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Flavobacterium psychrophilum is one of the most important bacterial pathogens affecting cultured rainbow trout (Oncorhynchus mykiss) and is increasingly causing problems in Atlantic salmon (Salmo salar L.) hatcheries. Little is known about the heterogeneity of F. psychrophilum isolates on UK salmonid farms. A total of 315 F. psychrophilum isolates, 293 of which were collected from 27 sites within the UK, were characterised using four genotyping methods and a serotyping scheme. A high strain diversity was identified among the isolates with 54 pulsotypes, ten (GTG)-PCR types, two 16S rRNA allele lineages, seven plasmid profiles and three serotypes. Seven PFGE groups and 27 singletons were formed at a band similarity of 80%. PFGE group P (n=75) was found to be numerically predominant in eight sites within the UK. Two major PFGE clusters and 13 outliers were found at the band similarity of 40%. The predominant profileobserved within the F. psychrophilum isolates examined was PFGE cluster II - (GTG)-PCR type r1-16S rRNA lineage II - serotype Th (70/156 isolates examined, 45%). Co-existence of genetically and serologically heterogeneous isolates within each farm was detected, confounding the ability to control RTFS outbreaks. The occurrence over time (up to 11 years) of F. psychrophilum pulsotypes in three representative sites (Scot I, Scot III and Scot V) within Scotland was examined, potentially providing important epidemiological data for farm management and the development of site-specific vaccines.

Rainbow trout fry syndrome (RTFS) is a disease caused by the Gram-negative bacterium Flavobacterium psychrophilum, responsible for significant economic losses in salmonid aquaculture worldwide. The diversity of F. psychrophilum isolates and the inherent difficulties in vaccinating juvenile fish has hampered the development of a vaccine for RTFS. Disease episodes tend to occur between 10–14 °C with necrotic lesions often seen on the skin surrounding the dorsal fin and tail. At present no commercial vaccines are available for RTFS in the UK, leaving antibiotics as the only course of action to control disease outbreaks. The current work was performed as a pilot study to assess the efficacy of a polyvalent, whole cell vaccine containing formalin-inactivated F. psychrophilum, to induce protective immunity in rainbow trout fry. Duplicate groups of 30 trout (5 g) were immersed in 1 L of the vaccine for 30 s. Samples were taken 4 h, day 2 and 7 post-vaccination (pv) of skin mucus, tissues for histology and gene expression analysis; serum and histology samples were taken 6 weeks pv. A booster vaccination was given at 315 degree days (dd) also by immersion. Challenge was by immersion with a heterologous isolate of F. psychrophilum 630 dd post primary vaccination. The vaccine provided significant protection to the trout fry with a RPS of 84% (p < 0.0001). Detection of increased numbers of IgT positive cells in systemic organs, up-regulation of IgT expression in hind-gut and an increase in total IgT in serum was observed in vaccinated fish; however a functional role of IgT in the observed protection remains to be demonstrated.Electronic supplementary materialThe online version of this article (doi:10.1186/s13567-017-0448-z) contains supplementary material, which is available to authorized users.

It is well known that lysine acetylation (Kace) modification is a common post-translational modification (PTM) that plays an important role in multiple biological and pathological functions in bacteria. However, few studies have focused on lysine acetylation modification in aquatic pathogens to date. In this study, the acetylome profiling of fish pathogen, Vibrio alginolyticus was investigated by combining affinity enrichment with LC MS/MS. A total of 2883 acetylation modification sites on 1178 proteins in this pathogen were identified. The Kace modification of several selected proteins were further validated by Co-immunocoprecipitation combined with Western blotting. Bioinformatics analysis showed that seven conserved motifs can be enriched among Kace peptides, and many of them were significantly enriched in metabolic processes such as biosynthesis of secondary metabolites, microbial metabolism in diverse environments, and biosynthesis of amino acids, which was similar to data previously published for V. parahaemolyticus. Moreover, we found at least 102 acetylation modified proteins predicted as virulence factors, which indicate the important role of PTM on bacterial virulence. In general, our results provide a promising starting point for further investigations of the biological role of lysine acetylation on bacterial virulence in V. alginolyticus.

Efficacy of an inactivated whole-cell injection vaccine for nile tilapia, Oreochromis niloticus (L), against multiple isolates of Francisella noatunensis subsp. orientalis from diverse geographical regions.

Vibrio alginolyticus, a bacterial pathogen in fish and humans, expresses a type III secretion system (T3SS) that is critical for pathogen virulence and disease development. However, little is known about the associated effectors (T3SEs) and their physiological role. In this study, the T3SE gene hopPmaJ (hop) was cloned from V. alginolyticus wild-type strain HY9901 and the mutant strain HY9901Δhop was constructed by the in-frame deletion method. The results showed that the deduced amino acid sequence of V. alginolyticus HopPmaJ shared 78-98% homology with other Vibrio spp. In addition, the HY9901Δhop mutant showed an attenuated swarming phenotype and a 2600-fold decrease in the virulence to grouper. However, the HY9901Δhop mutant showed no difference in morphology, growth, biofilm formation and ECPase activity. Finally, grouper vaccinated via intraperitoneal (IP) injection with HY9901Δhop induced a high antibody titer with a relative percent survival (RPS) value of 84% after challenging with the wild-type HY9901. Real-time PCR assays showed that vaccination with HY9901Δhop enhanced the expression of immune-related genes, including MHC-Iα, MHC-IIα, IgM, and IL-1β after vaccination, indicating that it is able to induce humoral and cell-mediated immune response in grouper. These results demonstrate that the HY9901Δhop mutant could be used as an effective live vaccine to combat V. alginolyticus in grouper.

Flavobacterium psychrophilum is the causative agent of Rainbow Trout Fry Syndrome which has had a major impact on global salmonid aquaculture. Recent outbreaks in Atlantic salmon in Scotland and Chile have added to the need for a vaccine to protect both salmon and trout. At present no licensed vaccines are available in Europe, leaving antibiotics as the only course of action to contain disease outbreaks. Outbreaks generally occur in fry at temperatures between 10 and 15 °C. Recently outbreaks in larger fish have given added impetus to the development of a vaccine which can provide long term protection from this highly heterogeneous pathogen. Most fish injectable vaccines are formulated with oil emulsion adjuvants to induce strong and long lasting immunity, but which are known to cause side effects. Alternative adjuvants are currently sought to minimise these adverse effects. The current study was performed to assess the efficacy of a polyvalent, whole cell vaccine containing formalin-inactivated F. psychrophilum to induce protective immunity in Atlantic salmon. The vaccine was formulated with an adjuvant containing squalene and aluminium hydroxide, and was compared to a vaccine formulated with a traditional oil adjuvant, Montanide ISA 760VG, and a non-adjuvanted vaccine. Duplicate groups of salmon (23.5 ± 6.8 g) were vaccinated with each of the vaccine formulations or phosphate buffered saline by intraperitoneal injection. Fish were challenged by intramuscular injection with F. psychrophilum six weeks post-vaccination to test the efficacy of the vaccines. Cumulative mortality reached 70% in the control salmon, while the groups of salmon that received vaccine had significantly lower mortality than the controls (p = 0.0001), with no significant difference in survival between vaccinated groups. The squalene/alum adjuvant was safe, more readily metabolised by the fish and induced less histopathological changes than the traditional oil adjuvant.

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