Yersinia ruckeri isolates recovered from diseased Atlantic Salmon (Salmo salar) in Scotland are more diverse than those from Rainbow Trout (Oncorhynchus mykiss) and represent distinct subpopulations

Yersinia ruckeri is the etiological agent of enteric redmouth (ERM) disease of farmed salmonids. Enteric redmouth disease is traditionally associated with rainbow trout (Oncorhynchus mykiss, Walbaum), but its incidence in Atlantic salmon (Salmo salar) is increasing. Yersinia ruckeri isolates recovered from diseased Atlantic salmon have been poorly characterized, and very little is known about the relationship of the isolates associated with these two species. Phenotypic approaches were used to characterize 109 Y. ruckeri isolates recovered over a 14-year period from infected Atlantic salmon in Scotland; 26 isolates from infected rainbow trout were also characterized. Biotyping, serotyping, and comparison of outer membrane protein profiles identified 19 Y. ruckeri clones associated with Atlantic salmon but only five associated with rainbow trout; none of the Atlantic salmon clones occurred in rainbow trout and vice versa. These findings suggest that distinct subpopulations of Y. ruckeri are associated with each species. A new O serotype (designated O8) was identified in 56 biotype 1 Atlantic salmon isolates and was the most common serotype identified from 2006 to 2011 and in 2014, suggesting an increased prevalence during the time period sampled. Rainbow trout isolates were represented almost exclusively by the same biotype 2, serotype O1 clone that has been responsible for the majority of ERM outbreaks in this species within the United Kingdom since the 1980s. However, the identification of two biotype 2, serotype O8 isolates in rainbow trout suggests that vaccines containing serotypes O1 and O8 should be evaluated in both rainbow trout and Atlantic salmon for application in Scotland

Larva of the greater wax moth, Galleria mellonella, is a suitable alternative host for studying virulence of fish pathogenic Vibrio anguillarum

Background: Microbial diseases cause considerable economic losses in aquaculture and new infection control measures often rely on a better understanding of pathogenicity. However, disease studies performed in fish hosts often require specialist infrastructure (e.g., aquaria), adherence to strict legislation and do not permit high-throughput approaches; these reasons justify the development of alternative hosts. This study aimed to validate the use of larvae of the greater wax moth (Galleria mellonella) to investigate virulence of the important fish pathogen, Vibrio anguillarum. Results: Using 11 wild-type isolates of V. anguillarum, these bacteria killed larvae in a dose-dependent manner and replicated inside the haemolymph, but infected larvae were rescued by antibiotic therapy. Crucially, virulence correlated significantly and positively in larva and Atlantic salmon (Salmo salar) infection models. Challenge studies with mutants knocked out for single virulence determinants confirmed conserved roles in larva and fish infections in some cases (pJM1 plasmid, rtxA), but not all (empA, flaA, flaE). Conclusions: The G. mellonella model is simple, more ethically acceptable than experiments on vertebrates and, crucially, does not necessitate liquid systems, which reduces infrastructure requirements and biohazard risks associated with contaminated water. The G. mellonella model may aid our understanding of microbial pathogens in aquaculture and lead to the timely introduction of new effective remedies for infectious diseases, while adhering to the principles of replacement, reduction and refinement (3Rs) and considerably reducing the number of vertebrates used in such studies

Complete genome sequence of Streptococcus agalactiae strain 01173, isolated from Kuwaiti wild fish

© 2020 Santi et al. Here, we report the complete genome of piscine Streptococcus agalactiae 01173 serotype Ia, which was generated using long-read sequencing technology. The bacteria were isolated from wild fish displaying signs of streptococcosis, from a fish kill incident in Kuwait

Genetic and serological diversity of Flavobacterium psychrophilum isolates from salmonids in United Kingdom

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)5-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 profile observed within the F. psychrophilum isolates examined was PFGE cluster II − (GTG)5-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

The promise of whole genome pathogen sequencing for the molecular epidemiology of emerging aquaculture pathogens

Aquaculture is the fastest growing food-producing sector, and the sustainability of this industry is critical both for global food security and economic welfare. The management of infectious disease represents a key challenge. Here, we discuss the opportunities afforded by whole genome sequencing of bacterial and viral pathogens of aquaculture to mitigate disease emergence and spread. We outline, by way of comparison, how sequencing technology is transforming the molecular epidemiology of pathogens of public health importance, emphasizing the importance of community-oriented databases and analysis tools

Antimicrobial susceptibility of Flavobacterium psychrophilum isolates from the United Kingdom

Routine application of antimicrobials is the current treatment of choice for rainbow trout fry syndrome (RTFS) or bacterial coldwater disease (BCWD) caused by Flavobacterium psychrophilum. In this study, the antimicrobial susceptibilities of 133 F. psychrophilum isolates, 118 of which were from the UK, were evaluated by broth microdilution and disc diffusion methods following VET04-A2 and VET03-A guidelines of Clinical and Laboratory Standards Institute (CLSI), respectively. Isolates were categorised as wild type (fully susceptible, WT) or non-wild type (NWT) using normalised resistance interpretation (NRI) determined cut-off values (COWT). Broth microdilution testing showed that only 12% of UK isolates were WT to oxolinic acid (MIC COWT ≤0.25 mg L-1) and 42% were WT for oxytetracycline (MIC COWT ≤0.25 mg L-1). In contrast, all the isolates tested were WT (MIC COWT ≤2 mg L-1) for florfenicol, the main antimicrobial for RTFS control in the UK. Disc diffusion-based COWT values were ≥51 mm for 10 µg amoxicillin, ≥44 mm for 30 µg florfenicol, ≥30 mm for 2 µg oxolinic acid and ≥51 mm for 30 µg oxytetracycline. There was a high categorical agreement between the classifications of the isolates by two testing methods for florfenicol (100%), oxytetracycline (93%), and oxolinic acid (99%)

High Prevalence of Multidrug-Tolerant Bacteria and Associated Antimicrobial Resistance Genes Isolated from Ornamental Fish and Their Carriage Water

Background: Antimicrobials are used to directly control bacterial infections in pet (ornamental) fish and are routinely added to the water these fish are shipped in to suppress the growth of potential pathogens during transport. Methodology/Principal Findings: To assess the potential effects of this sustained selection pressure, 127 Aeromonas spp. isolated from warm and cold water ornamental fish species were screened for tolerance to 34 antimicrobials. Representative isolates were also examined for the presence of 54 resistance genes by a combination of miniaturized microarray and conventional PCR. Forty-seven of 94 Aeromonas spp. isolates recovered from tropical ornamental fish and their carriage water were tolerant to >= 15 antibiotics, representing seven or more different classes of antimicrobial. The quinolone and fluoroquinolone resistance gene, qnrS2, was detected at high frequency (37% tested recent isolates were positive by PCR). Class 1 integrons, IncA/C broad host range plasmids and a range of other antibiotic resistance genes, including floR, blaTEM21, tet(A), tet(D), tet(E), qacE2, sul1, and a number of different dihydrofolate reductase and aminoglycoside transferase coding genes were also detected in carriage water samples and bacterial isolates. Conclusions: These data suggest that ornamental fish and their carriage water act as a reservoir for both multi-resistant bacteria and resistance genes

Comparative metagenomics reveals a diverse range of antimicrobial resistance genes in effluents entering a river catchment.

The aquatic environment has been implicated as a reservoir for antimicrobial resistance genes (ARGs). In order to identify sources that are contributing to these gene reservoirs, it is crucial to assess effluents that are entering the aquatic environment. Here we describe a metagenomic assessment for two types of effluent entering a river catchment. We investigated the diversity and abundance of resistance genes, mobile genetic elements (MGEs) and pathogenic bacteria. Findings were normalised to a background sample of river source water. Our results show that effluent contributed an array of genes to the river catchment, the most abundant being tetracycline resistance genes tetC and tetW from farm effluents and the sulfonamide resistance gene sul2 from wastewater treatment plant (WWTP) effluents. In nine separate samples taken across 3 years, we found 53 different genes conferring resistance to seven classes of antimicrobial. Compared to the background sample taken up river from effluent entry, the average abundance of genes was three times greater in the farm effluent and two times greater in the WWTP effluent. We conclude that effluents disperse ARGs, MGEs and pathogenic bacteria within a river catchment, thereby contributing to environmental reservoirs of ARGs.This research was funded by the Centre for Environment, Fisheries and Aquaculture Science, GlaxoSmithKline and the Biotechnology and Biological Sciences Research Council. Metagenomic sequencing was performed by the Sequencing Service and Wellcome Trust Biomedical Informatics Hub, Exeter (UK).This is the author accepted manuscript. It is currently under an indefinite embargo pending publication by IWA Publishing