RESUMEN
Acinetobacter baumannii is an emergent bacterial pathogen that provokes many types of infections in hospitals around the world. The genome of this organism consists of a chromosome and plasmids. These plasmids vary over a wide size range and many of them have been linked to the acquisition of antibiotic-resistance genes. Our bioinformatic analyses indicate that A. baumannii plasmids belong to a small number of plasmid lineages. The general structure of these lineages seems to be very stable and consists not only of genes involved in plasmid maintenance functions but of gene sets encoding poorly characterized proteins, not obviously linked to survival in the hospital setting, and opening the possibility that they improve the parasitic properties of plasmids. An analysis of genes involved in replication, suggests that members of the same plasmid lineage are part of the same plasmid incompatibility group. The same analysis showed the necessity of classifying the Rep proteins in ten new groups, under the scheme proposed by Bertini et al. (2010). Also, we show that some plasmid lineages have the potential capacity to replicate in many bacterial genera including those embracing human pathogen species, while others seem to replicate only within the limits of the Acinetobacter genus. Moreover, some plasmid lineages are widely distributed along the A. baumannii phylogenetic tree. Despite this, a number of them lack genes involved in conjugation or mobilization functions. Interestingly, only 34.6% of the plasmids analyzed here possess antibiotic resistance genes and most of them belong to fourteen plasmid lineages of the twenty one described here. Gene flux between plasmid lineages appears primarily limited to transposable elements, which sometimes carry antibiotic resistance genes. In most plasmid lineages transposable elements and antibiotic resistance genes are secondary acquisitions. Finally, broad host-range plasmids appear to have played a crucial role.
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OBJECTIVE: To describe the clinical features, outcomes, and molecular epidemiology of an outbreak of multidrug resistant (MDR) A. baumannii. METHODS: We performed a retrospective analysis of all MDR A. baumannii isolates recovered during an outbreak from 2011 to 2015 in a tertiary care cancer hospital. Cases were classified as colonized or infected. We determined sequence types following the Bartual scheme and plasmid profiles. RESULTS: There were 106 strains of A. baumannii isolated during the study period. Sixty-six (62.3%) were considered as infection and 40 (37.7%) as colonization. The index case, identified by molecular epidemiology, was a patient with a drain transferred from a hospital outside Mexico City. Ninety-eight additional cases had the same MultiLocus Sequence Typing (MLST) 758, of which 94 also had the same plasmid profile, two had an extra plasmid, and two had a different plasmid. The remaining seven isolates belonged to different MLSTs. Fifty-three patients (50%) died within 30 days of A. baumanniii isolation: 28 (20%) in colonized and 45 (68.2%) in those classified as infection (p<0.001). In multivariate regression analysis, clinical infection and patients with hematologic neoplasm, predicted 30-day mortality. The molecular epidemiology of this outbreak showed the threat posed by the introduction of MDR strains from other institutions in a hospital of immunosuppressed patients and highlights the importance of adhering to preventive measures, including contact isolation, when admitting patients with draining wounds who have been hospitalized in other institutions.
Asunto(s)
Infecciones por Acinetobacter/tratamiento farmacológico , Infecciones por Acinetobacter/mortalidad , Infección Hospitalaria/epidemiología , Acinetobacter baumannii/genética , Acinetobacter baumannii/aislamiento & purificación , Acinetobacter baumannii/patogenicidad , Adulto , Anciano , Estudios de Casos y Controles , Brotes de Enfermedades , Resistencia a Múltiples Medicamentos/fisiología , Farmacorresistencia Bacteriana Múltiple/efectos de los fármacos , Farmacorresistencia Bacteriana Múltiple/genética , Femenino , Hospitales Generales , Humanos , Masculino , México , Pruebas de Sensibilidad Microbiana , Persona de Mediana Edad , Epidemiología Molecular/métodos , Tipificación de Secuencias Multilocus/métodos , Plásmidos/efectos de los fármacos , Plásmidos/genética , Estudios Retrospectivos , Análisis de Secuencia de ADN/métodos , beta-Lactamasas/genéticaRESUMEN
Although Acinetobacter baumannii has become one of the most important nosocomial pathogens worldwide, very little is known about the genetic identity of isolates from less developed countries in Latin America. To alleviate this, we sequenced the genomes of 16 A. baumannii isolates from Honduras. Whole-genome sequencing was conducted on 16 isolates from five Honduran Hospitals. With the sequences of these Honduran isolates and other 42 publically available genomes, a maximum likelihood phylogeny was constructed to establish the relationship between the Honduran isolates and those belonging to the International Clones (ICs). In addition, sequence type (ST) assignation was conducted by the PubMLST, and antibiotic resistance genes were identified using ResFinder. The Honduran isolates are highly diverse and contain new allele combinations under the Bartual multilocus sequence typing scheme. The most common STs were STB447/STP10 and STB758/STP156. Furthermore, none of these isolates belongs to clonal complexes related to the ICs. Antibiotic susceptibility profiles of these isolates showed that they are multidrug resistant (MDR) or extensively drug resistant (XDR). In addition, the Honduran isolates had genes involved in resistance to seven antibiotic families. For instance, several blaOXA alleles were found, including blaOXA-23 and a gene encoding the metallo-beta-lactamase NDM-1. Notably, nine of the Honduran isolates have antibiotic resistance genes to three or more antibiotic families. In summary, in this study, we unveiled an untapped source of genetic diversity of MDR and XDR isolates; notably, these isolates did not belong to the well-known ICs.
Asunto(s)
Infecciones por Acinetobacter/epidemiología , Acinetobacter baumannii/genética , Farmacorresistencia Bacteriana Múltiple/genética , Genes Bacterianos , Genoma Bacteriano , beta-Lactamasas/genética , Infecciones por Acinetobacter/tratamiento farmacológico , Infecciones por Acinetobacter/microbiología , Acinetobacter baumannii/clasificación , Acinetobacter baumannii/efectos de los fármacos , Acinetobacter baumannii/aislamiento & purificación , Antibacterianos/farmacología , Células Clonales , Monitoreo Epidemiológico , Expresión Génica , Variación Genética , Honduras/epidemiología , Humanos , Pruebas de Sensibilidad Microbiana , Tipificación de Secuencias Multilocus , Filogenia , Plásmidos/química , Plásmidos/metabolismo , Centros de Atención Terciaria , Secuenciación Completa del Genoma , beta-Lactamasas/metabolismoRESUMEN
Although genome sequencing has become a very promising approach to conduct microbial taxonomy, few labs have the resources to afford this especially when dealing with data sets of hundreds to thousands of isolates. The goal of this study was to identify the most adequate loci for inferring the phylogeny of the species within the genus Staphylococcus; with the idea that those who cannot afford whole genome sequencing can use these loci to carry out species assignation confidently. We retrieved 177 orthologous groups (OGs) by using a genome-based phylogeny and an average nucleotide identity analysis. The top 26 OGs showed topologies similar to the species tree and the concatenation of them yielded a topology almost identical to that of the species tree. Furthermore, a phylogeny of just the top seven OGs could be used for species assignment. We sequenced four staphylococcus isolates to test the 26 OGs and found that these OGs were far superior to commonly used markers for this genus. On the whole, our procedure allowed identification of the most adequate markers for inferring the phylogeny within the genus Staphylococcus. We anticipate that this approach will be employed for the identification of the most suitable markers for other bacterial genera and can be very helpful to sort out poorly classified genera.
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Genome sequencing has been useful to gain an understanding of bacterial evolution. It has been used for studying the phylogeography and/or the impact of mutation and recombination on bacterial populations. However, it has rarely been used to study gene turnover at microevolutionary scales. Here, we sequenced Mexican strains of the human pathogen Acinetobacter baumannii sampled from the same locale over a 3 year period to obtain insights into the microevolutionary dynamics of gene content variability. We found that the Mexican A. baumannii population was recently founded and has been emerging due to a rapid clonal expansion. Furthermore, we noticed that on average the Mexican strains differed from each other by over 300 genes and, notably, this gene content variation has accrued more frequently and faster than the accumulation of mutations. Moreover, due to its rapid pace, gene content variation reflects the phylogeny only at very short periods of time. Additionally, we found that the external branches of the phylogeny had almost 100 more genes than the internal branches. All in all, these results show that rapid gene turnover has been of paramount importance in producing genetic variation within this population and demonstrate the utility of genome sequencing to study alternative forms of genetic variation.
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In this study, we present the complete genome sequence of a blaOXA-58-producing Acinetobacter baumannii strain, sampled from a Mexican hospital and not related to the international clones.
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Rhizobium etli CFN42 has a multipartite genome composed of one chromosome and six large plasmids with low copy numbers, all belonging to the repABC plasmid family. All elements essential for replication and segregation of these plasmids are encoded within the repABC operon. RepA and RepB direct plasmid segregation and are involved in the transcriptional regulation of the operon, and RepC is the initiator protein of the plasmid. Here we show that in addition to RepA (repressor) and RepB (corepressor), full transcriptional repression of the operon located in the symbiotic plasmid (pRetCFN42d) of this strain requires parS, the centromere-like sequence, and the operator sequence. However, the co-expression of RepA and RepB is sufficient to induce the displacement of the parental plasmid. RepA is a Walker-type ATPase that self associates in vivo and in vitro and binds specifically to the operator region in its RepA-ADP form. In contrast, RepA-ATP is capable of binding to non-specific DNA. RepA and RepB form high molecular weight DNA-protein complexes in the presence of ATP and ADP. RepA carrying ATP-pocket motif mutations induce full repression of the repABC operon without the participation of RepB and parS. These mutants specifically bind the operator sequence in their ATP or ADP bound forms. In addition, their expression in trans exerts plasmid incompatibility against the parental plasmid. RepA and RepB expressed in trans induce plasmid incompatibility because of their ability to repress the repABC operon and not only by their capacity to distort the plasmid segregation process.
Asunto(s)
Adenosina Trifosfatasas/genética , Proteínas Bacterianas/genética , Regulación Bacteriana de la Expresión Génica , Genoma Bacteriano , Operón , Plásmidos , Rhizobium etli/genética , Adenosina Difosfato/metabolismo , Adenosina Trifosfatasas/metabolismo , Adenosina Trifosfato/metabolismo , Secuencia de Aminoácidos , Proteínas Bacterianas/metabolismo , Datos de Secuencia Molecular , Mutación , Rhizobium etli/metabolismo , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Transcripción GenéticaRESUMEN
The repABC replication/partitioning systems are commonly found in alpha-proteobacteria plasmids and in secondary chromosomes. All of the elements required for their replication and stable maintenance are encoded within a single transcription unit: the repABC operon. The repC gene encodes an initiator protein, while RepA, RepB and centromere-like sequence (parS) direct plasmid segregation. Strains containing two or more repABC plasmids are a common feature in some alpha proteobacteria groups, indicating that the repABC plasmid family embraces several incompatibility groups. Genes encoded within repABC operons are highly dynamic: each one possess its own distinctive phylogeny and homologous recombination events are common within these operons. Additionally, alpha-proteobacterial genomes contain repAB genes not associated with the ctRNA or with repC as well as plasmids whose replication depends on a ctRNA-repC module without the participation of repAB genes. Some alphaproteobacteria have repC genes clustered with other genes that are not involved in replication/partitioning functions. These atypical associations of genes could have an important role in the origin and diversification of new plasmids. Here we evaluated the functionality and possible evolutionary consequences of one of these atypical gene associations: the repAC genes present in the Rhizobium leguminosarum plasmid pRL7. The repAC genes are organized in an operon and they are capable of sustaining replication but in an unstable manner. RepC was essential for replication, and the origin of replication resides within its coding region. In contrast, RepA plays a minor role in the negative regulation of its own transcription.