analysis. Collected samples were analyzed for pH, TA, turbidity, DO, BOD, TH, COD, TDS, fecal coliform, and E. coli. Analytical instruments AAS and GC-ECD were used for heavy metal and organochlorine pesticide analysis. DO was 3.2 ± 0.3 to 7.4 ± 0.4 (avg 5 ± 1.8) mg/l, BOD was 10.6 ± 0.5 to 41 ± 1.7 (avg 22.9 ± 13.3) mg/l, while COD was 26.6 ± 4.6 to 93.5 ± 4.6 (avg 56 ± 31.3) mg/l. WQI values gradually increased from S1 (56.7) to S5 (181.8) which denotes a significant deterioration of water quality downstream of the river. Cd, Mn, Fe, Pb, and Cr had increment of 64, 18, 12, 6, and 2 fold, respectively, as compared to individual BIS:10500 standards. Among the OCPs, concentrations were in the order: aldrin > pp-DDT > β-HCH > γ-HCH > α-endosulfan > pp-DDE > α-HCH. Statistically strong positive correlation (r > 0.95 at 0.01) was observed among physicochemical parameters. Furthermore, bacterial species Aeromonas salmonicida subsp. Achromogenes, Aeromonas salmonicida strain A527, Aeromonas encheleia strain NCTC12917, and Aeromonas salmonicida strain ATCC33658 were identified by 16S rRNA gene amplification and molecular phylogenetic analysis. The study indicated heavy contamination of the river with organic and inorganic pollutants owing to mixing of untreated sewage, industrial effluents, and associated toxicants along with pathogenic bacteria which are a barrier to the harmony of aquatic life. [ABSTRACT FROM AUTHOR]

Rainbow trout, one of the most economically important aquaculture fish species worldwide, is affected by the pathogenic bacteria A. salmonicida, which causes furunculosis outbreaks, leading to huge economic losses. In this study, an oligonucleotide microarray was applied to identify transcriptional changes in the skin of rainbow trout individuals in response to a bacterial infection. Overall, 656 and 434 differentially expressed genes (DEGs) were identified at 2 and 6 days after a bacterial challenge (dpi), respectively. A comparison of moribund (2 dpi) and survivor fish (6 dpi) revealed 169 DEGs. Between these were many genes involved in immune response, including lysozymes, pattern recognition receptors (c-type lectins), antimicrobial peptides (cathelicidin and hepcidin), acute-phase proteins (serum amyloids and haptoglobin), complement cascade proteins (c3, c4, c6 and c7), interleukins (il11 and il1b) and chemokines (ccl19 and cxcl8). Alterations of leptin, eicosanoids and prostaglandins have been found, which suggest metabolic remodeling in conjunction with immune response. Further, the regulation of programmed cell death genes (caspase 8, bcl2 apoptosis regulator, nfkb inhibitor alpha and heme oxygenase) and structural proteins (collagens, myosins, keratins and metalloproteinases) was observed. This study provides, for the first time, a gene expression analysis of rainbow trout skin in response to A. salmonicida infection, revealing the complexity of defense strategies in response to furunculosis. [ABSTRACT FROM AUTHOR]

The diversity of O-polysaccharides (O-antigens) among 28 Aeromonas salmonicida strains isolated from ill fish has been determined by using high-resolution magic angle spinning (HR MAS) NMR spectroscopy. The new O-polysaccharide has been identified in two isolates. This new structure was investigated by 1H and 13C NMR spectroscopy and matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The following structure of the linear hexasaccharide repeating unit of A. salmonicida O-antigen has been established: →3)-α-L-Rhap-(1→3)-α-D-ManpNAc-(1→2)-β-D-Glcp-(1→3)-α-L-Rhap2OAc4OAc-(1→3)-β-D-ManpNAc-(1→3)-α-D-Glcp-(1→. This new A. salmonicida O-polysaccharide was detected among two isolates collected from trout and turbot fish in 2010 and 2011, respectively. Further investigations should be conducted to evaluate the distribution of this new O-polysaccharide among a larger collection of isolates, depending on their geographic origin, the species of fish, and the health status of the fish.

Hsp40–Hsp70 typically function in concert as molecular chaperones, and their roles in post-infection immune responses are increasingly recognized. However, in the economically important fish species Scophthalmus maximus (turbot), there is still a lack in the systematic identification, interaction models, and binding site analysis of these proteins. Herein, 62 Hsp40 genes and 16 Hsp70 genes were identified in the turbot at a genome-wide level and were unevenly distributed on 22 chromosomes through chromosomal distribution analysis. Phylogenetic and syntenic analysis provided strong evidence in supporting the orthologies and paralogies of these HSPs. Protein–protein interaction and expression analysis was conducted to predict the expression profile after challenging with Aeromonas salmonicida. dnajb1b and hspa1a were found to have a co-expression trend under infection stresses. Molecular docking was performed using Auto-Dock Tool and PyMOL for this pair of chaperone proteins. It was discovered that in addition to the interaction sites in the J domain, the carboxyl-terminal domain of Hsp40 also plays a crucial role in its interaction with Hsp70. This is important for the mechanistic understanding of the Hsp40–Hsp70 chaperone system, providing a theoretical basis for turbot disease resistance breeding, and effective value for the prevention of certain diseases in turbot.

Aeromonas salmonicida is an ancient fish pathogen. Lysozymes are important molecules in the innate immune system that fight bacterial infections. The expression characteristics of C-type lysozyme in crucian carp infected with A. salmonicida and its antibacterial effect against A. salmonicida had not been investigated. Thus, we used bioinformatics to analyze the gene and protein characteristics of C-type lysozymes in crucian carp. Changes in C-type lysozyme expression before and after crucian carp infection with A. salmonicida were detected, and the in vitro antibacterial effect of recombinant carp C-type lysozyme on A. salmonicida was validated. The results showed that the coding DNA sequence region of the lysozyme gene sequence was 438 bp long, encoding 145 amino acids and containing two conserved catalytic sites: Glu53 and Asp69. Phylogenetic analysis revealed that crucian carp C-type lysozymes clustered with Cyprinus carpio lysozyme C. After crucian carp were infected with A. salmonicida, the gene and protein expression of C-type lysozymes in the liver, spleen, kidney, and hindgut were significantly upregulated, with the liver showing the highest upregulation that was 15 times higher than that in the uninfected group. In addition, recombinant C-type lysozyme exhibited significant antibacterial activity against A. salmonicida, with an average inhibition zone radius of 0.92 cm when using 40 μg recombinant lysozyme. In conclusion, this study reveals the important role of C-type lysozymes in the innate immune response of crucian carp and provides a theoretical basis for preventing crucian carp infection with A. salmonicida.

The plasmid known as pAsa5 is present in Aeromonas salmonicida subsp. salmonicida, a fish pathogen. The pAsa5 plasmid carries genes that are essential for the bacterium's virulence. Recombination events are known to occur in pAsa5, resulting in the loss of certain segments or the acquisition of additional genetic elements. For example, the transposon carried by the large pAsa8 plasmid was found to be inserted into the pAsa5 plasmid in the SHY16-3432 strain, enabling the addition of antibiotic resistance genes to this plasmid, which does not normally possess any. In this study, we present the isolation of additional strains carrying pAsa8. Further analyses of these strains revealed that a fusion between pAsa5 and the complete version of pAsa8 is possible. The pAsa8 transposon insertion in pAsa5 seen in the SHY16-3432 strain appears to be an aberrant event compared to the fusion of the two full-length plasmids. A 22-nucleotide sequence, present in both plasmids, serves as the site for the fusion of the two plasmids. Moreover, it is possible to introduce pAsa8 through conjugation into naive strains of A. salmonicida subsp. salmonicida and once the plasmid is within a new strain, the fusion with pAsa5 is detectable. This study reveals a previously unexplored aspect of pAsa5 plasmid biology, highlighting an additional risk for the spread of antibiotic resistance genes in A. salmonicida subsp. salmonicida. [ABSTRACT FROM AUTHOR]

Aeromonas salmonicida is the causative agent of septicemia in fish, and it is associated with significant economic losses in the aquaculture industry. While piscine Aeromonas infections are mainly treated with antibiotics, the emergence of resistance in bacterial populations requires the development of alternative methods of treatment. The use of phages can be one of them. A novel A. salmonicida jumbo phage, AerS_266, was isolated and characterized. This phage infects only mesophilic A. salmonicida strains and demonstrates a slow lytic life cycle. Its genome contains 243,674 bp and 253 putative genes: 84 encode proteins with predicted functions, and 3 correspond to tRNAs. Genes encoding two multisubunit RNA polymerases, chimallin and PhuZ, were identified, and AerS_266 was thus defined as a phiKZ-like phage. While similar phages with genomes >200 kb specific to Aeromonas hydrophila and Aeromonas veronii have been previously described, AerS_266 is the first phiKZ-like phage found to infect A. salmonicida. [ABSTRACT FROM AUTHOR]

We studied the phenotypic and genomic characteristics related to the virulence and antibiotic resistance of two Aeromonas strains, which were co-isolated before an outbreak of Aeromonas veronii among diseased seabass on Agathonisi Island, Greece, in April 2015. The first strain, AG2.13.2, is a potentially pathogenic mesophilic variant of Aeromonas salmonicida, and the second, AG2.13.5, corresponds to an Aeromonas rivipollensis related to A. rivipollensis KN-Mc-11N1 with an ANI value of 97.32%. AG2.13.2 lacks the type III secretion system just like other mesophilic strains of A. salmonicida. This characteristic has been associated with lower virulence. However, the genome of AG2.13.2 contains other important virulence factors such as type II and type VI secretion systems, and toxins such as rtxA, aerolysin aer/act, and different types of hemolysins. The strain also carries several genes associated with antibiotic resistance such as the tetE efflux pump, and exhibits resistance to tetracycline, ampicillin, and oxolinic acid. In an in vivo challenge test with gilthead seabream larvae, the A. veronii bv sobria strain AG5.28.6 exhibited the highest virulence among all tested strains. Conversely, both A. salmonicida and A. rivipollensis showed minimal virulence when administered alone. Interestingly, when A. veronii bv sobria AG5.28.6 was co-administered with A. rivipollensis, the larvae survival probability increased compared to those exposed to A. veronii bv sobria AG5.28.6 alone. This finding indicates an antagonistic interaction between A. veronii bv sobria AG5.28.6 and A. rivipollensis AG2.13.5. The co-administration of A. veronii bv sobria AG5.28.6 with Aeromonas salmonicida did not yield distinct survival probabilities. Our results validate that the primary pathogen responsible for European seabass aeromoniasis is Aeromonas veronii bv sobria. [ABSTRACT FROM AUTHOR]

Simple Summary: Aeromonas salmonicida represents an important cause of mortality in a broad diversity of fish species, in which it has been extensively reported. Nevertheless, aeromoniasis in sturgeons is far from being fully understood. Four juvenile sturgeons from a mortality outbreak in a farm located in the north of Spain were submitted to the VISAVET Health Surveillance Centre. Aeromonas salmonicida was isolated from the bacteriological culture. They were found to suffer from a hyperacute form of the disease (almost no clinical signs and non-specific gross or histological lesions), thus confirming the susceptibility of sturgeons to this pathogen. The findings described here hope to provide a better understanding of this potentially fatal disease in the industries of aquaculture and caviar. Aeromonas salmonicida is one of the major threats to world aquaculture, causing fish furunculosis and high mortality rates in cultured fish, particularly salmonids. Although Aeromonas spp. is a thoroughly studied pathogen, little is known regarding aeromoniasis in sturgeons. After a mortality outbreak, four juvenile sturgeons (Acipenser baerii) were submitted for autopsy and tissue samples were collected for histopathological and microbiological studies. The external examination revealed size heterogenicity, skin hyperpigmentation and reduced body condition of sturgeons. Within the abdominal cavity, mild hepatomegaly and splenomegaly were observed, as well as generalized organic congestion. Histology revealed severe multifocal haemorrhagic and ulcerative dermatitis, mainly localized in the dorsal and latero-ventral areas of fish. The histological study also showed moderate to severe inflammation of gills and organic lesions compatible with septicaemia. Bacterial isolates were identified as Aeromonas salmonicida subsp. salmonicida using MALDI-TOF MS and PCR. Overall, the lesions first described here are consistent with those previously reported in other cultured fish species and contribute to a better understanding of the pathogenesis of Aeromonas salmonicida subsp. salmonicida in the Siberian sturgeon, aside from providing new diagnostic tools for bacterial diseases impacting the fast-growing industry of caviar. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the compliance with Good Aquaculture Practices (GAPs) in the Health Compliance Units (UPAs) during the incubation of rainbow trout eggs and to identify the presence of Aeromonas salmonicida and Aeromonas hydrophila using the Polymerase Chain Reaction (PCR) technique. Design/Methodology/Approach: Seven and three egg incubation UPAs were identified in Puebla and Veracruz, respectively. An structured questionnaire was applied to evaluate the compliance with the GAPs. Samples were collected from the trout avelin batches and were analyzed in search of A. salmonicida and A. hydrophila using the PCR technique. The results were compared with the health characteristics of the fish of each batch. Results: The GAP-certified UPAs comply with the recommendations made by the authorities and do not show morbidity. UPAs that comply with less GAP points have health issues. A. hydrophila was detected in batches with a lower compliance with the GAPs. The presence of A. salmonicida was not identified. Study Limitations/Implications: The lack of compliance with the GAPs can lead to infection by other trout pathogens (not taken into consideration in this study). Findings/Conclusions: Compliance with the GAPs reduces the health risk at the rainbow trout egg incubation UPAs. [ABSTRACT FROM AUTHOR]

Lactic acid bacteria Ligilactobacillus salivarius has been shown to be a substitute for antibiotics in the treatment of bacterial disease in high animals. However, its beneficial mechanism in fish farming is still unclear. This study evaluated the antagonistic effects of the Ligilactobacillus salivarius GX118 strain on Aeromonas salmonicida and its regulation of rainbow trout growth in vivo and in vitro. The results found that GX118 produces an antibacterial substance that can directly destroy the cell wall of A. salmonicida. Whole-genome sequencing of GX118 revealed that Enterolysin A is a type III bacteriocin with antibacterial properties. An in vivo experiment showed that the supplementation of GX118 in diet competitively inhibited the colonization of A. salmonicida in the intestine. In addition, it was able to improve the growth performance of rainbow trout within a 21-day feeding experiment. The supplementation of GX118 increased the diversity of gut microbiota, in which the abundance of Bacteroidota, Blautia, and Rhodobacteraceae increased. In addition, the use of GX118 activated the expression of IFN-γ and NF-κB genes and reduced the expression level of IL-6 and IL-8, thus exhibiting a certain effect on activating the immunity of rainbow trout. This study provides a scientific basis for the development of antibacterial probiotics in the healthy farming of rainbow trout.

The Aeromonas salmonicida is responsible for causing furunculosis in various fish species. Furunculosis is a ubiquitous disease that affects the aquaculture industry and causes the mass mortality of turbot. Vibrio vulnificus is a pathogen that causes skin ulcers and hemorrhagic septicemia in fish, resulting in significant mortality in aquaculture. In this study, we have established a bivalent inactivated vaccine against A. salmonicida and V. vulnificus with Montanide™ ISA 763 AVG as an adjuvant. This bivalent inactivated vaccine was used to immunize turbot by intraperitoneal injection, and the relevant immune indexes were detected. The results demonstrate that the bivalent inactivated vaccine exhibited a relative percent survival (RPS) of 77% following A. salmonicida and V. vulnificus intraperitoneal challenge. The vaccinated group exhibited higher levels of acid phosphatase activity and lysozyme activity compared to the control group. ELISA results showed a significant increase in serum antibody levels in immunized turbot, which was positively correlated with immunity. In the kidney tissue, related immune genes (TLR5, CD4, MHCI and MHCII) were up-regulated significantly, showing that the vaccine can induce cellular and humoral immune responses in turbot. In conclusion, the bivalent inactivated vaccine against A. salmonicida and V. vulnificus was immunogenic, efficiently preventing turbot from infection, which has the potential to be applied in aquaculture.

Representatives of the bacterial genus Aeromonas are some of the most notorious aquaculture pathogens associated with a range of diseases in different fish species. As the world forges toward the post-antibiotic era, alternative options for combating bacterial pathogens are needed. One such alternative option is phage biocontrol. In this study, a novel podophage—JELG-KS1—infecting Aeromonas salmonicida was retrieved from wastewater along with its host strain. The genome of the JELG-KS1 phage is a 40,505 bp dsDNA molecule with a GC% of 53.42% and 185 bp direct terminal repeats and encodes 53 predicted proteins. Genomic analysis indicates that JELG-KS1 might represent a novel genus within the subfamily Studiervirinae. Podophage JELG-KS1 is a strictly lytic phage without any identifiable virulence or AMR genes that quickly adsorbs onto the surface of host cells to initiate a 48 min long infectious cycle, resulting in the release of 71 ± 12 JELG-KS1 progeny virions per infected cell. JELG-KS1 effectively lyses its host population in vitro, even at very low multiplicities of infection. However, when challenged against a panel of Aeromonas spp. strains associated with diseases in aquaculture, JELG-KS1 shows host-specificity that is confined only to its isolation strain, immediately compromising its potential for Aeromonas spp. biocontrol in aquaculture.

Turbot aquaculture production is an important economic activity in several countries around the world; nonetheless, the incidence of diseases, such furunculosis, caused by the etiological agent A. salmonicida subsp. salmonicida, is responsible for important losses to this industry worldwide. Given this perspective, this study aimed to evaluate early immune responses in turbot (S. maximus L.) following infection with A. salmonicida subsp. salmonicida. For this, 72 fish were individually weighed and randomly distributed into 6 tanks in a circulating seawater system. For the bacterial challenge, half of the individuals (3 tanks with 36 individuals) were infected using a peritoneal injection with the bacterial suspension, while the other half of individuals were injected with PBS and kept as a control group. Several factors linked to the innate immune response were studied, including not only haematological (white blood cells, red blood cells, haematocrit, haemoglobin, mean corpuscular volume, mean cell haemoglobin, mean corpuscular haemoglobin concentration, neutrophils, monocytes, lymphocytes, thrombocytes) and oxidative stress parameters, but also the analyses of the expression of 13 key immune-related genes (tnf-α, il-1β, il-8, pparα-1, acox1, tgf-β1, nf-kB p65, srebp-1, il-10, c3, cpt1a, pcna, il-22). No significant differences were recorded in blood or innate humoral parameters (lysozyme, anti-protease, peroxidase) at the selected sampling points. There was neither any evidence of significant changes in the activity levels of the oxidative stress indicators (catalase, glutathione S-transferase, lipid peroxidation, superoxide dismutase). In contrast, pro-inflammatory (tnf-α, il-1β), anti-inflammatory (il-10), and innate immune-related genes (c3) were up-regulated, while another gene linked with the lipid metabolism (acox1) was down-regulated. The results showed new insights about early responses of turbot following infection with A. salmonicida subsp. salmonicida. [ABSTRACT FROM AUTHOR]

Vibrio scophthalmi and Aeromonas salmonicida can cause high turbot mortality and huge economic losses. Presently, vaccination is the most promising method for preventing communicable diseases. In this study, we used formalin to kill V. scophthalmi and A. salmonicida cells, and mixed with the mineralized oil adjuvant (Montanide™ ISA 763 AVG) to prepare the bivalent inactivated vaccine. The results showed that turbot inoculated with the bivalent inactivated vaccine exhibited strong tolerance to the infection of V. scophthalmi and A. salmonicida, and no obvious clinical symptoms and pathological changes were observed. The activities of enzymes lysozyme, acid phosphatase and complement C3 had significantly increased after the vaccination. The antibody titer response of vaccinated turbot was greatly boosted, which was positively connected with the immunological impact according to ELISA results. Simultaneously, the expression levels of immune-related genes such as MHC-IIα, MHC-IIβ, CD4, CD8, TNF-α and IL-1β were up-regulated, demonstrating that it might stimulate humoral and cellular immunological response in turbot. These findings highlight the potential of the bivalent inactivated vaccine for controlling V. scophthalmi and A. salmonicida infections in turbot. [ABSTRACT FROM AUTHOR]

The Gram-negative bacterium A. salmonicida is the causal agent of furunculosis and used to be one of the most loss-causing bacterial infections in the salmonid aquaculture industry with a mortality rate of about 90% until the 1990s, when an inactivated vaccine with mineral oil as adjuvant was successfully implemented to control the disease. However, the use of this vaccine is associated with inflammatory side effects in the peritoneal cavity as well as autoimmune reactions in Atlantic salmon, and incomplete protection has been reported in rainbow trout. We here aimed at developing and testing a recombinant alternative vaccine based on virus-like particles (VLPs) decorated with VapA, the key structural surface protein in the outer A-layer of A. salmonicida. The VLP carrier was based on either the capsid protein of a fish nodavirus, namely red grouper nervous necrotic virus (RGNNV) or the capsid protein of Acinetobacter phage AP205. The VapA and capsid proteins were expressed individually in E. coli and VapA was fused to auto-assembled VLPs using the SpyTag/SpyCatcher technology. Rainbow trout were vaccinated/immunized with the VapA-VLP vaccines by intraperitoneal injection and were challenged with A. salmonicida 7 weeks later. The VLP vaccines provided protection comparable to that of a bacterin-based vaccine and antibody response analysis demonstrated that vaccinated fish mounted a strong VapA-specific antibody response. To our knowledge, this is the first demonstration of the potential use of antigendecorated VLPs for vaccination against a bacterial disease in salmonids. [ABSTRACT FROM AUTHOR]

Aeromonas salmonicida subsp. salmonicida (A. salmonicida), a Gram-negative bacterium causing furunculosis in fish, produces the siderophores acinetobactin and amonabactins in order to extract iron from its hosts. While the synthesis and transport of both systems is well understood, the regulation pathways and conditions necessary for the production of each one of these siderophores are not clear. The acinetobactin gene cluster carries a gene (asbI) encoding a putative sigma factor belonging to group 4 σ factors, or, the ExtraCytoplasmic Function (ECF) group. By generating a null asbI mutant, we demonstrate that AsbI is a key regulator that controls acinetobactin acquisition in A. salmonicida, since it directly regulates the expression of the outer membrane transporter gene and other genes necessary for Fe-acinetobactin transport. Furthermore, AsbI regulatory functions are interconnected with other iron-dependent regulators, such as the Fur protein, as well as with other sigma factors in a complex regulatory network. [ABSTRACT FROM AUTHOR]

This study aimed to identify histamine-forming bacteria (HFB) and the sources of introduction of such bacteria to recommend control measures to mitigate histamine formation in yellowfin tuna (YFT). Field samples were collected from multi-day boats that landed at Dikkowita, Negombo, Trincomalee and Dondra fishery harbours. Ice from the fish holds (n=63) and chill transport vehicles (n=63), and swabs from the fish holds (n=63), the boat decks (n=63) and the skin of YFT (n=63) were collected. Fish loin samples (n=15), ice samples (n=36) and swabs from the skin of YFT (n=18), floor (n=18) and chill transport vehicles (n=18) were collected from fish processing plants. Presumptive HFB isolated from Nivens medium and Violet Red Bile Glucose (VRBG) agar were screened for histamine forming ability in Tripticase soy broth (TSB) supplemented with 1.0% L-histidine. HFB isolates were characterized by sequencing approximately 1400 bp of the 16S rDNA. Seven isolates that produced histamine in the range of 3000--4000 ppm in TSB isolated from ice samples, and a swab sample collected from the boat deck, were confirmed as Klebsiella aerogens (n=6) and Morganella morganii (n=1) respectively. Hafnia alvei (n=1), Serratia sp. (n=2), Citrobacter freundii (n=1), Rahnella sp. (n=1) and Aeromonas salmonicida (n=8) were also among the isolated histamine forming bacteria. Pseudomonas sp. (n=24) and Shewanella baltica (n=7), which are known as spoilage bacteria were also isolated and showed weak histamine formation. Hence, it is evident that histamineforming bacteria could be introduced into the fish from ice and contacting surfaces. This necessitates the practice of rigorous cleaning procedures and adaptation of proper postharvest handling procedures to minimize contamination of the fish. [ABSTRACT FROM AUTHOR]

ABSTRACTCarbohydrates can both protect against infection and act as targets promoting infection. Mucins are major components of the slimy mucus layer covering the fish epithelia. Mucins can act as decoys for intimate pathogen interaction with the host afforded by binding to glycosphingolipids in the host cell membrane. We isolated and characterized glycosphingolipids from Atlantic salmon skin, gill, stomach, pyloric caeca, and intestine. We characterized the glycosphingolipids using liquid chromatography – mass spectrometry and tandem mass spectrometry and the glycan repertoire was compared with the glycan repertoire of mucins from the same epithelia. We also investigated Aeromonas salmonicida binding using chromatogram and microtiter well based binding assays. We identified 29 glycosphingolipids. All detected acid glycans were of the ganglio-series (unless shorter) and showed a high degree of polysialylation. The non-acid glycans were mostly composed of the neolacto, globo, and ganglio core structures. The glycosphingolipid repertoire differed between epithelia and the proportion of the terminal moieties of the glycosphingolipids did not reflect the terminal moieties on the mucins from the same epithelia. A. salmonicida did not bind the Atlantic salmon glycosphingolipids. Instead, we identified that A. salmonicida binding to sialic acid occurred to α2–6 Neu5Ac but not to α2–3 Neu5Ac. α2–6 Neu5Ac was present on mucins whereas mainly α2–3 Neu5Ac was found on the glycosphingolipids, explaining the difference in A. salmonicida binding ability between these host glycoconjugates. A. salmonicida´s ability to bind to Atlantic salmon mucins, but not the glycosphingolipids, is likely part of the host defence against this pathogen.