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Aeromonas salmonicida is studied using Dictyostelium discoideum as a model host, with predation resistance measured as a key parameter. Aeromonas salmonicida mesophilic isolates exhibit inconclusive results with the amoebic model. This study focuses on new mesophilic isolates (S24-S38, S26-S10, and S28-S20) from Alberta, Canada, and introduces an improved predation test method. Phylogenetic analysis reveals two subgroups, with S24-S38 and S26-S10 clustering with the subspecies pectinolytica from Argentina, and S28-S20 with strains from India (Y567) and Spain (AJ83), showcasing surprising mesophilic strain diversity across geographic locations. Predation tests were carried out with various mesophilic and psychrophilic strains of A. salmonicida, including Alberta isolates. The amoeba cell lines used were DH1-10 and AX2. Although the mesophilic isolates were very resistant to predation by the amoeba DH1-10, some lost this resistance to the AX2 strain, which appeared more voracious in the conditions tested. In addition, when diluting the culture medium used in a predation test with AX2, a loss of the capacity to predation resistance was observed for all the mesophilic isolates, including the highly resistant S28-S20 isolate. This study provides insights into the predation resistance of A. salmonicida isolates and offers avenues for better characterizing mesophilic isolates., (© The Author(s) 2024. Published by Oxford University Press on behalf of FEMS.)

All the 36 known species to date of the genus Aeromonas are mesophilic except the species Aeromonas salmonicida , which includes both psychrophilic and mesophilic subspecies. For 20 years, more and more mesophilic A. salmonicida strains have been discovered. Only A. salmonicida subsp. pectinolytica has officially been classified as a mesophilic subspecies. Most mesophiles have been isolated in hot countries. We present, for the first time, the characterization of two new mesophilic isolates from Quebec (Canada). Phenotypic and genomic characterizations were carried out on these strains, isolated from dead fish from a fish farm. Isolates 19-K304 and 19-K308 are clearly mesophiles, virulent to the amoeba Dictyostelium discoideum , a surrogate host, and close to strain Y577, isolated in India. To our knowledge, this is the first time that mesophilic strains isolated from different countries are so similar. The major difference between the isolates is the presence of plasmid pY47-3, a cryptic plasmid that sometimes presents in mesophilic strains. More importantly, our extensive phylogenetic analysis reveals two well-defined clades of mesophilic strains with psychrophiles associated with one of these clades. This helps to have a better understanding of the evolution of this species and the apparition of psychrophilic subspecies.

Aeromonas salmonicida subsp. salmonicida causes furunculosis, a major infection that affects fish farms worldwide. We isolated phage vB_AsaM_LPM4 (LPM4) from a diseased fish. Based on its DNA sequence, LPM4 is identical to the uncharacterized Prophage 3, a prophage present mostly in North American A. salmonicida subsp. salmonicida isolates that bear the genomic island AsaGEI2a. Prophage 3 and AsaGEI2a are inserted side by side in the bacterial chromosome. The LPM4/Prophage 3 sequence is similar to that of other prophages found in various members of the genus Aeromonas. LPM4 specifically infects A. salmonicida subsp. salmonicida strains that do not already bear Prophage 3. The presence of an A-layer on the surface of the bacteria is not necessary for the adsorption of phage LPM4 but seems to facilitate its infection process. We also successfully produced lysogenic strains that bear Prophage 3 using sensitive strains with different genetic backgrounds, suggesting that there is no interdependency between LPM4 and AsaGEIs. PCR analysis of the excision dynamics of Prophage 3 and AsaGEIs revealed that these genetic elements can spontaneously excise themselves from the bacterial chromosome independently of one another. Through the isolation and characterization of LPM4, this study reveals new facets of Prophage 3 and AsaGEIs., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

High-throughput genomic sequencing has helped to reveal the plasmidome of Aeromonas salmonicida. This literature review provides an overview of A. salmonicida's rich plasmidome by presenting all the plasmids identified so far, addressing their biological importance and the functional links between them. The plasmids of A. salmonicida, especially those bearing antibiotic resistance genes, can provide clues about interactions of this species with other pathogens (animals and humans), as is the case for pRAS3-3432 and Chlamydia suis or pSN254b and Salmonella enterica. In addition to antibiotic resistance, plasmids play an important role in the virulence of A. salmonicida, particularly for the subspecies salmonicida and the plasmid pAsa5, which carries genes for the type-three secretion system, a virulence factor essential for the bacterium. The A. salmonicida plasmidome also has many cryptic plasmids with no known biological function, but which can be used for the acquisition of new genetic elements. Striking examples are pAsa7 and pAsaXII that provide, respectively, resistance to chloramphenicol and formaldehyde and are derivatives of cryptic pAsa2., (© 2020 New York Academy of Sciences.)

Aeromonas salmonicida strains cause problematic bacterial infections in the aquaculture industry worldwide. The genus Aeromonas includes both mesophilic and psychrophilic species. Bacteriophages that infect Aeromonas spp. strains are usually specific for mesophilic or psychrophilic species; only a few bacteriophages can infect both types of strains. In this study, we characterized the podophage T7-Ah, which was initially found to infect the Aeromonas salmonicida HER1209 strain. The burst size of T7-Ah against its original host is 72 new virions per infected cell, and its burst time is 30 minutes. It has been found that this phage can lyse both mesophilic and psychrophilic A. salmonicida strains, as well as one strain of Escherichia coli. Its genome comprises 40,153 bp of DNA and does not contain any recognizable toxin or antibiotic resistance genes. The adsorption rate of the phage on highly sensitive bacterial strains was variable and could not be related to the presence or absence of a functional A-layer on the surface of the bacterial strains. The lipopolysaccharide migration patterns of both resistant and sensitive bacterial strains were also studied and compared to investigate the nature of the potential receptor of this phage on the bacterial surface. This study sheds light on the surprising diversity of lifestyles of the bacterial strains sensitive to phage T7-Ah and opens the door to the potential use of this phage against A. salmonicida infections in aquaculture.

The Gram-negative bacterium Aeromonas salmonicida subsp. salmonicida is an aquatic pathogen which causes furunculosis to salmonids, especially in fish farms. The emergence of strains of this bacterium exhibiting antibiotic resistance is increasing, limiting the effectiveness of antibiotherapy as a treatment against this worldwide disease. In the present study, we discovered an isolate of A. salmonicida subsp. salmonicida that harbors two novel plasmids variants carrying antibiotic resistance genes. The use of long-read sequencing (PacBio) allowed us to fully characterize those variants, named pAsa5-3432 and pRAS3-3432, which both differ from their classic counterpart through their content in mobile genetic elements. The plasmid pAsa5-3432 carries a new multidrug region composed of multiple mobile genetic elements, including a Class 1 integron similar to an integrated element of Salmonella enterica. With this new region, probably acquired through plasmid recombination, pAsa5-3432 is the first reported plasmid of this bacterium that bears both an essential virulence factor (the type three secretion system) and multiple antibiotic resistance genes. As for pRAS3-3432, compared to the classic pRAS3, it carries a new mobile element that has only been identified in Chlamydia suis. Hence, with the identification of those two novel plasmids harboring mobile genetic elements that are normally encountered in other bacterial species, the present study puts emphasis on the important impact of mobile genetic elements in the genomic plasticity of A. salmonicida subsp. salmonicida and suggests that this aquatic bacterium could be an important reservoir of antibiotic resistance genes that can be exchanged with other bacteria, including human and animal pathogens., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Aeromonas salmonicida subsp. salmonicida is a fish pathogen that causes furunculosis. Antibiotherapy used to treat furunculosis in fish has led to resistance. Virulent phages are increasingly seen as alternatives or complementary treatments against furunculosis in aquaculture environments. For phage therapy to be successful, it is essential to study the natural mechanisms of phage resistance in A. salmonicida subsp. salmonicida. Here, we generated bacteriophage-insensitive mutants (BIMs) of A. salmonicida subsp. salmonicida, using a myophage with broad host range and characterized them. Phage plaques were different depending on whether the A-layer surface array protein was expressed or not. The genome analysis of the BIMs helped to identify mutations in genes involved in the biogenesis of lipopolysaccharides (LPS) and on an uncharacterized gene (ASA_1998). The characterization of the LPS profile and gene complementation assays identified LPS as a phage receptor and confirmed the involvement of the uncharacterized protein ASA_1998 in phage infection. In addition, we confirmed that the presence of an A-layer at the bacterial surface could act as protection against phages. This study brings new elements into our understanding of the phage adsorption to A. salmonicida subsp. salmonicida cells., (© 2019 John Wiley & Sons Ltd.)

The bacterium Aeromonas salmonicida is known since long time as a major fish pathogen unable to grow at 37 °C. However, some cases of human infection by putative mesophilic A. salmonicida have been reported. The goal of the present study is to examine two clinical cases of human infection by A. salmonicida in Spain and to investigate the pathogenicity in mammals of selected mesophilic A. salmonicida strains. An evaluation of the pathogenicity in a mouse model of clinical and environmental A. salmonicida strains was performed. The genomes of the strains were sequenced and analyzed in order to find the virulence determinants of these strains. The experimental infection in mice showed a gradient in the virulence of these strains and that some of them can cause necrotizing fasciitis and tissue damage in the liver. In addition to demonstrating significant genomic diversity among the strains studied, bioinformatics analyses permitted also to shed light on crucial elements for the virulence of the strains, like the presence of a type III secretion system in the one that caused the highest mortality in the experimental infection. Clinicians and microbiologists should consider these results for the inclusion of A. salmonicida in diagnosis tests since it is now clear that some mesophilic strains are also pathogens for humans., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The Gram-negative bacterium Aeromonas salmonicida contains five subspecies: salmonicida, smithia, achromogenes, masoucida and pectinolytica. Pectinolytica is a mesophilic subspecies with the ability to thrive at a wide range of temperatures, including 37°C, while the four other subspecies are psychrophilic, restricted to lower temperatures. The psychrophilic subspecies are known to infect a wide range of fishes. However, there is no evidence of pathogenicity for the mesophilic subspecies pectinolytica. Study of the differences between the mesophilic and psychrophilic subspecies is hampered by the lack of completely sequenced and closed genomes from the mesophilic subspecies. A previous study reported that insertion sequences, which can induce genomic rearrangements at temperatures around 25°C, could be one of the determinants explaining the differences in lifestyle (mesophilic or psychrophilic) between the subspecies. In this study, the genome of mesophilic strain A527 of A. salmonicida was sequenced, closed and analyzed to investigate the mesophilic-psychrophilic discrepancy. This reference genome supports the hypothesis that insertion sequences are major determinants of the lifestyle differences between the A. salmonicida subspecies. Moreover, the phylogenetic analysis performed to position strain A527 within the taxonomy raises an issue regarding the intraspecies structure of A. salmonicida., (© FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Phages infecting Aeromonas salmonicida subsp. salmonicida, the causative agent of the fish disease furunculosis, have been isolated for decades but very few of them have been characterized. Here, the host range of 12 virulent phages, including three isolated in the present study, was evaluated against a panel of 65 A. salmonicida isolates, including representatives of the psychrophilic subspecies salmonicida, smithia, masoucida, and the mesophilic subspecies pectinolytica. This bacterial set also included three isolates from India suspected of being members of a new subspecies. Our results allowed to elucidate a lytic dichotomy based on the lifestyle of A. salmonicida (mesophilic or psychrophilic) and more generally, on phage types (lysotypes) for the subspecies salmonicida. The genomic analyses of the 12 phages from this study with those available in GenBank led us to propose an A. salmonicida phage pan-virome. Our comparative genomic analyses also suggest that some phage genes were under positive selection and A. salmonicida phage genomes having a discrepancy in GC% compared to the host genome encode tRNA genes to likely overpass the bias in codon usage. Finally, we propose a new classification scheme for A. salmonicida phages.

Background: Aeromonas salmonicida subsp. salmonicida is a ubiquitous psychrophilic waterborne bacterium and a fish pathogen. The numerous mobile elements, especially insertion sequences (IS), in its genome promote rearrangements that impact its phenotype. One of the main virulence factors of this bacterium, its type three secretion system (TTSS), is affected by these rearrangements. In Aeromonas salmonicida subsp. salmonicida most of the TTSS genes are encoded in a single locus on a large plasmid called pAsa5, and may be lost when the bacterium is cultivated at a higher temperature (25 °C), producing non-virulent mutants. In a previous study, pAsa5-rearranged strains that lacked the TTSS locus on pAsa5 were produced using parental strains, including 01-B526. Some of the generated deletions were explained by homologous recombination between ISs found on pAsa5, whereas the others remained unresolved. To investigate those rearrangements, short- and long-read high-throughput sequencing technologies were used on the A. salmonicida subsp. salmonicida 01-B526 whole genome., Results: Whole genome sequencing of the 01-B526 strain revealed that its pAsa5 has an additional IS copy, an ISAS5, compared to the reference strain (A449) sequence, which allowed for a previously unknown rearrangement to occur. It also appeared that 01-B526 bears a second large plasmid, named pAsa9, which shares 40 kbp of highly similar sequences with pAsa5. Following these discoveries, previously unexplained deletions were elucidated by genotyping. Furthermore, in one of the derived strains a fusion of pAsa5 and pAsa9, involving the newly discovered ISAS5 copy, was observed., Conclusion: The loss of TTSS and hence virulence is explained by one consistent mechanism: IS-driven homologous recombination. The similarities between pAsa9 and pAsa5 also provide another example of genetic diversity driven by ISs.

The bacterium Aeromonas salmonicida subsp. salmonicida is a common pathogen in fish farms worldwide. Since the antibiotic resistance of this bacterial species is on the increase, it is important to have a broader view on this issue. In the present study, we tested the presence of known plasmids conferring multi-drug resistance as well as antibiotic resistance genes by a PCR approach in 100 Canadian A. salmonicida subsp. salmonicida isolates. Our study highlighted the dominance of the conjugative pSN254b plasmid, which confers multi-drug resistance. We also identified a new multi-drug plasmid named pAsa8, which has been characterized by a combination of sequencing technologies (Illumina and Oxford nanopore). This new plasmid harbors a complex class 1 integron similar to the one of the Salmonella genomic island 1 (SGI1) found in Salmonella enterica and Proteus mirabilis. Consequently, in addition to providing an update on the A. salmonicida subsp. salmonicida isolates that are resistant to antibiotics, our data suggest that this bacterium is potentially an important reservoir of drug resistance genes and should consequently be monitored more extensively. In addition, we describe a screening method that has the potential to become a diagnostic tool that is complementary to other methods currently in use.

Aeromonas salmonicida subsp. salmonicida is a fish pathogen known to have a rich plasmidome. In the present study, we discovered an isolate of this bacterium bearing an additional unidentified small plasmid. After having sequenced the DNA of that isolate by next-generation sequencing, it appeared that the new small plasmid is a ColE1-type replicon plasmid, named here pAsa7. This plasmid bears a functional chloramphenicol-acetyltransferase-encoding gene (cat-pAsa7) previously unknown in A. salmonicida and responsible for resistance to chloramphenicol. A comparison of pAsa7 with pAsa2, the only known ColE1-type replicon plasmid usually found in A. salmonicida subsp. salmonicida, revealed that even if both plasmids share a high structural similarity, it is still unclear if pAsa7 is a derivative of pAsa2 since they showed several mutations at the nucleotide level. Transcriptomic analysis revealed that the cat-pAsa4 gene, another chloramphenicol-acetyltransferase-encoding gene, found on the large plasmid pAsa4, was significantly more transcribed than cat-pAsa7. This was correlated with a higher chloramphenicol resistance for isolates bearing pAsa4 compared with the one having pAsa7. Finally, a phylogenetic analysis showed that both CAT-pAsa4 and CAT-pAsa7 proteins were in different clusters. The clustering was supported by the identity of residues involved in the catalytic site. In addition, to give a better understanding of the large drug-resistance panel of A. salmonicida, this study reinforces the hypothesis that A. salmonicida subsp. salmonicida is a considerable reservoir for mobile genetic elements such as plasmids.

Background: Aeromonads make up a group of Gram-negative bacteria that includes human and fish pathogens. The Aeromonas salmonicida species has the peculiarity of including five known subspecies. However, few studies of the genomes of A. salmonicida subspecies have been reported to date., Results: We sequenced the genomes of additional A. salmonicida isolates, including three from India, using next-generation sequencing in order to gain a better understanding of the genomic and phylogenetic links between A. salmonicida subspecies. Their relative phylogenetic positions were confirmed by a core genome phylogeny based on 1645 gene sequences. The Indian isolates, which formed a sub-group together with A. salmonicida subsp. pectinolytica, were able to grow at either at 18 °C and 37 °C, unlike the A. salmonicida psychrophilic isolates that did not grow at 37 °C. Amino acid frequencies, GC content, tRNA composition, loss and gain of genes during evolution, pseudogenes as well as genes under positive selection and the mobilome were studied to explain this intraspecies dichotomy., Conclusion: Insertion sequences appeared to be an important driving force that locked the psychrophilic strains into their particular lifestyle in order to conserve their genomic integrity. This observation, based on comparative genomics, is in agreement with previous results showing that insertion sequence mobility induced by heat in A. salmonicida subspecies causes genomic plasticity, resulting in a deleterious effect on the virulence of the bacterium. We provide a proof-of-concept that selfish DNAs play a major role in the evolution of bacterial species by modeling genomes.

Aeromonas salmonicida subsp. salmonicida is the causal agent of furunculosis in salmonids. We recently identified a group of genomic islands (AsaGEI) in this bacterium. AsaGEI2a, one of these genomic islands, has almost exclusively been identified in isolates from North America. To date, Aeromonas salmonicida subsp. salmonicida JF3224, a strain isolated from a wild brown trout (Salmo trutta) caught in Switzerland, was the only European isolate that appeared to bear AsaGEI2a. We analyzed the genome of JF3224 and showed that the genomic island in JF3224 is a new variant of AsaGEI, which we have called AsaGEI2b. While AsaGEI2b shares the same integrase gene and insertion site as AsaGEI2a, it is very different in terms of many other features. Additional genomic investigations combined with PCR genotyping revealed that JF3224 is sensitive to growth at 25°C, leading to insertion sequence-dependent rearrangement of the locus on the pAsa5 plasmid that encodes a type three secretion system, which is essential for the virulence of the bacterium. The analysis of the JF3224 genome confirmed that AsaGEIs are accurate indicators of the geographic origins of A. salmonicida subsp. salmonicida isolates and is another example of the susceptibility of the pAsa5 plasmid to DNA rearrangements., (© FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The bacterium Aeromonas salmonicida is the etiological agent of furunculosis, a widespread fish disease causing important economic losses to the fish farming industry. Antibiotic treatments in fish farms may be challenging given the existence of multidrug-resistant isolates of this bacterium. Here, we report the draft genome sequences of the 2004-05MF26 and 2009-144K3 isolates, which harbor plasmids conferring antibiotic resistance. Both isolates also carry the large plasmid pAsa5, which is known to encode a type three secretion system (TTSS) and the pAsal1 plasmid which has the aopP gene producing a TTSS effector. These two isolates are good representatives of the plasmid diversity in A. salmonicida subsp. salmonicida., (© FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Aeromonas salmonicida subsp. salmonicida is a fish pathogen. Analysis of its genomic characteristics is required to determine the worldwide distribution of the various populations of this bacterium. Genomic alignments between the 01-B526 pathogenic strain and the A449 reference strain have revealed a 51-kb chromosomal insertion in 01-B526. This insertion (AsaGEI1a) has been identified as a new genomic island (GEI) bearing prophage genes. PCR assays were used to detect this GEI in a collection of 139 A. salmonicida subsp. salmonicida isolates. Three forms of this GEI (AsaGEI1a, AsaGEI1b, AsaGEI2a) are now known based on this analysis and the sequencing of the genomes of seven additional isolates. A new prophage (prophage 3) associated with AsaGEI2a was also discovered. Each GEI appeared to be strongly associated with a specific geographic region. AsaGEI1a and AsaGEI2a were exclusively found in North American isolates, except for one European isolate bearing AsaGEI2a. The majority of the isolates bearing AsaGEI1b or no GEI were from Europe. Prophage 3 has also a particular geographic distribution and was found only in North American isolates. We demonstrated that A. salmonicida subsp. salmonicida possesses unsuspected elements of genomic heterogeneity that could be used as indicators to determine the geographic origins of isolates of this bacterium., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Plasmids that carry antibiotic resistance genes occur frequently in Aeromonas salmonicida subsp. salmonicida, an aquatic pathogen with severe consequences in salmonid farming. Here, we describe a 67 kb plasmid found in the A. salmonicida subsp. salmonicida Strain SHY15-2939 from Quebec, Canada. This new plasmid, named pAsa-2939 and identified by high throughput sequencing, displays features never found before in this bacterial species. It contains a transposon related to the Tn21 family, but with an unusual organization. This transposon bears a catB3 gene (chloramphenicol resistance) that has not been detected yet in A. salmonicida subsp. salmonicida. The plasmid is transferable by conjugation into Aeromonas hydrophila, but not into Escherichia coli. Based on PCR analysis and genomic sequencing (Illumina and PacBio), we determined that the transposon is unstable in A. salmonicida subsp. salmonicida Strain SHY15-2939, but it is stable in A. hydrophila trans-conjugants, which explains the chloramphenicol resistance variability observed in SHY15-2939. These results suggest that this bacterium is likely not the most appropriate host for this plasmid. The presence of pAsa-2939 in A. salmonicida subsp. salmonicida also strengthens the reservoir role of this bacterium for antibiotic resistance genes, even those that resist antibiotics not used in aquaculture in Québec, such as chloramphenicol. [ABSTRACT FROM AUTHOR]

Aeromonas salmonicida subsp. salmonicida is a pathogenic bacterium responsible for furunculosis in salmonids. Following an outbreak of furunculosis, the infection can be treated with antibiotics, but it is common to observe ineffective treatment due to antibiotic resistance. This bacterium has a wide variety of plasmids responsible for this resistance. Among them, pRAS3 carries a tetracycline resistance gene. Several variants of this plasmid have been discovered over the years (pRAS3-3432 and pRAS3.1 to 3.4). During the present study, two new variants of the plasmid pRAS3 were identified (pRAS3.5 and pRAS3-3759) in strains of A. salmonicida subsp. salmonicida. Plasmid pRAS3-3759, which has been found in many strains from the same region over the past three years, has an additional genetic element identical to one found in pRAS3-3432. This genetic element was also found in Chlamydia suis, a swine pathogen. In this study, we analyzed the bacteria's resistance to tetracycline, the number of copies of the plasmids, and the growth of the strains that carry five of the pRAS3 variants (pRAS3.3 to 3.5, pRAS3-3432, and pRAS3-3759). The results show no particular trend despite the differences between the plasmids, except for the resistance to tetracycline when analyzed in an isogenic background. Blast analysis also revealed the presence of pRAS3 plasmids in other bacterial species, which suggests that this plasmid family has widely spread. This study once again highlights the ability of A. salmonicida subsp. salmonicida to adapt to furunculosis antibiotic treatments, and the still-growing family of pRAS3 plasmids. [ABSTRACT FROM AUTHOR]

Genomic islands (Aeromonas salmonicida genomic islands , AsaGEI s) are found worldwide in many isolates of Aeromonas salmonicida subsp. salmonicida , a fish pathogen. To date, five variants of AsaGEI (1a, 1b, 2a, 2b and 2c) have been described. Here, we investigate a sixth AsaGEI , which was identified in France between 2016 and 2019 in 20 A. salmonicida subsp. salmonicida isolates recovered from sick salmon all at the same location. This new AsaGEI shares the same insertion site in the chromosome as the other AsaGEI2 s as they all have a homologous integrase gene. This new AsaGEI was thus named AsaGEI2d , and has five unique genes compared to the other AsaGEI s. The isolates carrying AsaGEI2d also bear the plasmid pAsa7, which was initially found in an isolate from Switzerland. This plasmid provides resistance to chloramphenicol thanks to a cat gene. This study reveals more about the diversity of the AsaGEI s. [ABSTRACT FROM AUTHOR]

In Aeromonas salmonicida subsp. salmonicida, a bacterium that causes fish disease, there are two types of small plasmids (<15 kbp): plasmids without known function, called cryptic plasmids, and plasmids that bear beneficial genes for the bacterium. Four among them are frequently detected in strains of A. salmonicida subsp. salmonicida: pAsa1, pAsa2, pAsa3, and pAsal1. The latter harbors a gene which codes for an effector of the type three secretion system, while the three others are cryptic. It is currently unclear why these cryptic plasmids are so highly conserved throughout strains of A. salmonicida subsp. salmonicida. In this study, three small plasmids, named pAsa10, pAsaXI and pAsaXII, are described. Linked to tetracycline resistance, a partial Tn1721 occupies half of pAsa10. A whole Tn1721 is also present in pAsa8, another plasmid previously described in A. salmonicida subsp. salmonicida. The backbone of pAsa10 has no relation with other plasmids described in this bacterium. However, the pAsaXI and pAsaXII plasmids are derivatives of cryptic plasmids pAsa3 and pAsa2, respectively. pAsaXI is identical to pAsa3, but bears a transposon with a gene that encodes for a putative virulence factor. pAsaXII, also found in Aeromonas bivalvium, has a 95% nucleotide identity with the backbone of pAsa2. Like pAsa7, another pAsa2-like plasmid recently described, orf2 and orf3 are missing and are replaced in pAsaXII by genes that encode a formaldehyde detoxification system. These new observations suggest that transposons and particularly Tn1721 are frequent and diversified in A. salmonicida subsp. salmonicida. Moreover, the discovery of pAsaXI and pAsaXII expands the group of small plasmids that are derived from cryptic plasmids and have a function. Although their precise roles remain to be determined, the present study shows that cryptic plasmids could serve as moldable vectors to acquire mobile elements such as transposons. Consequently, they could act as key agents of the diversification of virulence and adaptive traits of Aeromonas salmonicida subsp. salmonicida. [ABSTRACT FROM AUTHOR]

Furunculosis, which is caused by Aeromonas salmonicida subsp. salmonicida, is a major salmonid disease in fish farms worldwide. Several plasmids found in this bacterium confer phenotypes such drug resistance and virulence. Small plasmids (pAsa1, pAsa2, pAsa3, and pAsal1) related to ColE1- and ColE2-type replicons are usually present in its normal plasmidome. In the present study, with the objective to investigate if these plasmids display particularities related to the origin of the isolates bearing them, a total of 153 isolates, including 78 new and 75 previously described, were analyzed for the presence of small plasmids by PCR and DNA restriction fragment profiling. A geographical dichotomy between Canadian and European isolates for their propensity to do not have pAsa3 or pAsal1 was found. In addition, the genotyping analysis led to the identification of two European isolates harboring an unusual pAsal1. An investigation by next-generation sequencing (NGS) of these two isolates shed light on two pAsal1 variants (pAsal1C and pAsal1D). As with pAsal1B, another pAsal1 variant previously described, these two new variants bore a second insertion sequence (ISAS5) in addition to the usual ISAS11. The characterization of these variants suggested that they could predominate over the wild-type pAsal1 in stressful conditions such as growth at temperatures of 25?C and above. To obtain a comprehensive portrait of the mutational pressure on small plasmids, 26 isolates whose DNA had been sequenced by NGS were investigated. pAsa3 and pAsal1 were more prone to mutations than pAsa1 and pAsa2, especially in the mobA gene, which encodes a relaxase and a primase. Lastly, the average copy number of each plasmid per cell was assessed using raw sequencing data. A clear trend with respect to the relative proportion per cell of each plasmid was identified. Our large-scale study revealed a geographical dichotomy in small plasmid repertoire in addition to a clear trend for pAsa3 and pAsal1 to be more frequently altered. Moreover, we present the discovery of two new variants of pAsal1: pAsal1C and pAsal1D. [ABSTRACT FROM AUTHOR]

The bacterium Aeromonas salmonicida has long been known to be one of the most feared pathogens in fish farming. However, the more we discover about this bacterial species, the more we question whether it is really exclusively an aquatic pathogen. In recent years, it has become obvious that this bacterial species includes a myriad of strains with various lifestyle and ecological niches, including the well-known strict psychrophiles, the first bacteria known of the species, and the newly described mesophilic strains. The mesophiles are able to grow at low temperatures, but even better at temperatures of approximately 37 °C, which strict psychrophiles cannot do. In this perspective article, we address some aspects surrounding this dual lifestyle in A. salmonicida, including the impact of mobile genetic elements, and how future research around this bacterial species may focus on the psychrophilic/mesophilic dichotomy, which makes A. salmonicida an increasingly interesting and relevant model for the study of speciation. [ABSTRACT FROM AUTHOR]

The type three secretion system (TTSS) locus of Aeromonas salmonicida subsp. salmonicida, located on the plasmid pAsa5, is known to be lost when the bacterium is grown at temperatures of 25 °C. The loss of the locus is due to the recombination of the insertion sequences flanking the TTSS region. However, the mechanism involved in this recombination is still elusive. Here, we analyzed 22 A. salmonicida subsp. salmonicida strains that had already lost their TTSS locus, and we systematically explored another 47 strains for their susceptibility to lose the same locus when grown at 25 °C. It appeared that strains from Europe were more prone to lose their TTSS locus compared to Canadian strains. More specifically, it was not possible to induce TTSS loss in Canadian strains that have AsaGEI2a, a genomic island, and prophage 3, or in Canadian strains without a genomic island. A comparative genomic approach revealed an almost perfect correlation between the presence of a cluster of genes, not yet characterized, and the susceptibility of various groups of strains to lose their locus. This cluster of genes encodes putative proteins with DNA binding capacity and phage proteins. This discovery creates new opportunities in the study of pAsa5 thermosensitivity. [ABSTRACT FROM AUTHOR]

Aeromonas salmonicida is a Gram-negative bacterium, known as a fish pathogen since its discovery. Although the species was initially considered psychrophilic, a mesophilic subspecies (pectinolytica) and many other mesophilic strains still not attributed to subspecies have been described in the last two decades. These mesophilic strains were sampled from various sources, including humans, and some of them are known to be pathogenic. In this study, we describe a strain, JF2480, which was isolated from the spleen, and also found the kidney and liver of a dead pied avocet (Recurvirostra avosetta), a type of migratory bird inhabiting aquatic environments. A core genome phylogenomic analysis suggests that JF2480 is taxonomically distant from other known A.salmonicida subspecies. The genome sequence confirms that the strain possesses key virulence genes that are present in the typical A. salmonicida psychrophilic subspecies, with the exception of the genes encoding the type three secretion system (T3SS). Bacterial virulence assays conducted on the surrogate host Dictyostelium discoideum amoeba confirmed that the strain is virulent despite the lack of T3SS. Bacterial growth curves showed that strain JF2480 grow well at 40 °C, the body temperature of the pied avocet, and even faster at 41 °C, compared to other mesophilic strains. Discovery of this strain further demonstrates the extent of the phylogenomic tree of this species. This study also suggests that A. salmonicida can infect a wider array of hosts than previously suspected and that we need to rethink the way we perceive A. salmonicida's natural environment. [ABSTRACT FROM AUTHOR]