Abstract Heritable hypermutable strains deficient in DNA repair genes (mutators) facilitate microbial adaptation as they may rapidly generate beneficial mutations. Mutators deficient in mismatch (MMR) and oxidised guanine (OG) repair are abundant in clinical samples and show increased adaptive potential in experimental infection models but their role in pathoadaptation is poorly understood. Here we investigate the role of mutators in epidemiology and evolution of the broad host pathogen, Streptococcus iniae, employing 80 strains isolated globally over 40 years. We determine phylogenetic relationship among S. iniae using 10,267 non-recombinant core genome single nucleotide polymorphisms (SNPs), estimate their mutation rate by fluctuation analysis, and detect variation in major MMR (mutS, mutL, dnaN, recD2, rnhC) and OG (mutY, mutM, mutX) genes. S. iniae mutation rate phenotype and genotype are strongly associated with phylogenetic diversification and variation in major streptococcal virulence determinants (capsular polysaccharide, hemolysin, cell chain length, resistance to oxidation, and biofilm formation). Furthermore, profound changes in virulence determinants observed in mammalian isolates (atypical host) and vaccine-escape isolates found in bone (atypical tissue) of vaccinated barramundi are linked to multiple MMR and OG variants and unique mutation rates. This implies that adaptation to new host taxa, new host tissue, and to immunity of a vaccinated host is promoted by mutator strains. Our findings support the importance of mutation rate dynamics in evolution of pathogenic bacteria, in particular adaptation to a drastically different immunological setting that occurs during host jump and vaccine escape events. Importance Host immune response is a powerful selective pressure that drives diversification of pathogenic microorganisms and, ultimately, evolution of new strains. Major adaptive events in pathogen evolution, such as transmission to a new host species or infection of vaccinated hosts, require adaptation to a drastically different immune landscape. Such adaptation may be favoured by hypermutable strains (or mutators) that are defective in normal DNA repair and consequently capable of generating multiple potentially beneficial and compensatory mutations. This permits rapid adjustment of virulence and antigenicity in a new immunological setting. Here we show that mutators, through mutations in DNA repair genes and corresponding shifts in mutation rate, are associated with major diversification events and virulence evolution in the broad host-range pathogen Streptococcus iniae. We show that mutators underpin infection of vaccinated hosts, transmission to new host species and the evolution of new strains.
Hypermutable strains (mutators) occur via mutations in DNA repair genes and facilitate microbial adaptation as they may rapidly generate stress-resistance mutations. Bacterial mutators deficient in mismatch repair (MMR) and oxidised guanine repair (OG) are abundant in clinical settings, however their role in epidemiology and evolution of virulence remains poorly understood. Here we determine phylogenetic relationship among 80 strains of Streptococcus iniae isolated globally over 40 years based using non-recombinant core genome single nucleotide polymorphisms (SNPs), estimate their mutation rate by fluctuation analysis, and identify variation in major MMR (mutS, mutL, dnaN, recD2, rnhC) and OG (yfhQ, mutM, mutT) genes. We find that S. iniae mutation rate phenotype and genotype are strongly associated with phylogenetic diversification and variation in major streptococcal virulence determinants (capsular polysaccharide, hemolysin, cell chain length, resistance to oxidation, and biofilm formation). Furthermore, we find that profound changes in virulence determinants observed in mammalian isolates (atypical host) and vaccine-escape isolates found in bone (atypical tissue) of vaccinated barramundi are linked to multiple MMR and OG variants and unique mutation rates. This implies that adaptation to new host taxa, to new host tissue, and to immunity of a vaccinated host may be facilitated by mutator strains. Our findings highlight the importance of mutators and mutation rate dynamics in evolution of pathogenic bacteria, in particular adaptation to a drastically different immunological setting that occurs during host jump and vaccine escape events.
Abstract Pathogens continuously adapt to changing host environments where variation in their virulence and antigenicity is critical to their long-term evolutionary success. The emergence of novel variants is accelerated in microbial mutator strains (mutators) deficient in DNA repair genes, most often from mismatch repair and oxidised-guanine repair systems (MMR and OG respectively). Bacterial MMR/OG mutants are abundant in clinical samples and show increased adaptive potential in experimental infection models, yet the role of mutators in the epidemiology and evolution of infectious disease is not well understood. Here we investigated the role of mutation rate dynamics in the evolution of a broad host range pathogen, Streptococcus iniae , using a set of 80 strains isolated globally over 40 years. We have resolved phylogenetic relationships using non-recombinant core genome variants, measured in vivo mutation rates by fluctuation analysis, identified variation in major MMR/OG genes and their regulatory regions, and phenotyped the major traits determining virulence in streptococci. We found that both mutation rate and MMR/OG genotype are remarkably conserved within phylogenetic clades but significantly differ between major phylogenetic lineages. Further, variation in MMR/OG loci correlates with occurrence of atypical virulence-associated phenotypes, infection in atypical hosts (mammals), and atypical tissue of a vaccinated primary hosts (barramundi bone). These findings suggest that mutators are likely to facilitate adaptations preceding major diversification events, and may promote emergence of variation permitting colonisation of a novel host tissue, novel host taxa (host jumps), and immune-escape in the vaccinated host.
Abstract Fish mortality caused by Streptococcus iniae is a major economic problem in fish aquaculture in warm and temperate regions globally. There is also risk of zoonotic infection by S. iniae through handling of contaminated fish. In this study, we present the complete genome sequence of S. iniae strain QMA0248, isolated from farmed barramundi in South Australia. The 2.12 Mb genome of S. iniae QMA0248 carries a 32 Kb prophage, a 12 Kb genomic island, and 92 discrete insertion sequence (IS) elements. These include 9 novel IS types that belong mostly to the IS 3 family. Comparative and phylogenetic analysis between S. iniae QMA0248 and publicly available complete S. iniae genomes revealed discrepancies that are likely due to misassembly in the genomes of isolates ISET0901 and ISNO. We also determined by long-range PCR that a tandem duplication of an rRNA region in the PacBio assembly of QMA0248 was an assembly error. A similar rRNA duplication in the PacBio genome of S. iniae 89353 may also be a misassembly. Our study not only highlights assembly problems in existing genomes, but provides a high quality reference genome for S. iniae QMA0248, including manually curated mobile genetic elements, that will assist future S. iniae comparative genomic and evolutionary studies.
Photobacterium damselae comprises two subspecies, P. damselae subsp. damselae and P. damselae subsp. piscicida, that contrast remarkably despite their taxonomic relationship. The former is opportunistic and free-living but can cause disease in compromised individuals from a broad diversity of taxa, while the latter is a highly specialized, primary fish pathogen. Here, we employ new closed curated genome assemblies from Australia to estimate the global phylogenetic structure of the species P. damselae. We identify genes responsible for the shift from an opportunist to a host-adapted fish pathogen, potentially via an arthropod vector as fish-to-fish transmission was not achieved in repeated cohabitation challenges despite high virulence for Seriola lalandi. Acquisition of ShdA adhesin and of thiol peroxidase may have allowed the environmental, generalist ancestor to colonize zooplankton and to occasionally enter in fish host sentinel cells. As dependence on the host has increased, P. damselae has lost nonessential genes, such as those related to nitrite and sulfite reduction, urea degradation, a type 6 secretion system (T6SS) and several toxin-antitoxin (TA) systems. Similar to the evolution of Yersinia pestis, the loss of urease may be the crucial event that allowed the pathogen to stably colonize zooplankton vectors. Acquisition of host-specific genes, such as those required to form a sialic acid capsule, was likely necessary for the emergent P. damselae subsp. piscicida to become a highly specialized, facultative intracellular fish pathogen. Processes that have shaped P. damselae subsp. piscicida from subsp. damselae are similar to those underlying evolution of Yersinia pestis from Y. pseudotuberculosis. IMPORTANCE Photobacterium damselae subsp. damselae is a ubiquitous marine bacterium and opportunistic pathogen of compromised hosts of diverse taxa. In contrast, its sister subspecies P. damselae subsp. piscicida (Pdp) is highly virulent in fish. Pdp has evolved from a single subclade of Pdd through gene loss and acquisition. We show that fish-to-fish transmission does not occur in repeated infection models in the primary host, Seriola lalandi, and present genomic evidence for vector-borne transmission, potentially via zooplankton. The broad genomic changes from generalist Pdd to specialist Pdp parallel those of the environmental opportunist Yersinia pseudotuberculosis to vector-borne plague bacterium Y. pestis and demonstrate that evolutionary processes in bacterial pathogens are universal between the terrestrial and marine biosphere.
Heritable hypermutable strains deficient in DNA repair genes (mutators) facilitate microbial adaptation as they may rapidly generate beneficial mutations. Bacterial mutators deficient in mismatch (MMR) and oxidised guanine (OG) repair are abundant in clinical samples and show increased adaptive potential in experimental models of infection in mice. However, their role in pathoadaptation is poorly understood. Here we investigate the role of mutators in epidemiology and evolution of the rapidly evolving aquatic pathogen, Streptococcus iniae, employing a collection of 80 strains isolated globally over 40 years. We determine phylogenetic relationship among S. iniae using 10,267 non-recombinant core genome single nucleotide polymorphisms (SNPs), estimate their mutation rate by fluctuation analysis, and detect variation in major MMR (mutS, mutL, dnaN, recD2, rnhC) and OG (mutY, mutM, mutT) genes. We find that S. iniae mutation rate phenotype and genotype are strongly associated with phylogenetic diversification and variation in major streptococcal virulence determinants (capsular polysaccharide, hemolysin, cell chain length, resistance to oxidation, and biofilm formation). Furthermore, profound changes in virulence determinants observed in mammalian isolates (atypical host) and vaccine-escape isolates found in bone (atypical tissue) of vaccinated barramundi are linked to multiple MMR and OG variants and unique mutation rates. This implies that adaptation to new host taxa, new host tissue, and to immunity of a vaccinated host is promoted by mutator strains. Our findings support the importance of mutation rate dynamics in evolution of pathogenic bacteria, in particular adaptation to a drastically different immunological setting that occurs during host jump and vaccine escape events.
Fish mortality caused by Streptococcus iniae is a major economic problem in fish aquaculture in warm and temperate regions globally. There is also risk of zoonotic infection by S. iniae through handling of contaminated fish. In this study, we present the complete genome sequence of S. iniae strain QMA0248, isolated from farmed barramundi inSouth Australia. The 2.12 Mb genome of S. iniae QMA0248 carries a 32 Kb prophage, a 12 Kb genomic island, and 92 discrete insertion sequence (IS) elements. These include 9 novel IS types that belong mostly to the IS3 family. Comparative and phylogenetic analysis between S. iniae QMA0248 and publicly available complete S. iniae genomes revealed discrepancies that are likely due to misassembly in the genomes of isolates ISET0901 and ISNO. We also determined by long-range PCR that a tandem duplication of an rRNA region in the PacBio assembly of QMA0248 was an assembly error. A similar rRNA duplication in the PacBio genome of S. iniae 89353 may also be a misassembly. Our study not only highlights assembly problems in existing genomes, but provides a high quality reference genome for S. iniae QMA0248, including manually curated mobile genetic elements, that will assist future S. iniae comparative genomic and evolutionary studies.
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