title: Evidence that compatibility of closely related replicons in Clostridium perfringens depends on linkage to parMRC-like partitioning systems of different subfamilies.
abstract: Clostridium perfringens produces an extensive repertoire of toxins and extracellular enzymes, many of which are intimately involved in the progression of disease and are encoded by genes on conjugative plasmids. In addition, many C. perfringens strains can carry up to five of these conjugative toxin or antimicrobial resistance plasmids, each of which has a similar 35kb backbone. This conserved backbone includes the tcp conjugation locus and the central control region (CCR), which encodes genes involved in plasmid regulation, replication and partitioning, including a parMRC partitioning locus. Most conjugative plasmids in C. perfringens have a conserved replication protein, raising questions as to how multiple, closely related plasmids are maintained within a single strain. Bioinformatics analysis has highlighted the presence of at least 10 different parMRC partitioning system families (parMRCA-J) in these plasmids, with differences in amino acid sequence identity between each ParM family ranging from 15% to 54%. No two plasmids that encode genes belonging to the same partitioning family have been observed in a single strain, suggesting that these families represent the basis for plasmid incompatibility. In an attempt to validate the proposed parMRC incompatibility groups, genetically marked C. perfringens plasmids encoding identical parMRCCor parMRCDhomologues or different combinations of parMRCA, parMRCCand parMRCDfamily homologues were introduced into a single strain via conjugation. The stability of each plasmid was determined using an incompatibility assay in which the plasmid profile of each strain was monitored over the course of two days in the absence of direct selection. The results showed that plasmids with identical parMRCCor parMRCDhomologues were incompatible and could not coexist in the absence of external selection. By contrast, plasmids that encoded different parMRC homologues were compatible and could coexist in the same cell in the absence of selection, with the exception of strains housing parMRCCand parMRCDcombinations, which showed a minor incompatibility phenotype. In conclusion, we have provided the first direct evidence of plasmid incompatibility in Clostridium spp. and have shown experimentally that the compatibility of conjugative C. perfringens plasmids correlates with the presence of parMRC-like partitioning systems of different phylogenetic subfamilies.
authors.affiliation: Infection and Immunity Program, Monash Biomedicine Discovery Institute, Department of Microbiology, Monash University, Victoria 3800, Australia.
authors.forename: Thomas D
authors.initials: TD
authors.lastname: Watts
authors.affiliation: Infection and Immunity Program, Monash Biomedicine Discovery Institute, Department of Microbiology, Monash University, Victoria 3800, Australia.
authors.forename: Priscilla A
authors.initials: PA
authors.lastname: Johanesen
authors.affiliation: Infection and Immunity Program, Monash Biomedicine Discovery Institute, Department of Microbiology, Monash University, Victoria 3800, Australia.
authors.forename: Dena
authors.initials: D
authors.lastname: Lyras
authors.affiliation: Infection and Immunity Program, Monash Biomedicine Discovery Institute, Department of Microbiology, Monash University, Victoria 3800, Australia.
authors.forename: Julian I
authors.initials: JI
authors.lastname: Rood
authors.affiliation: Infection and Immunity Program, Monash Biomedicine Discovery Institute, Department of Microbiology, Monash University, Victoria 3800, Australia. Electronic address: vicki.adams@monash.edu.
authors.forename: Vicki
authors.initials: V
authors.lastname: Adams
mesh_terms: D000199:Actins / Q000235:genetics* / Q000378:metabolism; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D003016:Clostridium perfringens / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D003227:Conjugation, Genetic; D004261:DNA Replication; D027101:DNA Topoisomerase IV / Q000235:genetics* / Q000378:metabolism; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D004352:Drug Resistance, Microbial / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D056426:Genetic Loci; D010957:Plasmids / Q000737:chemistry* / Q000378:metabolism; D012093:Replicon; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D000199:Actins; D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D004269:DNA, Bacterial; C086169:ParR protein, bacteria; D012097:Repressor Proteins; D027101:DNA Topoisomerase IV
keywords: Clostridium perfringens; Incompatibility; ParM; ParR; Partitioning; Toxin plasmids