Salmonid Rickettsial Septicemia (SRS) disease dynamics and Atlantic salmon immune response to Piscirickettsia salmonis LF-89 and EM-90 co-infection.

In Chile, Piscirickettsia salmonis contains two genetically isolated genogroups, LF-89 and EM-90. However, the impact of a potential co-infection with these two variants on Salmonid Rickettsial Septicemia (SRS) in Atlantic salmon (Salmo salar) remains largely unexplored. In our study, we evaluated the effect of P. salmonis LF-89-like and EM-90-like co-infection on post-smolt Atlantic salmon after an intraperitoneal challenge to compare changes in disease dynamics and host immune response. Co-infected fish had a significantly lower survival rate (24.1%) at 21 days post-challenge (dpc), compared with EM-90-like single-infected fish (40.3%). In contrast, all the LF-89-like single-infected fish survived. In addition, co-infected fish presented a higher presence of clinical lesions than any of the single-infected fish. The gene expression of salmon immune-related biomarkers evaluated in the head kidney, spleen, and liver showed that the EM-90-like isolate and the co-infection induced the up-regulation of cytokines (e.g., il-1ß, ifnγ, il8, il10), antimicrobial peptides (hepdicin) and pattern recognition receptors (PRRs), such as TLR5s. Furthermore, in serum samples from EM-90-like and co-infected fish, an increase in the total IgM level was observed. Interestingly, specific IgM against P. salmonis showed greater detection of EM-90-like antigens in LF-89-like infected fish serum (cross-reaction). These data provide evidence that P. salmonis LF-89-like and EM-90-like interactions can modulate SRS disease dynamics in Atlantic salmon, causing a synergistic effect that increases the severity of the disease and the mortality rate of the fish. Overall, this study contributes to achieving a better understanding of P. salmonis population dynamics.

Chitinolytic enzymes contribute to the pathogenicity of Aliivibrio salmonicida LFI1238 in the invasive phase of cold-water vibriosis.

BACKGROUND: Aliivibrio salmonicida is the causative agent of cold-water vibriosis in salmonids (Oncorhynchus mykiss and Salmo salar L.) and gadidae (Gadus morhua L.). Virulence-associated factors that are essential for the full spectrum of A. salmonicida pathogenicity are largely unknown. Chitin-active lytic polysaccharide monooxygenases (LPMOs) have been indicated to play roles in both chitin degradation and virulence in a variety of pathogenic bacteria but are largely unexplored in this context. RESULTS: In the present study we investigated the role of LPMOs in the pathogenicity of A. salmonicida LFI238 in Atlantic salmon (Salmo salar L.). In vivo challenge experiments using isogenic deletion mutants of the two LPMOs encoding genes AsLPMO10A and AsLPMO10B, showed that both LPMOs, and in particular AsLPMO10B, were important in the invasive phase of cold-water vibriosis. Crystallographic analysis of the AsLPMO10B AA10 LPMO domain (to 1.4 Å resolution) revealed high structural similarity to viral fusolin, an LPMO known to enhance the virulence of insecticidal agents. Finally, exposure to Atlantic salmon serum resulted in substantial proteome re-organization of the A. salmonicida LPMO deletion variants compared to the wild type strain, indicating the struggle of the bacterium to adapt to the host immune components in the absence of the LPMOs. CONCLUSION: The present study consolidates the role of LPMOs in virulence and demonstrates that such enzymes may have more than one function.

Freshwater salmon aquaculture in Chile and transferable antimicrobial resistance.

Large amounts of antimicrobials are used in salmonid aquaculture in Chile. Most are used in marine aquaculture, but appreciable amounts are also employed in freshwater aquaculture. Much research and many publications have examined transferable antimicrobial resistance in bacteria isolated from marine salmon farms, but much less attention has been paid to this area in freshwater salmon farming. A recent paper by Domínguez et al. (2019) has as least in part remedied this situation. We now comment on some of its interpretations and have attempted to point out its areas of strength and weakness in light of the published scientific literature. Seen in this setting, the important results presented by Domínguez et al. (2019) underline the need for increased awareness of the challenge to animal and human health posed by excessive use of antimicrobials in aquaculture.

Repeated Isolation of Extended-Spectrum-ß-Lactamase-Positive Escherichia coli Sequence Types 648 and 131 from Community Wastewater Indicates that Sewage Systems Are Important Sources of Emerging Clones of Antibiotic-Resistant Bacteria.

Antibiotic resistance in bacteria is an emerging problem globally. Resistant bacteria are found in human and animal microbiota, as well as in the environment. Wastewater receives bacteria from all these sources and thus can provide a measurement of abundance and diversity of antibiotic-resistant bacteria circulating in communities. In this study, water samples were collected from a wastewater pump station in a Norwegian suburban community over a period of 15 months. A total of 45 daily samples were cultured and analyzed for the presence of Escherichia coli Eighty E. coli-like colonies were collected from each daily sample and then phenotyped and analyzed for antibiotic resistance using the PhenePlate-AREB system. During the sampling period, two unique E. coli phenotypes with resistance to cefotaxime and cefpodoxime indicating carriage of extended-spectrum ß-lactamases (ESBL) were observed repeatedly. Whole-genome sequencing of 15 representative isolates from the two phenotypes identified these as two distinct clones belonging to the two globally spread E. coli multilocus sequence types (STs) ST131 and ST648 and carrying blaCTX-M-15 The number of ESBL-positive E. coli strains in the community wastewater pump station was 314 of 3,123 (10%) analyzed E. coli strains. Of the ESBL-positive isolates, 37% belonged to ST648, and 7% belonged to ST131. Repeated findings of CTX-M-15-positive ST648 and ST131 over time indicate that these STs are resident in the analyzed wastewater systems and/or circulate abundantly in the community.

An outbreak of acute disease and mortality in Atlantic salmon (Salmo salar) post-smolts in Norway caused by Tenacibaculum dicentrarchi.

An outbreak of disease characterized by skin ulcers, fin rot and mortality was observed a few days after the transfer of Atlantic salmon (Salmo salar) from a freshwater smolt production facility to a land-based seawater post-smolt site. Dead and moribund fish had severe skin and muscle ulcers, often 2-6 cm wide, particularly caudal to the pectoral fins. Microscopic examination of smears from ulcers and head kidney identified long, slender Gram-negative rods. Histopathological analysis revealed abundance of long, slender Tenacibaculum-like bacteria in ulcers and affected fins. Genetic characterization using multilocus sequence analysis (MLSA) of seven housekeeping genes, including atpA, dnaK, glyA, gyrB, infB, rlmN and tgt, revealed that the isolates obtained during the outbreak were all clustered with the Tenacibaculum dicentrarchi-type strain (USC39/09T ) from Spain. Two bath challenge experiments with Atlantic salmon and an isolate of T. dicentrarchi from the outbreak were performed. No disease or mortality was observed in the first trial. In the second trial with a higher challenge dose of T. dicentrarchi and longer challenge time, we got 100% mortality within 48 hr. This is the first reported outbreak of disease caused by T. dicentrarchi in Norwegian farmed Atlantic salmon.

Development of piscirickettsiosis in Atlantic salmon (Salmo salar L.) smolts after intraperitoneal and cohabitant challenge using an EM90-like isolate: A comparative study.

Piscirickettsiosis, caused by the intracellular Gram-negative bacteria Piscirickettsia salmonis, is at present the most devastating disease in the Chilean salmon industry. The aim of this study was to analyse disease development after challenge with a P. salmonis strain (EM90-like) under a controlled environment by comparing intraperitoneal challenge with cohabitation challenge. The P. salmonis EM90-like isolate was cultured in a liquid medium for the challenge of 400 Atlantic salmon (Salmo salar) smolts. Cumulative mortality was registered, necropsy was performed, and bacterial distribution in the tissues and histopathological changes were analysed. The results revealed a similar progression of the disease for the two different challenge models. Pathological and histopathological changes became more visible during the development of the clinical phase of the disease. Bacterial DNA was identified in all the analysed tissues indicating a systemic infection. Bacterial tropism to visceral organs was demonstrated by real-time quantitative PCR and immunohistochemistry. Better knowledge of disease development during P. salmonis infection may contribute to further development of challenge models that mimic the field situation during piscirickettsiosis outbreaks. The models can be used to develop and test future preventive measures against the disease.

Comparative evaluation of experimental challenge by intraperitoneal injection and cohabitation of Atlantic salmon (Salmo salar L) after vaccination against Piscirickettsia salmonis (EM90-like).

The Chilean aquaculture has been challenged for years by piscirickettsiosis. A common prophylactic measurement to try to reduce the impact from this disease is vaccination, but the development of vaccines that induce satisfactory protection of the fish in the field has so far not been successful. Experimental challenge models are used to test vaccine efficacy. The aim of this study was to evaluate the performance of experimental vaccines after challenge by the two most widely used challenge routes, intraperitoneal injection and cohabitation. A total of 1,120 Atlantic salmon were vaccinated with non-commercial experimental vaccines with increasing amounts of an inactivated Piscirickettsia salmonis EM90-like isolate. Differences in mortality, macroscopic and microscopic pathological changes, bacterial load and immune gene expression were compared after challenge by different routes. The results revealed a similar progression of the diseases after challenge by both routes and no gross differences reflecting the efficacy of the vaccines could be identified. The analysis of the immune genes suggests a possible suppression of the cellular immunity by CD8 T cell and with this stimulation of bacterial survival and replication. Comparative studies of experimental challenge models are valuable with regard to identifying the best model to mimic real-life conditions and vaccines' performance.

Aliivibrio salmonicida requires O-antigen for virulence in Atlantic salmon (Salmo salar L.).

Aliivibrio salmonicida is the causative agent of cold-water vibriosis, a hemorrhagic septicemia of salmonid fish. The bacterium has been shown to rapidly enter the fish bloodstream, and proliferation in blood is seen after a period of latency. Although the pathogenesis of the disease is largely unknown, shedding of high quantities of outer-membrane complex VS-P1, consisting of LPS and a protein moiety, has been suggested to act as decoy and contribute to immunomodulation. To investigate the role of LPS in the pathogenesis, we constructed O-antigen deficient mutants by knocking out the gene encoding O-antigen ligase waaL. As this gene exists in two copies in the Al. salmonicida genome, we constructed single and double in-frame deletion mutants to explore potential effects of copy number variation. Our results demonstrate that the LPS structure of Al. salmonicida is essential for virulence in Atlantic salmon. As the loss of O-antigen did not influence invasive properties of the bacterium, the role of LPS in virulence applies to later stages of the pathogenesis. One copy of waaL was sufficient for O-antigen ligation and virulence in experimental models. However, as a non-significant decrease in mortality was observed after immersion challenge with a waaL single mutant, it is tempting to suggest that multiple copies of the gene are beneficial to the bacterium at lower challenge doses. The loss of O-antigen was not found to affect serum survival in vitro, but quantification of bacteria in blood following immersion challenge suggested a role in in vivo survival. Furthermore, fish challenged with the waaL double mutant induced a more transient immune response than fish challenged with the wild type strain. Whether the reduction in virulence following the loss of waaL is caused by altered immunomodulative properties or impaired survival remains unclear. However, our data demonstrate that LPS is crucial for development of disease.

Multilocus sequence analysis revealed a high genotypic diversity of Aeromonas hydrophila infecting fish in Uganda.

A multilocus sequence analysis (MLSA) was carried out to delineate Aeromonas hydrophila from fish in Uganda. Five housekeeping genes including recA, gyrB, metG, gltA and pps; and the 16S rRNA gene were amplified and sequenced from a total of nine A. hydrophila isolates. The obtained sequences were edited, and consensus sequences generated for each gene locus. The housekeeping gene sequences were concatenated and phylogenetic analysis performed in MEGA version 7.0.2. Pairwise distances ranged from 0.000 to 0.118, highest within the gltA gene locus and lowest within the 16S rRNA gene. The average evolutionary diversity within isolates from the same source ranged between 0.002 and 0.037, and it was 0.033 between the different sources. Similar tree topologies were obtained from the different gene loci with recA, metG and gyrB being more consistent in discriminating isolates according to sources while the 16S rRNA gene had the lowest resolution. The concatenated tree had the highest discriminatory power. This study revealed that A. hydrophila strains infecting fish in Uganda are of diverse genotypes suggesting different sources of infection in a given outbreak. Efforts to minimize spread of the bacteria across sources should be emphasized to control infections of mixed genotypes.

A unique role of flagellar function in Aliivibrio salmonicida pathogenicity not related to bacterial motility in aquatic environments.

Aliivibrio salmonicida is the causative agent of cold-water vibriosis, a septicemia of farmed salmonid fish. The mechanisms of disease are not well described, and few virulence factors have been identified. However, a requirement for motility in the pathogenesis has been reported. Al. salmonicida is motile by the means of lophotrichous polar flagella, consisting of multiple flagellin subunits that are expressed simultaneously. Here we show that flagellin subunit FlaA, but not FlaD, is of major importance for motility in Al. salmonicida. Deletion of flaA resulted in 62% reduction in motility, as well as a reduction in the fraction of flagellated cells and number of flagella per cell. Similarly, deletion of the gene encoding motor protein motA gave rise to an aflagellate phenotype and cessation of motility. Surprisingly, we found that Al. salmonicida does not require motility for invasion of Atlantic salmon. Nevertheless, in-frame deletion mutants defective of motA and flaA were less virulent in Atlantic salmon challenged by immersion, whereas an effect on virulence after i.p. challenge was only seen for the latter. Our results indicate a complex requirement for motility and/or flagellation in the pathogenesis of cold-water vibriosis, but the mechanisms involved remain unknown. We hypothesize that the differences in virulence observed after immersion and i.p. challenge are related to the immune response of the host.

Vibrio tapetis from wrasse used for ectoparasite bio-control in salmon farming: phylogenetic analysis and serotyping.

So-called 'cleaner fish', including various wrasse (Labridae) species, have become increasingly popular in Norwegian salmon farming in recent years for biocontrol of the salmon louse Lepeophtheirus salmonis. Cleaner fish mortalities in salmon farms are, however, often high. Various bacterial agents are frequently associated with episodes of increased cleaner fish mortality, and Vibrio tapetis is regularly cultured from diseased wrasse. In the present study, we investigated the genetic relationships among 54 V. tapetis isolates (34 from wrasse species) by multilocus sequence analysis (MLSA; rpoD, ftsZ, pyrH, rpoA and atpA). In the resulting phylogenetic tree, all wrasse isolates belonged to sub-clusters within V. tapetis subsp. tapetis. Slide agglutination testing further confirmed the complete dominance amongst these isolates of 4 O-antigen serotypes, designated here as V. tapetis subsp. tapetis serotypes O1, O3, O4 and O5, respectively. A pilot challenge trial using serotypes O3, O4 and O5 did not indicate high pathogenicity towards ballan wrasse Labrus bergylta, thus questioning the role of V. tapetis as a primary pathogen of this fish species.

Co-cultivation and transcriptome sequencing of two co-existing fish pathogens Moritella viscosa and Aliivibrio wodanis.

BACKGROUND: Aliivibrio wodanis and Moritella viscosa have often been isolated concurrently from fish with winter-ulcer disease. Little is known about the interaction between the two bacterial species and how the presence of one bacterial species affects the behaviour of the other. RESULTS: The impact on bacterial growth in co-culture was investigated in vitro, and the presence of A. wodanis has an inhibitorial effect on M. viscosa. Further, we have sequenced the complete genomes of these two marine Gram-negative species, and have performed transcriptome analysis of the bacterial gene expression levels from in vivo samples. Using bacterial implants in the fish abdomen, we demonstrate that the presence of A. wodanis is altering the gene expression levels of M. viscosa compared to when the bacteria are implanted separately. CONCLUSIONS: From expression profiling of the transcriptomes, it is evident that the presence of A. wodanis is altering the global gene expression of M. viscosa. Co-cultivation studies showed that A. wodanis is impeding the growth of M. viscosa, and that the inhibitorial effect is not contact-dependent.

Phylogenetic analysis and serotyping of Vibrio splendidus-related bacteria isolated from salmon farm cleaner fish.

Cleaner fish, i.e. various wrasse (Labridae) species and lumpsucker Cyclopterus lumpus, are to an increasing extent used for biocontrol of the salmon louse Lepeophtheirus salmonis in European salmon farming. Although efficient de-licers, cleaner fish mortality levels in salmon farms are often high. Bacterial infections are common, and Vibrio splendidus-related strains are frequently identified during diagnostic investigations. The population structure of 112 V. splendidus-related isolates, derived primarily from wrasse species, was investigated by means of multilocus sequence analysis using 5 housekeeping genes (rpoD, ftsZ, pyrH, rpoA and atpA). Most isolates were found to be closely related to the V. splendidus type strain, yet displayed extensive genetic microdiversity. Slide agglutination testing using polyclonal rabbit antisera further indicated O-antigen variability. Intra-outbreak genetic and antigenic diversity suggests direct infection from seawater, rather than fish-to-fish transmission, as the main route of infection. The variable nature of isolates involved complicates qualified selection of representative candidate strains, e.g. for infection and vaccine trials.

Antimicrobial resistance and antimicrobial resistance genes in marine bacteria from salmon aquaculture and non-aquaculture sites.

Antimicrobial resistance (AR) detected by disc diffusion and antimicrobial resistance genes detected by DNA hybridization and polymerase chain reaction with amplicon sequencing were studied in 124 marine bacterial isolates from a Chilean salmon aquaculture site and 76 from a site without aquaculture 8 km distant. Resistance to one or more antimicrobials was present in 81% of the isolates regardless of site. Resistance to tetracycline was most commonly encoded by tetA and tetG; to trimethoprim, by dfrA1, dfrA5 and dfrA12; to sulfamethizole, by sul1 and sul2; to amoxicillin, by blaTEM ; and to streptomycin, by strA-strB. Integron integrase intl1 was detected in 14 sul1-positive isolates, associated with aad9 gene cassettes in two from the aquaculture site. intl2 Integrase was only detected in three dfrA1-positive isolates from the aquaculture site and was not associated with gene cassettes in any. Of nine isolates tested for conjugation, two from the aquaculture site transferred AR determinants to Escherichia coli. High levels of AR in marine sediments from aquaculture and non-aquaculture sites suggest that dispersion of the large amounts of antimicrobials used in Chilean salmon aquaculture has created selective pressure in areas of the marine environment far removed from the initial site of use of these agents.

Host specificity and clade dependent distribution of putative virulence genes in Moritella viscosa.

Moritella viscosa is the aetiological agent of winter-ulcer disease in farmed salmonids in the North Atlantic. Previously, two major (typical and variant) genetic clades have been demonstrated within this bacterial species, one of which is almost solely related to disease in Atlantic salmon (Salmo salar). In the present study infection trials demonstrated that 'typical' M. viscosa isolated from Norwegian Atlantic salmon was highly virulent in this fish species but resulted in lower levels of mortality in rainbow trout. 'Variant' M. viscosa isolated from rainbow trout resulted in modest mortality levels in both Atlantic salmon and rainbow trout. To investigate the possible genetic background for inter-strain virulence differences, 38 M. viscosa isolates of diverse geographical origin and host species and a number of other Moritella spp. were investigated for the presence/absence of putative virulence related homologs. All isolates were positive for DNA sequences coding for; the Type VI secretion ATPase (clpV), hemolysin co-regulated protein (hcp), bacterioferritins (bfrA and bfrB), lectin (hemG), phospholipase D (pld), multifunctional autoprocessing repeats-in-toxin (martxA), aerolysin (aer), invasin (inv), and cytotoxic necrotizing factor (cnf), with the exception of one isolate in which cnf could not be confirmed. The product of an ABC transporter metal-binding lipoprotein (mat) was consistently detected although 11 isolates, all phylogenetically related, appear to produce a truncated version. A putative insecticidal toxin complex (mitABC) was detected almost exclusively in 'typical' Atlantic salmon isolates, and our data indicate that this complex of genes is expressed and co-transcribed. Transmission electron microscopy investigation revealed pili and flagella surface structures on nine M. viscosa representing both typical and variant isolates. Our results provide strong support for the existence of host specificity/high virulence in 'typical' M. viscosa related to Atlantic salmon. The gene distribution also provides further support for the genetic division within M. viscosa, and constitutes a basis for further study of the importance of the mitABC complex in winter-ulcer pathogenesis.

Integrated multi-omics approach reveals novel associations in the rapeseed diet-microbiota-host axis in pigs.

Diet-mediated host-microbiota interplay is a key factor in optimizing the gut function and overall health of the host. Gaining insight into the biological mechanisms behind this relationship is fundamental to finding sustainable, environment-friendly feed solutions in livestock production systems. Here, we apply a multi-omics integration approach to elucidate sustainable diet-associated host-gut microbiota interactions in pigs and we demonstrate novel and biologically relevant host-microbe associations in the gut, driven by a rapeseed meal-based feed (RSF). Interestingly, RSF-diet promoted the abundance of segmented filamentous bacteria Candidatus Arthromitus that was associated with the maintenance of mucosal immunity in the ileum of pigs. In the colon, RSF diet affected host mRNA splicing functions, which may result in different host gene products, through host-microbiota associations, particularly with the Faecalibacterium population, and through the interaction of dietary components such as sinapic acid with the host cells. Moreover, telomere maintenance and organization functions that may determine the overall health of the host were upregulated and notably associated with Subdoligranulum population in the colon of RSF diet-fed pigs. This integrative multi-omics approach provides more insight into the diet-microbiota-host axis, and a better understanding of mechanisms and opportunities to find new strategies for modulating host health and potentially improving caloric and nutritional efficiency in animal production.

Bacteria in Normal Canine Milk Analyzed by Blood Agar Medium.

Studies of microbiota in normal canine milk from healthy dams are sparse. As is the case with blood and urine, it was considered that milk contains no microbiota. Any discovery of bacteria in canine milk is, therefore, often noted to be a result of contamination during sampling or interpreted as mastitis and treated with antibiotics. Milk was collected twice within 19 days after natural parturition from 11 lactating dams, with no general or local clinical signs of mastitis or other disease. The skin and teats were prepared with an antimicrobial protocol prior to each milk sampling. In total, 210 milk samples were collected and assessed for a number of bacterial colonies grown on each plate. Bacterial growth was detected in 180 samples (86%). Staphylococcus pseudintermedius, Enterococcus spp., Clostridium spp., Coagulase-Negative Staphylococci (CoNS), Streptococcus spp., Streptococcus canis, Bacillus spp., Pasteurella spp., and Escherichia coli were identified from pure and/or mixed bacterial growth, listed in descending order of occurrence. Despite the small sample size, the consistent occurrence of bacteria in early postpartum dams indicates a genuine occurrence of bacteria in canine milk, rather than random contamination. The finding of bacteria in the milk of dams should not, therefore, be the sole argument for the diagnosis of mastitis.

Ductal Architecture: Corrosion Casting of Canine Mammary Glands Using an Intraductal Approach.

Detailed knowledge related to the morphology, anatomy, physiology, and pathology of the canine mammary gland is scarce. Mammary tissue undergoes massive changes instructed by hormones multiple times within the lifespan of every bitch, affecting its appearance. To address the ductal system's appearance and to present how different our findings regarding the canine mammary gland are in comparison with the available literature, we obtained cadaveric specimens after euthanasia and mastectomy. All bitches were euthanised due to poor prognosis for their recovery from maladies unrelated to mammae. Using intraductal cannulation ex vivo, milk- or fluid-yielding ducts were perfused using VasQtec (polyurethane resin), which revealed casts, i.e., imprints of ducts and glandular structures in situ. We observed primary, vertically positioned ducts that ascended throughout the teat and continued branching to secondary, tertiary, etc., horizontally positioned ducts, which drained mammary gland lobes under the skin located close to the abdominal wall. The ascendant teat canal could be split into two and could be connected to gland alveoli or end blind. Alveolar formations were located along ducts and ductules in bigger and/or smaller clusters. This study is the first to generate a 3D image of canine ducts and glandular tissue using an intraductal approach.

Cohabitation of Piscirickettsia salmonis genogroups (LF-89 and EM-90): synergistic effect on growth dynamics.

Piscirickettsia salmonis, the biological agent of Salmonid Rickettsial Septicemia (SRS), is a facultative intracellular bacterium that can be divided into two genogroups (LF-89 and EM-90) with different virulence levels and patterns. Studies have found co-infection of these genogroups in salmonid farms in Chile, but it is essential to assess whether this interaction within the host is related to virulence and changes in pathogen dynamics. In this study, we studied four isolates from EM-90 and one LF-89 isolate chosen based on their genomic differences. The aim was to evaluate how co-cultivation affects bacterial growth performance and virulence factor expression in Atlantic salmon (Salmo salar) in vitro and in vivo. In vitro results using FN2 medium, showed a similar growth curve between co-cultures of LF-89 and EM-90 compared to EM-90 monocultures. This was explained by the higher ratio of EM-90 to LF-89 in all co-cultures. When evaluating the expression of virulence factors, it was discovered that the luxR gene was expressed only in EM-90-like isolates and that there were significant differences between mono- and co-cultures for flaA and cheA, suggesting a response to cohabitation. Moreover, during in vivo co-cultures, transcriptomic analysis revealed an upregulation of transposases, flagellum-related genes (fliI and flgK), transporters, and permeases that could unveil novel virulence effectors used in the early infection process of P. salmonis. Thus, our work has shown that cohabitation of P. salmonis genogroups can modulate their behavior and virulence effector expression. These data can contribute to new strategies and approaches to improve the current health treatments against this salmonid pathogen.

The mobile gene cassette carrying tetracycline resistance genes in Aeromonas veronii strain Ah5S-24 isolated from catfish pond sediments shows similarity with a cassette found in other environmental and foodborne bacteria.

Aeromonas veronii is a Gram-negative bacterium ubiquitously found in aquatic environments. It is a foodborne pathogen that causes diarrhea in humans and hemorrhagic septicemia in fish. In the present study, we used whole-genome sequencing (WGS) to evaluate the presence of antimicrobial resistance (AMR) and virulence genes found in A. veronii Ah5S-24 isolated from catfish pond sediments in South-East, United States. We found cphA4, dfrA3, mcr-7.1, valF, bla FOX-7, and bla OXA-12 resistance genes encoded in the chromosome of A. veronii Ah5S-24. We also found the tetracycline tet(E) and tetR genes placed next to the IS5/IS1182 transposase, integrase, and hypothetical proteins that formed as a genetic structure or transposon designated as IS5/IS1182/hp/tet(E)/tetR/hp. BLAST analysis showed that a similar mobile gene cassette (MGC) existed in chromosomes of other bacteria species such as Vibrio parahaemolyticus isolated from retail fish at markets, Aeromonas caviae from human stool and Aeromonas media from a sewage bioreactor. In addition, the IS5/IS1182/hp/tet(E)/tetR/hp cassette was also found in the plasmid of Vibrio alginolyticus isolated from shrimp. As for virulence genes, we found the tap type IV pili (tapA and tapY), polar flagellae (flgA and flgN), lateral flagellae (ifgA and IfgL), and fimbriae (pefC and pefD) genes responsible for motility and adherence. We also found the hemolysin genes (hylII, hylA, and TSH), aerA toxin, biofilm formation, and quorum sensing (LuxS, mshA, and mshQ) genes. However, there were no MGCs encoding virulence genes found in A. veronii AhS5-24. Thus, our findings show that MGCs could play a vital role in the spread of AMR genes between chromosomes and plasmids among bacteria in aquatic environments. Overall, our findings are suggesting that MGCs encoding AMR genes could play a vital role in the spread of resistance acquired from high usage of antimicrobials in aquaculture to animals and humans.