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ABSTRACT Edwardsiella ictaluri and Flavobacterium covae are two bacteria species that cause diseases in farm‐raised channel catfish (Ictalurus punctatus) that cause heavy economic damage to the aquaculture industry, particularly to the channel catfish farming. In search for environmentally benign antibacterial compounds active against E. ictaluri and F. covae, we investigated the constituents isolated from Brazilian red, brown and green propolis. We have also synthetically modified active constituents to see if lipophilicity plays a role in enhancing antibacterial activities. Vestitol, neovestitol and methylvestitol were found to be the active constituents with minimum inhibitory concentration (MIC) relative to drug control florfenicol (RDCF) values (MIC−RDCF) of 7.6, 7.6 and 7.9 mg/L, respectively, against F. covae. The activity against E. ictaluri was not significant. [ABSTRACT FROM AUTHOR]

ABSTRACT Flavobacterium psychrophilum, the causative agent of bacterial cold water disease (BCWD), is one of the leading pathogens in rainbow trout (Oncorhynchus mykiss) aquaculture. To date, there is little knowledge of the transmission kinetics of F. psychrophilum over the course of infection. In particular, how transmission is affected by host genotype and pathogen exposure dosage are not well studied. In order to fill in these knowledge gaps, we exposed two divergently selected lines of rainbow trout (ARS‐Fp‐R and ARS‐Fp‐S) to a range of dosages of F. psychrophilum (strain CSF117‐10). We then measured mortality and bacterial shedding to estimate transmission risk at multiple time points since initial infection. As dosage increased, the number of fish shedding and the amount of bacteria shed increased ranging from 0% to 100% and 103 to 108 cells fish−1 h−1, respectively. In addition, we found that disease resistance (survival) was not correlated with transmission risk blocking, in that 67% of fish which shed bacteria experienced no clinical disease. In general, fish mortality began on Day 3, peaked between Days 5–7 and was higher in the ARS‐Fp‐R line. Results from this study could be used to develop epidemiological models and improve disease management, particularly in the context of aquaculture and selective breeding. [ABSTRACT FROM AUTHOR]

The intricate relationship between regeneration and microbiota has recently gained attention, spanning diverse model organisms. Axolotl (Ambystoma mexicanum) is a critically endangered salamander species and a model organism for regenerative and developmental biology. Despite its significance, a noticeable gap exists in understanding the interplay between axolotl regeneration and its microbiome. Here, we analyse in depth bacterial 16S rRNA amplicon dataset that we reported before as data resource and profile fungal community by sequencing ITS amplicons at the critical stages of limb regeneration (0–1–4–7–30–60 days post amputation, ‘dpa’). Results reveal a decline in richness and evenness in the course of limb regeneration, with bacterial community richness recovering beyond 30 dpa unlike fungi community. Beta diversity analysis reveals precise restructuring of the bacterial community along the three phases of limb regeneration, contrasting with less congruent changes in the fungal community. Temporal dynamics of the bacterial community highlight prevalent anaerobic bacteria in initiation phase and Flavobacterium bloom in the early phase correlating with limb blastema proliferation. Predicted functional analysis mirrors these shifts, emphasising a transition from amino acid metabolism to lipid metabolism control. Fungal communities shift from Blastomycota to Ascomycota dominance in the late regeneration stage. Our findings provide ecologically relevant insights into stage specific role of microbiome contributions to axolotl limb regeneration. [ABSTRACT FROM AUTHOR]

Petroleum hydrocarbons (PHCs) can be biodegraded into CO2, and PHC‐contaminated aquifers are always deemed as carbon sources. Fortunately, some carbon fixation microorganisms have been found in PHC‐contaminated sites. However, most of the studies are related to volatile short‐chain PHC, and few studies focus on long‐chain PHC‐contaminated sites. To reveal the carbon fixation microorganisms in these sites, in the study, a long‐chain PHC polluted site in North China was selected. Through hydrochemical and metagenomics analysis, the structure and capacity of carbon fixing microorganisms in the site were revealed. Results showed that there were many kinds of carbon fixed microorganisms that were identified such as Flavobacterium, Pseudomonas. HP/4HB, rTCA, and DC/4HB cycles were dominated carbon fixation pathways. The long‐chain PHC were weakly correlated with carbon fixation microorganisms, but it may stimulate the growth of some carbon fixation microorganisms, such as microorganisms involved in rTCA cycle. Practitioner Points: The microorganisms with carbon fixation gene exist in the aquifer contaminated by long‐chain petroleum hydrocarbon.Microorganisms that have the ability to degrade petroleum also have the ability to carbon fixation.Long‐chain petroleum hydrocarbon may promote the growth of carbon fixation microorganisms. [ABSTRACT FROM AUTHOR]

Glycoside hydrolase family 97 (GH97) comprises enzymes like anomer‐inverting α‐glucoside hydrolases (i.e., glucoamylase) and anomer‐retaining α‐galactosidases. In a soil bacterium, Flavobacterium johnsoniae, we previously identified a GH97 enzyme (FjGH97A) within the branched dextran utilization locus. It functions as an α‐glucoside hydrolase, targeting α‐(1→6)‐glucosidic linkages in dextran and isomaltooligosaccharides (i.e., glucodextranase). FjGH97A exhibits a preference for α‐(1→6)‐glucoside linkages over α‐(1→4)‐linkages, while Bacteroides thetaiotaomicron glucoamylase SusB (with 69% sequence identity), which is involved in the starch utilization system, exhibits the highest specificity for α‐(1→4)‐glucosidic linkages. Here, we examined the crystal structures of FjGH97A in complexes with glucose, panose, or isomaltotriose, and analyzed the substrate preferences of its mutants to identify the amino acid residues that determine the substrate specificity for α‐(1→4)‐ and α‐(1→6)‐glucosidic linkages. The overall structure of FjGH97A resembles other GH97 enzymes, with conserved catalytic residues similar to anomer‐inverting GH97 enzymes. A comparison of active sites between FjGH97A and SusB revealed differences in amino acid residues at subsites +1 and +2 (specifically Ala195 and Ile378 in FjGH97A). Among the three mutants (A195S, I378F, and A195S‐I378F), A195S and A195S‐I378F exhibited increased activity toward α‐(1→4)‐glucoside bonds compared to α‐(1→6)‐glucoside bonds. This suggests that Ala195, located on the Gly184‐Thr203 loop (named loop‐N) conserved within the GH97 subgroup, including FjGH97A and SusB, holds significance in determining linkage specificity. The conservation of alanine in the active site of the GH97 enzymes, within the same gene cluster as the putative dextranase, indicates its crucial role in determining the specificity for α‐(1→6)‐glucoside linkage. [ABSTRACT FROM AUTHOR]

Both above‐ and below‐ground parts of plants are constantly challenged with microbes and interact closely with them. Many plant‐growth‐promoting rhizobacteria, mostly interacting with the plant's root system, enhance the immunity of plants in a process described as induced systemic resistance (ISR). Here, we characterized local induced resistance (IR) triggered by the model PGPR Pseudomonas simiae WCS417r (WCS417) in Arabidopsis thaliana. Hydroponic application of WCS417 to Arabidopsis roots resulted in propagation of WCS417 in/on leaves and the establishment of local IR. WCS417‐triggered local IR was dependent on salicylic acid (SA) biosynthesis and signalling and on functional biosynthesis of pipecolic acid and monoterpenes, which are classically associated with systemic acquired resistance (SAR). WCS417‐triggered local IR was further associated with a priming of gene expression changes related to SA signalling and SAR. A metabarcoding approach applied to the leaf microbiome revealed a significant local IR‐associated enrichment of Flavobacterium sp.. Co‐inoculation experiments using WCS417 and At‐LSPHERE Flavobacterium sp. Leaf82 suggest that the proliferation of these bacteria is influenced by both microbial and immunity‐related, plant‐derived factors. Furthermore, application of Flavobacterium Leaf82 to Arabidopsis leaves induced SAR in an NPR1‐dependent manner, suggesting that recruitment of this bacterium to the phyllosphere resulted in propagation of IR. Together, the data highlight the importance of plant‐microbe‐microbe interactions in the phyllosphere and reveal Flavobacterium sp. Leaf82 as a new beneficial promoter of plant health. [ABSTRACT FROM AUTHOR]

Flavobacterium psychrophilum (Fp) causes Bacterial Cold Water Disease in salmonids. During host‐pathogen interactions, gram‐negative bacteria, such as Fp, release external membrane vesicles (OMVs) harbouring cargos, such as DNA, RNA and virulence factors. This study aimed to characterise the potential role of the OMVs' small RNAs (sRNAs) in the Fp‐rainbow trout host‐pathogen interactions. sRNAs carried within OMVs were isolated from Fp. RNA‐Seq datasets from whole‐cell Fp and their isolated OMVs indicated substantial enrichment of specific sRNAs in the OMVs compared to the parent cell. Many of the OMV‐packaged sRNAs were located in the pathogenicity islands of Fp. Conservation of sRNAs in 65 strains with variable degrees of virulence was reported. Dual RNA‐Seq of host and pathogen transcriptomes on day 5 post‐infection of Fp ‐resistant and ‐susceptible rainbow trout genetic lines revealed correlated expression of OMV‐packaged sRNAs and their predicted host's immune gene targets. In vitro, treatment of the rainbow trout epithelial cell line RTgill‐W1 with OMVs showed signs of cytotoxicity accompanied by dynamic changes in the expression of host genes when profiled 24 h following treatment. The OMV‐treated cells, similar to the Fp ‐resistant fish, showed downregulated expression of the suppressor of cytokine signalling 1 (SOCS1) gene, suggesting induction of phagosomal maturation. Other signs of modulating the host gene expression following OMV‐treatment include favouring elements from the phagocytic, endocytic and antigen presentation pathways in addition to HSP70, HSP90 and cochaperone proteins, which provide evidence for a potential role of OMVs in boosting the host immune response. In conclusion, the study identified novel microbial targets and inherent characteristics of OMVs that could open up new avenues of treatment and prevention of fish infections. [ABSTRACT FROM AUTHOR]

Nodular gill disease (NGD) is an emerging condition associated with amoeba trophozoites in freshwater salmonid farms. However, unambiguous identification of the pathogens still must be achieved. This study aimed to identify the amoeba species involved in periodic NGD outbreaks in two rainbow trout (Oncorhynchus mykiss) farms in Northeastern Italy. During four episodes (February–April 2023), 88 fish were euthanized, and their gills were evaluated by macroscopic, microscopic and histopathological examination. The macroscopic and microscopic severity of the lesions and the degree of amoebae infestation were scored and statistically evaluated. One gill arch from each animal was put on non‐nutrient agar (NNA) Petri dishes for amoeba isolation, cultivation and subsequent identification with SSU rDNA sequencing. Histopathology confirmed moderate to severe lesions consistent with NGD and mild to moderate amoeba infestation. The presence of amoebae was significantly correlated with lesion severity. Light microscopy of cultured amoebae strains and SSU rDNA analysis revealed the presence of a previously characterized amoeba Naegleria sp. strain GERK and several new strains: two strains from Hartmannelidae, three vannelid amoebae from the genus Ripella and cercozoan amoeba Rosculus. Despite the uncertainty in NGD etiopathogenesis and amoebae pathogenic role, identifying known and new amoebae leans towards a possible multi‐aetiological origin. [ABSTRACT FROM AUTHOR]

Objective: Doxycycline (DO) has been used in fish for a long time, but there are some factors that have not yet been clarified regarding its pharmacokinetic (PK) and pharmacodynamic (PD) properties. Therefore, the aim of this study was to investigate the PK and PK/PD targets of DO after 20 mg/kg intravascular (IV), intramuscular (IM) and oral (OR) gavage administration in rainbow trout (Oncorhynchus mykiss). Methods: Plasma samples were collected at specific time points and subsequently analysed by HPLC‐ultraviolet. The PK/PD indices were calculated based on the MIC90 (Aeromonas hydrophila and Aeromonas sobria) values obtained for the respective bacteria and the PK parameters obtained for DO following both IM and OR administration. Results: After IV administration, the elimination half‐life (t1/2ʎz), area under the concentration vs. time curve (AUC), apparent volume of distribution at steady‐state and total body clearance of DO were 34.81 h, 723.82 h µg/mL, 1.24 L/kg and 0.03 L/kg/h, respectively. The t1/2λz of the DO was found to be 37.39 and 39.78 h after IM, and OR administration, respectively. The bioavailability was calculated 57.02% and 32.29%, respectively, after IM and OR administration. The MIC90 of DO against A. hydrophila and A. sobria was 4 µg/mL. The PK/PD integration showed that DO (20 mg/kg dose) for A. hydrophila and A. sobria with MIC90 ≤4 µg/mL achieved target AUC/MIC value after IM administration. Conclusions: These results suggest that when rainbow trout was treated with 20 mg/kg IV and IM administered DO, therapeutically effective concentrations were reached in the control of infections caused by A. hydrophila and A. sobria. [ABSTRACT FROM AUTHOR]

Background: Annual or seasonal killifishes (Cyprinodontiformes: Nothobranchiidae) are unique among fish in their ability to enter into developmental arrests (diapauses: DI, DII, and DIII). They have a short lifespan and their embryos are exceptionally tolerant to a variety of environmental stresses. These traits make them a popular model for studying vertebrate diapause, aging, stress tolerance, genome adaptation, and evolution. In such issues, in a comparative evolutionary framework, Fundulopanchax gardneri, a popular aquarium fish from Africa, is commonly used as a representative non‐annual model though its development is not studied in detail and whether it includes diapauses remains uncertain. Results: We described in detail for the first time embryonic development of F. gardneri and revealed it to resemble that in the undoubtedly annual Austrofundulus limnaeus killifish in displaying two developmental depressions. However, if compared with A. limnaeus, these developmental states look like "less intense" versions of DII and DIII rather than true diapauses. Conclusions: To determine whether developmental depressions in F. gardneri represent "true" diapauses or only their functional equivalents, detailed studies of embryonic development of different killifish both annual and non‐annual are needed. Before that, acceptance of F. gardneri as a representative non‐annual fish seems premature. Key Findings: Aplocheloid annual killifish (Cyprinodontiformes) are unique among fish in the ability to arrest development at different embryonic stages and prolong it for months and years. These and other traits make them a popular model in studying of aging, stress tolerance etc. In these studies, in a comparative framework, aplocheloid Fundulopanchax gardneri is usually used as a representative non‐annual fish though its developmental life‐history remains obscure.This study provides detailed description of F. gardneri development from early embryonic stages till sexual maturation with focus on developmental arrests, divergent developmental trajectories, duration of embryonic period, etc.The data provided in this study indicate F. gardneri to display developmental traits characteristic for annual killifish but in the "less expressed" form. Acceptance of F. gardneri as annual vs non‐annual is premature until the detailed study of embryonic development in other killifish both annual and non‐annual. [ABSTRACT FROM AUTHOR]

The seed‐endophytic bacterial community is a potentially beneficial and heritable fraction of the plant microbiome. Its utilization as a sustainable crop improvement strategy could be especially valuable for species such as hemp, where production is being scaled up and new challenges will be faced in managing crop productivity and health. However, little is known about the makeup and variation of the hemp seed microbiome. This study profiled the endophytic bacterial communities harboured by 16 hemp cultivars sourced from commercial suppliers in Europe. A 16S rDNA amplicon sequencing approach identified 917 amplicon sequence variants across samples. Taxonomic classification of sequences revealed 4 phyla and 87 genera to be represented in the dataset. Several genera were widespread while some were specific to one or a few cultivars. Flavobacterium, Pseudomonas, and Pantoea were notable in their high overall abundance and prevalence, but community composition was variable and no one taxon was universally abundant, suggesting a high degree of flexibility in community assembly. Taxonomic composition and alpha diversity differed among cultivars, though further work is required to understand the relative influence of hemp genetic factors on community structure. The taxonomic profiles presented here can be used to inform further work investigating the functional characteristics and potential plant‐growth‐promoting traits of seed‐borne bacteria in hemp. [ABSTRACT FROM AUTHOR]

Objective: Columnaris disease is a leading cause of disease‐related losses in the catfish industry of the southeastern United States. The term "columnaris‐causing bacteria" (CCB) has been coined in reference to the four described species that cause columnaris disease: Flavobacterium columnare, F. covae, F. davisii, and F. oreochromis. Historically, F. columnare, F. covae, and F. davisii have been isolated from columnaris disease cases in the catfish industry; however, there is a lack of knowledge of which CCB species are most prevalent in farm‐raised catfish. The current research objectives were to (1) sample columnaris disease cases from the U.S. catfish industry and identify the species of CCB involved and (2) determine the virulence of the four CCB species in Channel Catfish Ictalurus punctatus in controlled laboratory challenges. Methods: Bacterial isolates or swabs of external lesions from catfish were collected from 259 columnaris disease cases in Mississippi and Alabama during 2015–2019. The DNA extracted from the samples was analyzed using a CCB‐specific multiplex polymerase chain reaction to identify the CCB present in each diagnostic case. Channel Catfish were challenged by immersion with isolates belonging to each CCB species to determine virulence at ~28°C and 20°C. Result: Flavobacterium covae was identified as the predominant CCB species impacting the U.S. catfish industry, as it was present in 94.2% (n = 244) of diagnostic case submissions. Challenge experiments demonstrated that F. covae and F. oreochromis were highly virulent to Channel Catfish, with most isolates resulting in near 100% mortality. In contrast, F. columnare and F. davisii were less virulent, with most isolates resulting in less than 40% mortality. Conclusion: Collectively, these results demonstrate that F. covae is the predominant CCB in the U.S. catfish industry, and research aimed at developing new control and prevention strategies should target this bacterial species. The methods described herein can be used to continue monitoring the prevalence of CCB in the catfish industry and can be easily applied to other industries to identify which Flavobacterium species have the greatest impact. Impact statementColumnaris disease is a leading cause of disease‐related losses in the southeastern U.S. catfish industry. Recently, four species of columnaris‐causing bacteria (CCB) have been described, but their prevalence in the industry was unknown. In this research, Flavobacterium covae was identified as the predominant species of CCB impacting the industry and was demonstrated to be highly virulent in Channel Catfish. Research aimed at developing new control and prevention strategies should target this bacterial species. [ABSTRACT FROM AUTHOR]

Flavobacterium psychrophilum, the causative agent of bacterial coldwater disease, causes substantial economic losses in salmonid farms and hatcheries. Some multilocus sequence types (ST) of F. psychrophilum are more likely to be associated with fish farms and hatcheries, but it is unclear if these patterns of association represent genetic lineages that are more adapted to aquaculture environments. Towards elucidating the disease ecology of F. psychrophilum, the culturability of 10 distinct F. psychrophilum STs was evaluated for 13 weeks in three microcosms including sterilized well water, sterilized well water with commercial trout feed, or sterilized well water with raceway detritus. All STs remained culturable in each of the microcosms for at least 8 weeks, with bacterial concentrations often highest in the presence of raceway detritus. In addition, most (e.g., 90%) STs remained culturable for at least 13‐weeks. Significant differences in log10 cfus were observed among STs, both within and between microcosms, suggesting potential variability in environmental persistence capacity among specific variants. Collectively, results highlight the ability of F. psychrophilum to not only persist for weeks under nutrient‐limited conditions but also thrive in the presence of organic substrates common in fish farms and hatchery‐rearing units. [ABSTRACT FROM AUTHOR]

Flavobacterium plurextorum is a potential fish pathogen of interest, previously isolated from diseased rainbow trout (Oncorhynchus mykiss) and oomycete‐infected chum salmon (Oncorhynchus keta) eggs. We report here the first complete genome sequence of F. plurextorum RSG‐18 isolated from the gut of Schlegel's black rockfish (Sebastes schlegelii). The genome of RSG‐18 consists of a circular chromosome of 5,610,911 bp with a 33.57% GC content, containing 4858 protein‐coding genes, 18 rRNAs, 63 tRNAs and 1 tmRNA. A comparative analysis was conducted on 11 Flavobacterium species previously reported as pathogens or isolated from diseased fish to confirm the potential pathogenicity of RSG‐18. In the SEED classification, RSG‐18 was found to have 36 genes categorized in 'Virulence, Disease and Defense'. Across all Flavobacterium species, a total of 16 antibiotic resistance genes and 61 putative virulence factors were identified. All species had at least one phage region and type I, III and IX secretion systems. In pan‐genomic analysis, core genes consist of genes linked to phages, integrases and matrix‐tolerated elements associated with pathology. The complete genome sequence of F. plurextorum RSG‐18 will serve as a foundation for future research, enhancing our understanding of Flavobacterium pathogenicity in fish and contributing to the development of effective prevention strategies. [ABSTRACT FROM AUTHOR]

In this study, 2 yeasts, Candida zeylanoides isolate Y12‐3 (MN073455.1) and Hyphopichia pseudoburtonii isolate Y12‐1 (MN073489.1), were isolated from rainbow trout hindguts. Their identity was confirmed by amplification of internal transcribed spacer (ITS) gene regions. Their in vitro probiotic properties showed that they were hydrophobic and did not inhibit bacterial growth. However, they displayed positive co‐aggregation with several bacterial pathogens. It was interesting that Y12‐1 and Y12‐3 isolates survived and grew well in bile salt concentrations ranging from 0.6% to 1.5% and at low pH conditions. After a 35‐day feeding trial, the results of dietary incorporation of the yeast isolates showed that serum myeloperoxidase activity was unchanged in the treated and control groups. Meanwhile, serum lysozyme activity in yeast groups was decreased significantly compared to the control. However, fish fed with Y12‐1 had higher total protein, albumin, and globulin than other groups. Total cholesterol levels were decreased significantly in yeast groups compared to the control. Interestingly, the Y12‐1 and Y12‐3 groups showed higher survival (69.05% and 80.95%) after the challenge with Lactococcus garvieae compared to the control (45.24%). In brief, the aforementioned findings showed that Y12‐1 and Y12‐3 strains could be potential probiotic candidates for use in rainbow trout farming. [ABSTRACT FROM AUTHOR]

BACKGROUND: Evolving climatic conditions impact the behavior of microorganisms. The lack of efficiency of beneficial microorganisms against pathogens can be due to these evolving abiotic factors more favorable to the development and adaptation of pathogens. It is therefore of great interest to understand their impact (especially temperature increase and relative humidity (RH) variation) on pathogenic and non‐pathogenic microorganisms. This work aimed to examine the possible effects of increasing temperature (20, 25, 30 and 33 °C) and RH (40%, 50%, 60% and 80%) on the growth and mycotoxin production (deoxynivalenol (DON) and zearalenone (ZEN)) of Fusarium graminearum, on the growth of three commercial biocontrol agents (BCAs; Mycostop®, Xedavir® and Polyversum®) and on the pathogen–BCA interaction. RESULTS: Results demonstrated that BCAs have contrasting impacts on the growth and mycotoxinogenesis of F. graminearum depending on abiotic factors. At 25 °C and regardless of RH, commercial BCAs limit DON production by F. graminearum, but at 30 °C and intermediate RH, Xedavir® is no longer effective. The ability of Xedavir® to control the production of ZEN production by F. graminearum is also affected by abiotic factors. However, increasing temperature has an opposite effect on its ability to control the accumulation of ZEN. Polyversum® oomycete is the BCA with the most resilient efficacy against F. graminearum toxinogenesis under the different abiotic factors. CONCLUSION: This work provides new knowledge of the effect of these abiotic parameters on the interaction between BCA and F. graminearum, especially on the production of mycotoxins. It paves the way for the development of efficient and resilient mycotoxin biocontrol strategies using beneficial microorganisms against F. graminearum, thus contributing to global food security. © 2023 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Maize grain production in the tropics is affected by Fusarium verticillioides, leading to significant losses in yield and quality. Fungicides are commonly used to control foliar diseases and increase grain yield, but have limited efficacy against F. verticillioides and may even stimulate the fungus to produce mycotoxins. Biocontrol can reduce F. verticillioides populations and fumonisin, but little is known about the resulting microbial interactions on the phylloplane and their relationship with F. verticillioides. This study aimed at evaluating the impact of different foliar treatments on the cultivable phylloplane community of fungi and bacteria and their potential in controlling F. verticillioides in maize. Results showed that two sprays of Bacillus subtilis BIOUFLA2 increased the percentage of native phylloplane antagonist bacteria and fungi against F. verticillioides by 25.0% and 27.3%, respectively, compared with the water control. However, two sprays with a triazole + strobilurin fungicide at different phenological stages in maize reduced such antagonistic communities. From field trials, 34 bacteria and 13 fungi from the phylloplane were selected for their inhibitory activity against F. verticillioides, with most coming from the exclusive treatment with BIOUFLA2. The study also revealed that the biocontrol agent‐recruited phylloplane indigenous lineages act through antibiosis, competition, and parasitism. Therefore, the adoption of biocontrol to limit F. verticillioides population build‐up not only has a direct activity on the pathogen but also recruits fungal and bacterial antagonists that may act additively or synergistically with the applied biocontrol agent. [ABSTRACT FROM AUTHOR]

Bacterial coldwater disease (BCWD), caused by Flavobacterium psychrophilum, results in significant losses among multiple salmonid (family Salmonidae) species. Molecular epidemiology and serotyping studies have suggested that some variants are host specific; however, these associations have not been evaluated by cross‐challenging fish species with putatively host‐associated F. psychrophilum isolates via more natural (i.e. immersion) exposure routes. To this end, F. psychrophilum isolates US19‐COS, US62‐ATS and US87‐RBT, each originally recovered from diseased coho salmon (Oncorhynchus kisutch), Atlantic salmon (Salmo salar) or rainbow trout (O. mykiss), and belonging to a host‐associated multilocus sequence typing clonal complex (e.g. CC‐ST9, CC‐ST232 or CC‐ST10), were PCR‐serotyped, evaluated for proteolytic activity, and used to challenge adipose fin‐clipped 4‐month old Atlantic salmon, coho salmon and rainbow trout via immersion. Findings showed US87‐RBT caused disease and mortality only in rainbow trout (e.g. 56.7% survival probability). US19‐COS and US62‐ATS caused more mortality in coho salmon and Atlantic salmon but also caused disease in both other host species, albeit to a lesser extent. Observed survival differences may be due to variant antigenic/virulence determinants as differences in serotype and proteolytic activity were discovered. Collectively, results highlight the intricacies of F. psychrophilum‐host interactions and provide further in vivo evidence that some F. psychrophilum MLST variants are host specific, which may have implications for the development of BCWD prevention and control strategies. [ABSTRACT FROM AUTHOR]

Widespread distribution of a highly pathogenic Edwardsiella ictaluri strain in farmed tilapia in northern Vietnam has recently been reported. The subsequent investigation noticed a disease outbreak occurred at five nearby tilapia farms with floating cages, in which the clinical signs of both edwardsiellosis and columnaris diseases were observed on the same infected fish and caused 65% to 85% fish mortality. Naturally diseased fish (n = 109) were sampled from the five infected farms for bacterial identification and conducting challenge tests. The two bacteria Edwardsiella ictaluri and Flavobacterium oreochromis were identified by a combination of biochemical tests, PCR and 16SrRNA sequencing methods. Experimental challenge tests on Nile tilapia resulted in the median lethal dose (LD50) of E. ictaluri and F. oreochromis at 70 CFU/fish by intraperitoneal (i.p.) injection and 3.6 × 106 CFU/mL by immersion, respectively. The experimentally co‐infected challenged fish exposed to LD50 doses resulted in 83% ± 6% mortality, with the infected fish exhibiting clinical signs of both edwardsiellosis and columnaris diseases, mimicking the naturally diseased fish. This finding suggests that the co‐infection of E. ictaluri and F. oreochromis may interact in a synergistic manner, to enhance the overall severity of the infection and elevates the need for efficient methods to control both pathogens. [ABSTRACT FROM AUTHOR]

Flavobacterium psychrophilum is a pathogenic bacterium affecting Chilean salmonid farms. High antigenic and genetic diversity exists among Chilean F. psychrophilum isolates, but the distribution thereof among farms is poorly understood. These epidemiological data are key for developing isolate‐specific vaccines. The present study isolated F. psychrophilum in diseased Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss) from five freshwater farms between 2018 and 2019. Each farm only raised one salmonid species and was geographically separated from and did not share culturing water with the other farms. Antigenic and genetic analyses were conducted to shed light on the possibility of isolates coexisting within the same farm during outbreaks. A total of 68 Chilean F. psychrophilum isolates were recovered from skin lesions, gills, fins, kidney and spleen of moribund and live fish. Among the 68 Chilean isolates, mPCR‐serotyping indicated three major serotypes (i.e. 23.5% type 0; 47.1% type 2; and 26.5% type 4) and, to a lesser degree, serotype 1 (2.9%). Sixteen antigenic groups were detected by slide agglutination. Genetic characterizations by 16S rRNA alleles identified 71% of the isolates as the virulent genogroup CSF259‐93 allele. A predominant serotype was associated with each farm, with types 0 and 4 related to Atlantic salmon and types 1 and 2 to rainbow trout. Notwithstanding, several antigenic groups coexisted within some farms. Likewise, the experimental intramuscular challenges (n = 20) demonstrated that the type‐2 isolates from rainbow trout were the most pathogenic among isolates recovered from infectious outbreaks in Atlantic salmon, especially as compared to those from types‐0 and ‐4. These results allow us to suggest that prevention measures, specifically vaccines, should be developed according to dominant isolates and with specificity to each farm, that is the use of autogenous or site‐specific vaccines. [ABSTRACT FROM AUTHOR]

Objective: Globally, flavobacteria (family Flavobacteriaceae and Weeksellaceae) are leading causes of disease‐related losses in fish‐farms and hatcheries. One route flavobacteria gain access to aquaculture facilities is via source water. Ultraviolet (UV) light treatment of source water has been effective in reducing the risk of disease outbreaks caused by nonflavobacteria; however, the UV dose required to inactivate flavobacteria has been understudied. The primary objective of this study was to examine the efficacy of UV light treatments for reducing the viability of fish‐pathogenic and fish‐associated Flavobacterium and Chryseobacterium species in a planktonic form. Methods: Sixty‐five flavobacterial isolates belonging to ten Flavobacterium spp. and Chryseobacterium spp. were exposed to a low (25 mJ/cm2) and high (126 mJ/cm2) dose of UV light via a collimating beam apparatus under in vitro conditions, after which treatment efficacy was determined via culture. Result: All assayed flavobacteria were reduced by an average of ~1000‐fold or ~100,000‐fold at the low and high UV doses, respectively; however, substantial differences in reduction at the same UV dose were noted among isolates of the same flavobacterial species, including F. psychrophilum, F. columnare, and F. oreochromis. In addition, F. psychrophilum multilocus sequence typing variants ST10 and ST78, which are two of the most widespread variants in the United States of America, were among the least susceptible to ultraviolet light. Conclusion: Overall, results demonstrate that viable flavobacteria can be reduced substantially by ultraviolet doses of 25–126 mJ/cm2, suggesting such treatments represent a promising tool for minimizing flavobacterial loads in hatcheries and aquaculture facilities, thereby enhancing biosecurity and reducing the risk of epizootics. Impact statementIn this study, ultraviolet light effectively reduced multiple fish disease‐causing flavobacteria under laboratory conditions. Thus, ultraviolet light treatment of water is a promising tool for reducing harmful flavobacteria in fish farms and hatcheries, thereby potentially improving fish health and aquaculture sustainability. [ABSTRACT FROM AUTHOR]

Background: Colorectal cancer (CRC) is known to present a distinct microbiome profile compared to healthy mucosa. Non‐targeted deep‐sequencing strategies enable nowadays full microbiome characterization up to species level. Aim: We aimed to analyze both bacterial and viral communities in CRC using these strategies. Materials & Methods: We analyzed bacterial and viral communities using both DNA and RNA deep‐sequencing (Novaseq) in colorectal tissue specimens from 10 CRC patients and 10 matched control patients. Following taxonomy classification using Kraken 2, different metrics for alpha and beta diversities as well as relative and differential abundance were calculated to compare tumoral and healthy samples. Results: No viral differences were identified between tissue types, but bacterial species Polynucleobacter necessarius had a highly increased presence for DNA in tumors (p = 0.001). RNA analyses showed that bacterial species Arabia massiliensis had a highly decreased transcription in tumors (p = 0.002) while Fusobacterium nucleatum transcription was highly increased in tumors (p = 0.002). Discussion: Sequencing of both DNA and RNA enables a wider perspective of micriobiome profiles. Lack of RNA transcription (Polynucleobacter necessarius) casts doubt on possible role of a microorganism in CRC. The association of F. nucleatum mainly with transcription, may provide further insights on its role in CRC. Conclusion: Joint assessment of the metagenome (DNA) and the metatranscriptome (RNA) at the species level provided a huge coverage for both bacteria and virus and identifies differential specific bacterial species as tumor associated. [ABSTRACT FROM AUTHOR]

Nodular gill disease (NGD) is an infectious condition characterized by proliferative gill lesions leading to respiratory problems, oxygen deficiency and mortality in fish. Globally, NGD primarily impacts freshwater salmonids in intensive aquaculture systems. In recent years, numerous outbreaks of severe gill disease have affected more than half of the larger rainbow trout (Oncorhynchus mykiss) farms in Switzerland, mainly during spring and early summer. Mortality has reached up to 50% in cases where no treatment was administered. Freshwater amoeba are the presumed aetiologic agent of NGD. The gross gill score (GS) categorising severity of gill pathology is a valuable first‐line diagnostic tool aiding fish farmers in identifying and quantifying amoebic gill disease (AGD) in farmed marine salmonids. In this study, the GS was adapted to the NGD outbreak in farmed trout in Switzerland. In addition to scoring disease severity, gill swabs from NGD‐affected rainbow trout were sampled and amoeba were cultured from these swabs. Morphologic and molecular methods identified six amoeba strains: Cochliopodium sp., Naegleria sp., Vannella sp., Ripella sp., Saccamoeba sp. and Mycamoeba sp. However, the importance of the different amoeba species for the onset and progression of NGD still has to be evaluated. This paper presents the first description of NGD with associated amoeba infection in farmed rainbow trout in Switzerland. [ABSTRACT FROM AUTHOR]

Flavobacterium psychrophilum affects many cultured fish species and is considered one of the most important bacterial pathogens causing substantial economic losses in salmonid aquaculture worldwide. Here, F. psychrophilum was identified by matrix‐assisted laser desorption ionization time‐of‐flight mass spectrometry (MALDI‐TOF MS) and nested PCR as the aetiological agent causing mortality in diseased juvenile Siberian sturgeons (Acipenser baerii) reared on a freshwater fish farm. Diseased sturgeons were lethargic and displayed dark skin pigmentation, increased mucus production and the presence of skin ulcerations and haemorrhages specially on the ventral side and the base of fins. The histological examination of fish revealed proliferative branchitis, ulcerative and necrotizing dermatitis and myositis, lymphoid tissue atrophy, liver and kidney degeneration and thrombosis. To the best of our knowledge, this is the first report describing the infection of Siberian sturgeons by F. psychrophilum. The detection of F. psychrophilum in diseased Siberian sturgeons and the description of the pathological findings observed during the outbreak may contribute to a better understanding of the bacterium pathogenicity and the range of fish species susceptible to infection. [ABSTRACT FROM AUTHOR]

This study evaluated the probiotic potential of the biofilm formed by the strain Pseudomonas sp. RGM2144 on rainbow trout survival. When challenged with the fish pathogen Flavobacterium psychrophilum, Pseudomonas sp. RGM2144 increased rainbow trout survival to 92.7 ± 1.2% (control: 35.3 ± 9.5%, p <.0001). The draft genome of Pseudomonas sp. RGM2144 is 6.8 Mbp long, with a completeness 100% and a contamination of 0.4%. The genome contains 6122 protein‐coding genes of which 3564 (~60%) have known functions. The genome and phylogeny indicate that Pseudomonas sp. RGM2144 is a new species in the Pseudomonas genus, with few virulence factors, plasmids, and genes associated with antimicrobial resistance, suggesting a non‐pathogenic bacterium with protective potential. In addition, the genome encodes for 11 secondary metabolite biosynthetic gene clusters that could be involved in the inhibition of F. psychrophilum. We suggest that Pseudomonas sp. RGM2144 may be applied as a probiotic in salmonid fish farming. [ABSTRACT FROM AUTHOR]

Flavobacterium psychrophilum is the causative agent of bacterial cold‐water disease (CWBD) and rainbow trout fry syndrome (RTFS), which affect salmonids. To better understand this pathogen and its interaction with the host during infection, including to support the development of resistant breeds and new vaccines and treatments, there is a pressing need for reliable and reproducible immersion challenge models that more closely mimic natural routes of infection. The aim of this present study was to evaluate a challenge model developed previously for rainbow trout for use in Atlantic salmon. First, preliminary challenges were conducted in Atlantic salmon (n = 120) and rainbow trout (n = 80) fry using two F. psychrophilum isolates collected from each fish species, respectively; fish had been pretreated with 200 mg/L hydrogen peroxide for 1 h. Thereafter, the main challenge was performed for just one F. psychrophilum isolate for each species (at 2 × 107 CFU/mL) but using larger cohorts (Atlantic salmon: n = 1187; rainbow trout: n = 2701). Survival in the main challenge was 81.2% in Atlantic salmon (21 days post‐challenge) and 45.3% in rainbow trout (31 days post‐challenge). Mortalities progressed similarly during the preliminary and main challenges for both species, demonstrating the reproducibility of this model. This is the first immersion challenge model of F. psychrophilum to be developed successfully for Atlantic salmon. [ABSTRACT FROM AUTHOR]

Flavobacteria are among the most important pathogens in freshwater salmonid aquaculture worldwide. Due to concerns regarding development of antibiotic resistance, phage therapy has been proposed as a solution to decrease pathogen load. However, application of phages is challenged by the development of phage resistance, and knowledge of the mechanisms and implications of phage resistance is therefore required. To study this, 27 phage‐resistant isolates of F. psychrophilum were genome sequenced and characterized to identify genetic modifications and evaluate changes in phenotypic traits, including virulence against rainbow trout. Phage‐resistant isolates showed reduction or loss of gliding motility, proteolytic activity, and adhesion to surfaces, and most isolates were completely non‐virulent against rainbow trout fry. Genomic analysis revealed that most phage‐resistant isolates had mutations in genes associated with gliding motility and virulence. Reversal of these mutations in a sub‐set of isolates led to regained motility, proteolytic activity, virulence and phage susceptibility. Although costly, the fast generation of phage resistance driven by single, reversible mutations likely represents a flexible and efficient phage defence mechanism in F. psychrophilum. The results further suggest that phage administration in aquaculture systems to prevent F. psychrophilum outbreaks selects for non‐virulent phage‐resistant phenotypes. [ABSTRACT FROM AUTHOR]

Calcium and manganese transporters play important roles in regulating Ca2+ and Mn2+ homeostasis in cells, which is necessary for the normal physiological activities of eukaryotes. Gdt1 and Pmr1 function as calcium/manganese transporters in the Golgi apparatus. However, the functions of Gdt1 and Pmr1 have not been previously characterized in the plant pathogenic fungus Fusarium graminearum. Here, we identified and characterized the biological functions of FgGdt1 and FgPmr1 in F. graminearum. Our study shows that FgGdt1 and FgPmr1 are both localized to the cis‐ and medial‐Golgi. Disruption of FgGdt1 or FgPmr1 in F. graminearum caused serious defects in vegetative growth, conidiation, sexual development and significantly decreased virulence in wheat but increased deoxynivalenol (DON) production. Importantly, FgGdt1 is involved in Ca2+ and Mn2+ homeostasis and the severe phenotypic defects of the ΔFggdt1 mutant were largely due to loss of FgGdt1 function in Mn2+ transportation. FgGdt1‐mCherry colocalizes with FgPmr1‐GFP at the Golgi, and FgGDT1 exerts its biological function upstream of FgPMR1. Taken together, our results collectively demonstrate that the cis‐ and medial‐Golgi‐localized proteins FgGdt1 and FgPmr1 regulate Ca2+ and Mn2+ homeostasis of the Golgi apparatus, and this function is important in modulating the growth, development, DON biosynthesis and pathogenicity of F. graminearum. [ABSTRACT FROM AUTHOR]

Lake whitefish (Coregonus clupeaformis; LWF) is an economically and ecologically valuable native species to the Great Lakes, but recent declines in their recruitment have generated significant concern about their future viability. Although studies have sought to identify factors contributing to declining recruitment, the potential role(s) of infectious diseases has not been thoroughly investigated. In 2018 and 2019, adult LWF were collected from Lakes Superior, Michigan, and Huron for clinical examination and bacteriological analyses. Herein, we describe the first isolation of Flavobacterium psychrophilum, aetiological agent of bacterial coldwater disease (BCWD) and rainbow trout fry syndrome (RTFS), from systemically infected adult LWF. Bacterial isolates were yellow‐orange, Gram‐negative, filamentous bacilli that were oxidase and catalase positive, and produced a flexirubin‐type pigment in 3% potassium hydroxide. Isolate identity was confirmed via F. psychrophilum‐specific PCR, and multilocus sequence typing revealed three new singleton sequence types (STs) that were distinct from all previously described F. psychrophilum STs. The prevalence of F. psychrophilum infections was 3.3, 1.7, and 0.0% in Lakes Superior, Michigan and Huron respectively. Findings illustrate the potential for F. psychrophilum to cause systemic infections in adult LWF and highlight the need for future studies to investigate the bacterium's potential role in declining LWF recruitment. [ABSTRACT FROM AUTHOR]

Two related salmonids, Arctic char (Salvelinus alpinus) and lake whitefish (Coregonus clupeaformis) sampled from the high Arctic region of Nunavut, Canada are anadromous fish, migrating annually from the same ice‐covered freshwater waterbodies to spend summers in the marine waters of the Arctic Ocean. Microbiota associated with the skin‐associated mucus undergo community change coincident with migration, and irrespective of this turnover, antibiotic resistance was detected in mixed bacterial cultures initiated with mucus samples. Although as expected most bacteria were unculturable, however, 5/7 isolates showed susceptibility to a panel of five common antibiotics. The fish were sampled under severe conditions and at remote locations far from human habitation. Regardless, two isolates, 'Carnobacterium maltaromaticum sm‐2' and 'Arthrobacter citreus sm', showed multi‐resistance to two or more antibiotics including ampicillin and streptomycin indicating multiple resistance genes. It is unknown if these fish bacteria have 'natural' resistance phenotypes or if resistance has been acquired. As result of these observations, we urge long‐term monitoring of drug‐resistant bacteria in the region and caution the assumption of a lack of drug‐resistant organisms even in such extreme environments. [ABSTRACT FROM AUTHOR]

Aquatic pathogens are a major concern for fish hatchery production, fisheries management, and conservation, and disease control needs to be addressed. Two important salmonid pathogens are Myxobolus cerebralis and Flavobacterium psychrophilum that cause whirling disease and bacterial coldwater disease (BCWD), respectively. Innate disease resistance is a potential option for reducing disease‐related mortality in hatchery‐reared rainbow trout (Oncorhynchus mykiss, Walbaum). Two experiments were conducted to assess pathogen resistance of first‐generation (F1) rainbow trout created by crossing M. cerebralis‐ and F. psychrophilum‐resistant strains. In the first experiment, we exposed two rainbow trout strains and one F1 cross to six treatments: control (no exposure), mock injection, F. psychrophilum only, M. cerebralis only, F. psychrophilum then M. cerebralis, and M. cerebralis then F. psychrophilum. Results indicated that the F1 cross was not resistant to either pathogen. In the second experiment, we exposed five rainbow trout strains and four rainbow trout crosses to F. psychrophilum. The second experiment indicated that at least one rainbow trout cross was F. psychrophilum‐resistant. Achieving dual resistance may be possible using selective breeding but only some multigenerational strains are suitable candidates for further evaluation. [ABSTRACT FROM AUTHOR]

Summary: Intensive aquaculture conditions expose fish to bacterial infections, leading to significant financial losses, extensive antibiotic use and risk of antibiotic resistance in target bacteria. Flavobacterium columnare causes columnaris disease in aquaculture worldwide. To develop a bacteriophage‐based control of columnaris disease, we isolated and characterized 126 F. columnare strains and 63 phages against F. columnare from Finland and Sweden in 2017. Bacterial isolates were virulent on rainbow trout (Oncorhynchus mykiss) and fell into four previously described genetic groups A, C, E and G, with genetic groups C and E being the most virulent. Phage host range studied against a collection of 227 bacterial isolates (from 2013 to 2017) demonstrated modular infection patterns based on host genetic group. Phages infected contemporary and previously isolated bacterial hosts, but bacteria isolated most recently were generally resistant to previously isolated phages. Despite large differences in geographical origin, isolation year or host range of the phages, whole‐genome sequencing of 56 phages showed high level of genetic similarity to previously isolated F. columnare phages (Ficleduovirus, Myoviridae). Altogether, this phage collection demonstrates a potential for use in phage therapy. [ABSTRACT FROM AUTHOR]

Summary: The physicochemical and microbiological parameters of postmortem crayfish stored at 4 °C and 25 °C were evaluated in order to reduce safety risks of crayfish from temperature abuse during transportation and storage. Results showed that hepatopancreas of postmortem crayfish had significantly higher contents of total volatile basic nitrogen, biogenic amines (BAs) and higher microbial loads than tail meat, regardless of storage temperature. Enterobacteriaceae and Pseudomonas counts reached log 6.41 log10 CFU g−1 and 6.31 log10 CFU g−1 in hepatopancreas of crayfish at 25 ℃ for 6 h. Putrescine and cadaverine were the main BAs in tail meat with levels of 28.18 ± 0.73 mg kg−1 and 187.32 ± 3.57 mg kg−1, respectively, whilst cadaverine, spermidine and spermine were the major BAs in hepatopancreas, reaching 283.45 ± 3.95 mg kg−1, 62.87 ± 9.02 mg kg−1 and 155.31 ± 4.55 mg kg−1, respectively, after postmortem storage at 25 °C for 12 h. With time increasing, Acinetobacter, Flavobacterium, Aeromonas and Chryseobacterium at 25 °C and Acinetobacter, Flavobacterium, Psychrobacter at 4 °C in tail meat as well as Bacteroides and Muribaculaceae at 25 °C, and Acinetobacter, Psychrobacter at 4 °C in hepatopancreas, gradually became the major genus at the end of storage. Based on the results of spoilage microorganisms and biogenic amines, crayfish stored at 4 °C and 25 °C could be edible within 24 h and 6 h, respectively. [ABSTRACT FROM AUTHOR]

Previously, rainbow trout fed deoxynivalenol (DON) or partially fed (pair‐fed) for 4 weeks before and during experimental infection with Flavobacterium psychrophilum had significantly decreased mortality rates. Similar results were obtained in the present study after 12 days, but not after 6 days, feeding 5 ppm DON or pair‐fed before infection. Furthermore, feeding 250 ppm chloroquine (CQ) also reduced mortality (p =.052) compared with controls and may have promise for treatment of some fish disease. Parallel groups of fish were maintained on the respective treatments for 15 days, with an additional group that was fasted, but were not infected to monitor autophagy. Fish that were fasted or fed DON had significantly increased LC3II in the liver and fasted fish had significantly decreased LC3II in muscle compared with controls using western blot. There was no difference in LC3II signal in the spleen of any treatment group. Fish that were fasted or pair‐fed had significant up‐regulation of the Atg genes atg4, atg7, lc3, gabarap and atg12 in muscle using quantitative PCR. Less alteration of Atg expression was seen in liver. Fish treated with CQ had significantly increased expression of atg4, becn1, lc3 and atg12 in the liver. Fish fed DON for 15 days had few alterations of Atg genes in either the liver or muscle. It is still not clear if autophagy is responsible for the resistance of rainbow trout fed DON, CQ or pair‐fed before F. psychrophilum infection. [ABSTRACT FROM AUTHOR]

This study examined supplementation of a novel yeast product containing Pichia guilliermondii on juvenile rainbow trout, Oncorhynchus mykiss, growth, and disease resistance. In two separate trials, fish (initial weight 6.2 and 10.6 g, respectively), were fed an extruded diet (41% digestible protein and 18% crude lipid) supplemented with 0.3 or 0.6% P. guilliermondii alone or 0.3% P. guilliermondii with a dietary gut supplement at 0.1%. Prior to the end of each trial, a subsample of fish from each tank was exposed by immersion to Yersinia ruckeri (Trial 1) or injection with Flavobacterium pyschrophilum (Trial 2). At the conclusion of the trials, supplementation of either P. guilliermondii or the gut supplement improved growth and food conversion efficiency in rainbow trout when fed for 16‐weeks. No effects of supplementation on mortality were observed. These results suggest potential as functional additives; however, additional investigation regarding the efficacy of whole‐cell and disrupted wall components of P. guilliermondii is needed to better assess their potential effects on rainbow trout immune responses. [ABSTRACT FROM AUTHOR]

Keywords: Flavobacterium psychrophilum; immersion challenge; immersion vaccination; proteomics; rainbow trout; skin mucus EN Flavobacterium psychrophilum immersion challenge immersion vaccination proteomics rainbow trout skin mucus 491 495 5 02/10/22 20220301 NES 220301 I Flavobacterium psychrophilum i is one of the most important pathogens affecting rainbow trout ( I Oncorhynchus mykiss i ) worldwide at the fry stage of their life cycle. Briefly, skin mucus was sampled by placing three fish (from each duplicate tank giving 2 pools of mucus/group) into a plastic bag containing 5 ml of Tris-buffered saline (TBS: 10 mM Tris base and 0.5 M NaCl, pH 7.5) and gently massaging for 2 min. In the current study, we examined the proteins of skin mucus of rainbow trout following immersion vaccination with I F. psychrophilum i . [Extracted from the article]

Histopathological assessments of young‐of‐the‐year (age‐0) Smallmouth Bass Micropterus dolomieu in the Susquehanna River drainage identified a high prevalence of the myxozoan Myxobolus inornatus. This myxozoan infects the connective tissue of the muscle below the skin but is sometimes observed in the esophagus and buccal cavity. In some instances, shallow infections cause breaks in the skin, which could increase the chance of opportunistic bacterial infections. Several microbial pathogens, including Flavobacterium columnare, Aeromonas spp., and Largemouth Bass virus, have also been cultured from clinically diseased young of year. A multiplex fluorescence in situ hybridization (FISH) assay was developed to determine potential colocalization of M. inornatus, Flavobacterium spp., and Aeromonas spp. infections. With FISH, 75% of age‐0 Smallmouth Bass exhibited M. inornatus infections, 10% had Aeromonas spp. infections, and 5% had Flavobacterium spp. infections, while 3% had coinfections with both bacterial species and M. inornatus. The results of the multiplex FISH assay revealed a low occurrence of coinfections of Flavobacterium spp. and/or Aeromonas spp. with M. inornatus in randomly sampled individuals. [ABSTRACT FROM AUTHOR]

Summary: Francisella novicida is a facultative intracellular pathogen and the causative agent of tularemia. Although cases of infection caused by exposure to contaminated water have been reported, its natural host and ecology in the environment remain unclear. In this study, we investigated in vitro the possibility that Paramecium bursaria may be a useful tool as a protist host model of F. novicida. Experimental infection with F. novicida resulted in a stable intracellular relationship within P. bursaria. This symbiotic intracellular relationship was not observed in experimental infections with other Francisella species and Legionella pneumophila. We found that F. novicida showed similar behaviour to that of the eukaryotic endosymbiont of P. bursaria, the green algae Chlorella, in the internalization process. In addition, stable intracellular localization of F. novicida was possible only when Chlorella was not present. Although we investigated the type VI secretion system of F. novicida as a candidate for the bacterial factor, we found that it was not involved in the establishment of an intracellular relationship with P. bursaria. These results suggested that P. bursaria is potentially a protist host model for F. novicida and may be a useful tool for understanding the relationship between protist hosts and their symbionts. [ABSTRACT FROM AUTHOR]

This study aimed to isolate waterborne heterotrophic organisms indigenous to water treatment processes with suitable ultraviolet (UV) dose–response profiles for estimation of the UV doses delivered by operational UV disinfection systems. The UV dose–response profiles of two isolated organisms, Flavobacterium succinicans and Sphingopyxis chilensis, were determined. S. chilensis has a UV dose–response with potential for use as an indigenous surrogate for Cryptosporidium inactivation, up to 2.2‐logs in the dose range 2–7 mJ/cm2 of monochromatic UV light at 253.7 nm. F. succinicans was more sensitive and tailing was observed above 4 mJ/cm2 making it unsuitable for UV dose verification above this dose. As a single species in the water samples used in this study, S. chilensis is unlikely to be present in sufficient numbers for routine use as an indigenous surrogate. However, the Sphingomonadaceae family to which it belongs is abundant in various drinking water sources and warrants further investigation. [ABSTRACT FROM AUTHOR]

Flavobacterium psychrophilum, the causative agent of bacterial coldwater disease (BCWD), causes significant economic losses worldwide, particularly in farmed Rainbow Trout Oncorhynchus mykiss. Over the last decade, multilocus sequence typing has revealed >30 clonal complexes (CCs) globally, comprised of >320 F. psychrophilum sequence types (STs). Despite the large number of CCs worldwide, CC‐ST10, which is currently the largest CC affecting Rainbow Trout, has been the primary focus of F. psychrophilum virulence studies, leaving the role of other CCs as primary causes of BCWD epizootics unclear. To this end, fingerling Rainbow Trout were experimentally challenged with F. psychrophilum strains belonging to the CC now recognized as the second largest in the world (CC‐ST191) alongside CC‐ST10 strains. Cumulative percent mortality was 100% in 7‐month‐old Rainbow Trout and between 27.8% and 61.1% in 8‐month‐old Rainbow Trout. All examined F. psychrophilum STs were virulent to Rainbow Trout, and no significant differences in virulence between CC‐ST10 and CC‐ST191 were detected. Due to their wide distribution and high pathogenic potential, both CC‐ST191 and CC‐ST10 F. psychrophilum strains are excellent candidates for further research aimed at preventing and controlling BCWD. [ABSTRACT FROM AUTHOR]

Summary: The fish pathogen Flavobacterium psychrophilum infects farmed salmonids worldwide, and application of bacteriophages has been suggested for controlling disease outbreaks in aquaculture. Successful application of phages requires detailed knowledge about the variability in phage susceptibility of the host communities. In this study, we analysed the genetic diversity of F. psychrophilum hosts and phages from the Baltic Sea area to identify genetic determinants of phage‐host interaction patterns. A host range analysis of 103 phages tested against 177 F. psychrophilum strains (18 231 phage–host interactions) identified nine phage clusters, infecting from 10% to 91% of the strain collection. The core genome‐based comparison of 35 F. psychrophilum isolates revealed an extremely low overall genomic diversity (>99.5% similarity). However, a small subset of 16 ORFs, including genes involved in the type IX secretion system (T9SS), gliding motility and hypothetical cell‐surface related proteins, exhibited a highly elevated genetic diversity. These specific genetic variations were linked to variability in phage infection patterns obtained from experimental studies, indicating that these genes are key determinants of phage susceptibility. These findings provide novel insights on the molecular mechanisms determining phage susceptibility in F. psychrophilum and emphasizes the importance of phages as drivers of core genomic diversity in this pathogen. [ABSTRACT FROM AUTHOR]

Iron is essential for growth and virulence in most pathogenic bacterial strains. In some cases, the hosts for these pathogenic bacteria develop specialized strategies to sequester iron and limit infectivity. This in turn may result in the invading pathogens utilizing high‐affinity iron transport mechanisms, such as the use of iron‐chelating siderophores, to extend beyond the host defences. Flavobacterium psychrophilum, the causative agent of bacterial coldwater disease (BCWD) in salmonids, relies on iron metabolism for infectivity, and the genome of the model CSF‐259‐93 strain has recently been made available. Further, this strain serves as a parent strain for a live‐attenuated vaccine strain, B.17, which has been shown to provide rainbow trout with protection against BCWD. To elucidate specific gene expression responses to iron metabolism and compare strain differences, both F. psychrophilum strains were grown under iron‐limiting conditions and 26 genes related to iron metabolism were mapped for 96 hr in culture via qPCR analyses. Results indicate increased production of the ferrous iron transport protein B (FITB; p =.008), and ferric receptor CfrA (FR 1; p =.012) in the wild‐type CSF‐259‐93 strain at 72 hr and 96 hr post‐exposure to iron‐limiting media. In the B.17 vaccine strain, siderophore synthase (SS) expression was found to be downregulated at 72 hr, in comparison with 0h (p =.018). When strains were compared, FITB (p =.021), FR1 (p =.009) and SS (p =.016) were also elevated in B.17 at 0 hr and TonB outer protein membrane receptor 1 (TBomr1; p =.005) had a lower expression at 96 hr. Overall, this study demonstrated strain‐related gene expression changes in only a fraction of the iron metabolism genes tested; however, results provide insight on potential virulence mechanisms and clarification on iron‐related gene expression for F. psychrophilum. [ABSTRACT FROM AUTHOR]

While Flavobacterium columnare is a well‐known fish pathogen that has been studied for over 100 years, columnaris disease remains poorly controlled and is in need of improved management solutions for U.S. catfish producers. Outside of therapeutants, the disease remains relatively unchallenged in the commercial catfish industry. Attempts to improve the management of this disease have led to the development of a live‐attenuated genetic group 2 F. columnare vaccine (hereafter, "17‐23 vaccine") administered via immersion to fingerling‐stage Channel Catfish Ictalurus punctatus. The 17‐23 vaccine was confirmed to protect against F. columnare in Channel Catfish fry during previous studies. In the present study, we investigated and compared the efficacy of the 17‐23 vaccine in Channel Catfish fingerlings versus nonvaccinated (control) fingerlings grown to food‐size fish in earthen ponds. Data were collected from the two treatments (n = 5 ponds/treatment) to evaluate impacts on survival, growth, feed conversion, antibody development, and economic benefit. There were no natural outbreaks of columnaris disease observed during the trial and no significant difference in survival of vaccinated versus control fish. Indirect enzyme‐linked immunosorbent assay results determined that the anti‐F. columnare 17‐23 antibody concentrations were significantly higher in vaccinated fish at 4 weeks postvaccination but not at 12 weeks. Vaccinated fish had a significantly higher average weight at harvest (mean ± SE = 357.5 ± 30 g) than control fish (289.3 ± 20 g), and the feed conversion ratio of vaccinated fish (1.35) was significantly better than that of control fish (2.13). Partial budget analysis of changing production from use of nonvaccinated fingerlings to use of 17‐23 vaccinated fingerlings for grow out resulted in a net benefit of US$1,443/ha. Based on these results, the vaccine appears to show potential economic benefit to producers but should be further substantiated under both research and commercial settings. [ABSTRACT FROM AUTHOR]

GldL is an inner‐membrane protein that is essential for the function of the type IX secretion system (T9SS) in Flavobacterium johnsoniae. The complex that it forms with GldM is supposed to act as a new rotary motor involved in the gliding motility of the bacterium. In the context of structural studies of GldL to gain information on the assembly and function of the T9SS, two camelid nanobodies were selected, produced and purified. Their interaction with the cytoplasmic domain of GldL was characterized and their crystal structures were solved. These nanobodies will be used as crystallization chaperones to help in the crystallization of the cytoplasmic domain of GldL and could also help to solve the structure of the complex using molecular replacement. [ABSTRACT FROM AUTHOR]

Summary: Sulfonolipids (SLs) are bacterial lipids that are structurally related to sphingolipids. Synthesis of this group of lipids seems to be mainly restricted to Flavobacterium, Cytophaga and other members of the phylum Bacteroidetes. These lipids have a wide range of biological activities: they can induce multicellularity in choanoflagellates, act as von Willebrand factor receptor antagonists, inhibit DNA polymerase, or function as tumour suppressing agents. In Flavobacterium johnsoniae, their presence seems to be required for efficient gliding motility. Until now, no genes/enzymes involved in SL synthesis have been identified, which has been limiting for the study of some of the biological effects these lipids have. Here, we describe the identification of the cysteate‐fatty acyl transferase Fjoh_2419 required for synthesis of the SL precursor capnine in F. johnsoniae. This enzyme belongs to the α‐oxoamine synthase family similar to serine palmitoyl transferases, 2‐amino‐3‐oxobutyrate coenzyme A ligase and 8‐amino‐7‐oxononanoate synthases. Expression of the gene fjoh_2419 in Escherichia coli caused the formation of a capnine‐derived molecule. Flavobacterium johnsoniae mutants deficient in fjoh_2419 lacked SLs and were more sensitive to many antibiotics. Mutant growth was not affected in liquid medium but the cells exhibited defects in gliding motility. [ABSTRACT FROM AUTHOR]

The continuing decline of many natural plant and animal populations emphasizes the importance of conservation strategies. Hybridization as a management tool has proven successful in introducing gene flow to small, inbred populations, but can be also associated with health risks. For example, hybridization can change susceptibility to infection in either direction due to heterosis (hybrid vigor) and outbreeding depression, but such health effects have rarely been considered in the genetic management of populations. Here, we investigated the effects of experimental outcrossing between the critically endangered Saimaa landlocked salmon (Salmo salar m. sebago) and the genetically more diverse Atlantic salmon (S. salar) on infection susceptibility. We exposed the parent populations and their hybrids to two parasites that commonly infect these species, the bacterium Flavobacterium columnare and the macroparasitic fluke Diplostomum pseudospathaceum. We found that landlocked salmon had lower survival during the bacterial epidemic, but higher resistance against the fluke, compared with Atlantic salmon. Hybrids showed intermediate survival and resistance, suggesting that hybridization decreased susceptibility to one parasite, but concurrently increased it to another. Our results emphasize the importance of considering health effects of different types of infections when employing hybridization for conservation. [ABSTRACT FROM AUTHOR]

Salmonid diseases caused by infections of Flavobacterium psychrophilum, the causative agent of bacterial coldwater disease, remain difficult to manage as novel, pathogenic strains continue to emerge in aquaculture settings globally. To date, much of the research regarding treatment options and vaccine development has focused on rainbow trout (Oncorhynchus mykiss), but other inland‐reared salmonids are also impacted by this Gram‐negative bacterium. As such, Atlantic salmon (Salmo salar) and brook trout (Salvelinus fontinalis) were injection‐challenged with a variety of previously reported F. psychrophilum strains isolated from disease diagnostic cases in salmonids, as well as a standard and well‐studied F. psychrophilum strain (CSF 259–93) known to be virulent in rainbow trout. In three separate virulence assessments (Trials A, B and C), strains US063 (isolated from lake trout; Salvelinus namaycush) and US149 (isolated from Atlantic salmon) caused a significantly higher cumulative per cent mortality (CPM) relative to other strains in Atlantic salmon (p <.001 for all trials), with US149 causing significantly greater mortality than US063 in Trials A (CPM 97% vs. 65%, p =.008) and B (CPM 96% ± 2.3% vs. 81.33% ± 4.8%, p =.014). Trial C used a lower dose (1.86 × 108 CFU/mL) for US149, resulting in a lower mortality (78.67% ± 9.33%) relative to Trials A and B. CSF259‐93 did not cause significant mortality in any trials. In brook trout, the strain 03–179 (originally isolated from steelhead trout; Oncorhynchus mykiss) was significantly more virulent than any other (CPM 100% ± 0%, p <.001), followed by US063 (73% ± 3.8%) and US149 (40% ± 6.1%,) respectively. Again, CSF259‐93 did not cause significant mortality relative to a mock challenge treatment. Results provide information about the applicability of strain selection in F. psychrophilum virulence testing in Atlantic salmon and brook trout, demonstrating the high virulence of US063 and US149 for these salmonid species. This information is applicable for the development of therapeutics and vaccines against F. psychrophilum infections and demonstrates the reproducibility of the experimental challenge model. [ABSTRACT FROM AUTHOR]

Flavobacterium columnare (Fc) is the causative agent for columnaris disease (CD) in several fish species and an emerging problem for rainbow trout aquaculture. We characterize the virulence phenotype of two Fc isolates, CSF‐298–10 and MS‐FC‐4, against trout from two sources, NCCCWA and a production stock (PS), at the eyed egg and alevin life stages. Immersion challenges demonstrated that NCCCWA eyed eggs were susceptible to the Fc isolate MS‐FC‐4 (>97% mortality) but no mortality was observed against PS eyed eggs. The CSF‐298–10 had little effect on any eyed eggs tested and was not highly virulent to any alevin till day six post‐hatch, up to 38% for NCCCWA and ~80% PS alevin. The MS‐FC‐4 strain produced ≥80% mortality any day an immersion challenge occurred post‐hatch. Significant difference in CFU counts was recorded between the Fc strains on 2 days post‐hatch immersion challenges. Counts for the NCCCWA alevin were 4.4 × 103 CFU/ml‐1 and 1.8 × 106 CFU/ml‐1 for the CSF‐298–10 strain and MS‐FC‐4 strain, respectively, and for the PS alevin CSF‐298–10 measured 9.9 × 101 CFU/ml‐1 and 3.8 × 105 CFU/ml‐1 for MS‐FC‐4. These two Fc isolates present stark differences in virulence phenotypes to both eyed eggs and alevin and present an interesting model system for virulence kinetics and potentially alternative pathogenic pathways. [ABSTRACT FROM AUTHOR]

Columnaris disease generates substantial losses of many freshwater fish species; one is the hybrid striped bass. The ubiquitous aquatic bacterium Flavobacterium columnare can be highly effective in biofilm formation on fish skin and gills. Previous research showed a difference between columnaris disease susceptibility of hybrid striped bass (Morone saxatilis × M. chrysops) and white bass (M. chrysops). To understand these differential susceptibilities and possible mucosal relationship, we assessed total bacterial growth and biofilm formation with mucus derived from each moronid parental species: white bass and striped bass (M. saxatilis). Differential susceptibility was confirmed of the other parent species, the striped bass (M. saxatilis). In addition to intraspecies investigations, individual hybrid striped bass mucosal affects were also studied for deferential responses to bacterial growth and biofilm formation. Species‐ and concentration‐dependent differences were detected in the total growth of the bacteria to host mucus. Our data suggest that bass mucus can significantly affect biofilm formation with the F. columnare isolate tested. There appears to be a correlation between the bacteria's response of growth and biofilms and bass species susceptibility. This study provides insight into our understanding of the host–pathogen interaction between F. columnare and moronids. [ABSTRACT FROM AUTHOR]

Burbot (Lota lota maculosa) are a potential new species for commercial aquaculture. As burbot culture expands, there is a need to further define pathogen susceptibility and characterize aspects of the burbot immune response in an effort to assess fish health. A recent clinical diagnostic case from juvenile burbot reared at a commercial production facility resulted in the isolation and identification of Flavobacterium columnare along with several Aeromonas spp. The F. columnare isolate was assigned to genetic group 1 via multiplex PCR, a genetic group commonly associated with columnaris disease cases in rainbow trout (Oncorhynchus mykiss). Virulence of the F. columnare isolate was assessed in vivo in both juvenile burbot and rainbow trout. Additionally, several of the Aeromonas sp. case isolates were identified via sequencing (16S rRNA, gyrB and rpoD) and a putative A. sobria isolate (BI‐3) was used to challenge burbot, along with a known virulent Aeromonas sp. (A141), but BI‐3 was not found to be virulent. Burbot were refractory to F. columnare when challenged by immersion, and it is likely that this is a secondary pathogen for burbot. Although refractory in burbot, the identified F. columnare isolate (BI‐1) was found to be virulent in rainbow trout. [ABSTRACT FROM AUTHOR]