Araceli Dávalos

Antibiotic-, biocide-, and metal-resistance genes co-occur in the genomes of multidrug-resistant bacteria. This relationship is more complex than mere physical co-localization. Instead, it arises from co-selection pressures that favor co-regulation and co-resistance. The genomic basis of this co-occurrence remains unclear. We predicted the co-occurrence of Cu(I)-ATPases with other metal-extrusion systems in Methylobacterium extorquens. In this study, we identified a more complex co-occurrence of Cu(I)-ATPases in the pathogen Brucella anthropi ATCC 49188. It comprises five collinear gene clusters spanning 48.5 kb and may be involved in multimetal homeostasis, redox homeostasis, conjugation, and the transport of miscellaneous molecules. We found highly homologous clusters in 43 strains across 3 alpha-proteobacterial orders, isolated from diverse habitats in different countries. Despite these geographical differences and phylogenetic distances, most orthologous genes remain syntenic. We called this gene arrangement a Collinear Synteny Block (CSB). In 20 species, these CSBs reside within an inferred ~110 kb genomic island, which may have contributed to their dispersion. CSBs co-occur with potential antibiotic-resistance genes scattered across the same replicon. This study will advance our understanding of the organization, evolution, and dispersal of resistance genes in bacteria. This genome-mining study is entirely predictive, and the proposed hypothesis requires experimental validation.

Antibiotic-, biocide-, and metal-resistance genes co-occur in the genomes of multidrug-resistant bacteria. This relationship is more complex than mere physical co-localization. Instead, it arises from co-selection pressures that favor co-regulation and co-resistance. The genomic basis of this co-occurrence remains unclear. Our studies on the co-occurrence of Cu(I)-ATPases with other metal-extrusion systems in the Methylobacterium extorquens AM1 genome revealed 3 Multimetal Homeostasis Gene Clusters (MHGCs) presumed to confer resistance to Cu/Ni/Co/Zn/Cd/Pb. In this study, we identified a more complex co-occurrence of Cu(I)-ATPases in the pathogen Brucella anthropi ATCC 49188. It comprises five collinear gene clusters (CGCs) spanning 48.5 kb and involved in multimetal and redox homeostasis, conjugation, and the transport of miscellaneous molecules. Highly homologous clusters were found in 43 strains across 3 alpha-proteobacterial orders, isolated from diverse habitats in different countries. Despite these geographical differences and phylogenetic distances, most orthologous genes remain syntenic. This arrangement of genes was named a Collinear Synteny Block (CSB). In some species, these CSBs reside within a ~110-kb genomic island, which explains their widespread occurrence. The CSB co-occurs with potential antibiotic-resistant genes scattered across the same replicon. This study will advance our understanding of the organization, evolution, and dispersal of resistance genes in bacteria.

Rhizobia are alpha-proteobacteria commonly found in soil and root nodules of legumes. It was recently reported that nitrogen-fixing rhizobia also inhabit legume seeds. In this study, we examined whole-genome sequences of seven strains of rhizobia isolated from seeds of common bean (Phaseolus vulgaris). Rhizobial strains included in this study belonged to three different species, including Rhizobium phaseoli, R. leguminosarum, and R. grahamii. Genome sequence analyses revealed that six of the strains formed three pairs of highly related strains. Both strains comprising a pair shared all but one plasmid. In two out of three pairs, one of the member strains was effective in nodulation and nitrogen fixation, whereas the other was ineffective. The genome of the ineffective strain in each pair lacked several genes responsible for symbiosis, including nod, nif, and fix genes, whereas that of the effective strain harbored the corresponding genes in clusters, suggesting that recombination events provoked gene loss in ineffective strains. Comparisons of genomic sequences between seed strains and nodule strains of the same species showed high conservation of chromosomal sequences and lower conservation of plasmid sequences. Approximately 70% of all genes were shared among the strains of each species. However, paralogs were more abundant in seed strains than in nodule strains. Functional analysis showed that seed strains were particularly enriched in genes involved in the transport and metabolism of amino acids and carbohydrates, biosynthesis of cofactors and in transposons and prophages. Genomes of seed strains harbored several intact prophages, one of which was inserted at exactly the same genomic position in three strains of R. phaseoli and R. leguminosarum. The R. grahamii strain carried a prophage similar to a gene transfer agent (GTA); this represents the first GTA reported for this genus. Seeds represent a niche for bacteria; their access by rhizobia possibly triggered the infection of phages, recombination, loss or gain of plasmids, and loss of symbiosis genes. This process probably represents ongoing evolution that will eventually convert these strains into obligate endophytes.

Background: A traditional concept in bacterial genetics states that housekeeping genes, those involved in basic metabolic functions needed for maintenance of the cell, are encoded in the chromosome, whereas genes required for dealing with challenging environmental conditions are located in plasmids. Exceptions to this rule have emerged from genomic sequence data of bacteria with multipartite genomes. The genome sequence ofR. etliCFN42 predicts the presence ofpanCandpanBgenes clustered together on the 642 kb plasmid p42f and a second copy ofpanBon plasmid p42e. They encode putative pantothenate biosynthesis enzymes (pantoate-β-alanine ligase and 3-methyl-2-oxobutanoate hydroxymethyltransferase, respectively). Due to their ubiquitous distribution and relevance in the central metabolism of the cell, these genes are considered part of the core genome; thus, their occurrence in a plasmid is noteworthy. In this study we investigate the contribution of these genes to pantothenate biosynthesis, examine whether their presence in plasmids is a prevalent characteristic of theRhizobialeswith multipartite genomes, and assess the possibility that thepanCBgenes may have reached plasmids by horizontal gene transfer.Results: Analysis of mutants confirmed that thepanCandpanBgenes located on plasmid p42f are indispensable for the synthesis of pantothenate. A screening of the location ofpanCBgenes among members of theRhizobialesshowed that onlyR. etliandR. leguminosarumstrains carrypanCBgenes in plasmids. ThepanCBphylogeny attested a common origin for chromosomal and plasmid-bornepanCBsequences, suggesting that theR. etliandR. leguminosarum panCBgenes are orthologs rather than xenologs. ThepanCBgenes could not totally restore the ability of a strain cured of plasmid p42f to grow in minimal medium.Conclusions: This study shows experimental evidence that corepanCBgenes located in plasmids ofR. etliandR. leguminosarumare indispensable for the synthesis of pantothenate. The unusual presence ofpanCBgenes in plasmids ofRhizobialesmay be due to an intragenomic transfer from chromosome to plasmid. Plasmid p42f encodes other functions required for growth in minimal medium. Our results support the hypothesis of cooperation among different replicons for basic cellular functions in multipartite rhizobia genomes.