Maxime Déraspe

The Resistome of Pseudomonas aeruginosa in Relationship to Phenotypic Susceptibility

ABSTRACT Many clinical isolates of Pseudomonas aeruginosa cause infections that are difficult to eradicate due to their resistance to a wide variety of antibiotics. Key genetic determinants of resistance were identified through genome sequences of 390 clinical isolates of P. aeruginosa, obtained from diverse geographic locations collected between 2003 and 2012 and were related to microbiological susceptibility data for meropenem, levofloxacin, and amikacin. β-Lactamases and integron cassette arrangements were enriched in the established multidrug-resistant lineages of sequence types ST111 (predominantly O12) and ST235 (O11). This study demonstrates the utility of next-generation sequencing (NGS) in defining relevant resistance elements and highlights the diversity of resistance determinants within P. aeruginosa. This information is valuable in furthering the design of diagnostics and therapeutics for the treatment of P. aeruginosa infections.

The initial state of the human gut microbiome determines its reshaping by antibiotics.

Microbiome studies have demonstrated the high inter-individual diversity of the gut microbiota. However, how the initial composition of the microbiome affects the impact of antibiotics on microbial communities is relatively unexplored. To specifically address this question, we administered a second-generation cephalosporin, cefprozil, to healthy volunteers. Stool samples gathered before antibiotic exposure, at the end of the treatment and 3 months later were analysed using shotgun metagenomic sequencing. On average, 15 billion nucleotides were sequenced for each sample. We show that standard antibiotic treatment can alter the gut microbiome in a specific, reproducible and predictable manner. The most consistent effect of the antibiotic was the increase of Lachnoclostridium bolteae in 16 out of the 18 cefprozil-exposed participants. Strikingly, we identified a subgroup of participants who were enriched in the opportunistic pathogen Enterobacter cloacae after exposure to the antibiotic, an effect linked to lower initial microbiome diversity and to a Bacteroides enterotype. Although the resistance gene content of participants’ microbiomes was altered by the antibiotic, the impact of cefprozil remained specific to individual participants. Resistance genes that were not detectable prior to treatment were observed after a 7-day course of antibiotic administration. Specifically, point mutations in beta-lactamase blaCfxA-6 were enriched after antibiotic treatment in several participants. This suggests that monitoring the initial composition of the microbiome before treatment could assist in the prevention of some of the adverse effects associated with antibiotics or other treatments.

Complete Sequence of pOZ176, a 500-Kilobase IncP-2 Plasmid Encoding IMP-9-Mediated Carbapenem Resistance, from Outbreak Isolate Pseudomonas aeruginosa 96

ABSTRACT Pseudomonas aeruginosa 96 (PA96) was isolated during a multicenter surveillance study in Guangzhou, China, in 2000. Whole-genome sequencing of this outbreak strain facilitated analysis of its IncP-2 carbapenem-resistant plasmid, pOZ176. The plasmid had a length of 500,839 bp and an average percent G+C content of 57%. Of the 618 predicted open reading frames, 65% encode hypothetical proteins. The pOZ176 backbone is not closely related to any plasmids thus far sequenced, but some similarity to pQBR103 of Pseudomonas fluorescens SBW25 was observed. Two multiresistant class 1 integrons and several insertion sequences were identified. The blaIMP-9-carrying integron contained aacA4→blaIMP-9→aacA4, flanked upstream by Tn21 tnpMRA and downstream by a complete tni operon of Tn402 and a mer module, named Tn6016. The second integron carried aacA4→catB8a→blaOXA-10 and was flanked by Tn1403-like tnpRA and a sul1-type 3′ conserved sequence (3′-CS), named Tn6217. Other features include three resistance genes similar to those of Tn5, a tellurite resistance operon, and two pil operons. The replication and maintenance systems exhibit similarity to a genomic island of Ralstonia solanacearum GM1000. Codon usage analysis suggests the recent acquisition of blaIMP-9. The origins of the integrons on pOZ176 indicated separate horizontal gene transfer events driven by antibiotic selection. The novel mosaic structure of pOZ176 suggests that it is derived from environmental bacteria.

Predictive computational phenotyping and biomarker discovery using reference-free genome comparisons.

BackgroundThe identification of genomic biomarkers is a key step towards improving diagnostic tests and therapies. We present a reference-free method for this task that relies on a k-mer representation of genomes and a machine learning algorithm that produces intelligible models. The method is computationally scalable and well-suited for whole genome sequencing studies.ResultsThe method was validated by generating models that predict the antibiotic resistance of C. difficile, M. tuberculosis, P. aeruginosa, and S. pneumoniae for 17 antibiotics. The obtained models are accurate, faithful to the biological pathways targeted by the antibiotics, and they provide insight into the process of resistance acquisition. Moreover, a theoretical analysis of the method revealed tight statistical guarantees on the accuracy of the obtained models, supporting its relevance for genomic biomarker discovery.ConclusionsOur method allows the generation of accurate and interpretable predictive models of phenotypes, which rely on a small set of genomic variations. The method is not limited to predicting antibiotic resistance in bacteria and is applicable to a variety of organisms and phenotypes. Kover, an efficient implementation of our method, is open-source and should guide biological efforts to understand a plethora of phenotypes (http://github.com/aldro61/kover/).

Genome and Plasmid Analysis of blaIMP-4 -Carrying Citrobacter freundii B38

ABSTRACT Sequencing of the blaIMP-4-carrying C. freundii B38 using the PacBio SMRT technique revealed that the genome contained a chromosome of 5,134,500 bp and three plasmids, pOZ172 (127,005 bp), pOZ181 (277,592 bp), and pOZ182 (18,467 bp). Plasmid pOZ172 was identified as IncFIIY, like pP10164-NDM and pNDM-EcGN174. It carries a class 1 integron with four cassettes (blaIMP-4-qacG2-aacA4-aphA15) and a complete hybrid tni module (tniR-tniQ-tniB-tniA). The recombination of tniR from Tn402 (identical) with tniQBA from Tn5053 (99%) occurred within the res site of Tn402/5053. The Tn402/5053-like integron, named Tn6017, was inserted into Tn1722 at the res II site. The replication, partitioning, and transfer systems of pOZ181 were similar to those of IncHI2 plasmids (e.g., R478) and contained a sul1-type class 1 integron with the cassette array orf-dfrA1-orf-gcu37-aadA5 linked to an upstream Tn1696 tnpA-tnpR and to a downstream 3′ conserved sequence (3′-CS) and ISCR1. A Tn2 transposon encoding a blaTEM-1 β-lactamase was identified on pOZ182. Other interesting resistance determinants encoded on the B38 chromosome included multidrug resistance (MDR) efflux pumps, an AmpC β-lactamase, and resistances to Cu, Ag, As, and Zn. This is the first report of a complete tni module linked to a blaIMP-4-carrying class 1 integron, which, together with other recently reported non-sul1 integrons, represents the emergence of a distinct evolutionary lineage of class 1 integrons lacking a 3′-CS (qacEΔ1-sul1). The unique cassette array, complete tni module of Tn6017, and incompatibility group of pOZ172 suggest a blaIMP-4 evolutionary pathway in C. freundii B38 different from that for other blaIMP-4 genes found in Gram-negative bacteria in the Western Pacific region.

A blaVIM-2 Plasmid Disseminating in Extensively Drug-Resistant Clinical Pseudomonas aeruginosa and Serratia marcescens Isolates

Infections caused by carbapenem-resistant Enterobacteriaceae isolates are an issue of major global concern ([1][1]). Genes coding for metallo-β-lactamases (MβLs) identified in clinical isolates are associated with mobile elements and subject to horizontal genetic transfer (HGT) events ([2][2

Phenetic Comparison of Prokaryotic Genomes Using k-mers

Abstract Bacterial genomics studies are getting more extensive and complex, requiring new ways to envision analyses. Using the Ray Surveyor software, we demonstrate that comparison of genomes based on their k-mer content allows reconstruction of phenetic trees without the need of prior data curation, such as core genome alignment of a species. We validated the methodology using simulated genomes and previously published phylogenomic studies of Streptococcus pneumoniae and Pseudomonas aeruginosa. We also investigated the relationship of specific genetic determinants with bacterial population structures. By comparing clusters from the complete genomic content of a genome population with clusters from specific functional categories of genes, we can determine how the population structures are correlated. Indeed, the strain clustering based on a subset of k-mers allows determination of its similarity with the whole genome clusters. We also applied this methodology on 42 species of bacteria to determine the correlational significance of five important bacterial genomic characteristics. For example, intrinsic resistance is more important in P. aeruginosa than in S. pneumoniae, and the former has increased correlation of its population structure with antibiotic resistance genes. The global view of the pangenome of bacteria also demonstrated the taxa-dependent interaction of population structure with antibiotic resistance, bacteriophage, plasmid, and mobile element k-mer data sets.

Partial recovery of microbiomes after antibiotic treatment.

ABSTRACT Antibiotics profoundly affect the gut microbiome and modulate microbial communities. We recently observed that antimicrobial drugs also impact the abundance and distribution of antibiotic resistance genes. In this addendum, we reanalyze our ∼1 trillion nucleotide shotgun metagenomic dataset to quantify comprehensive genomic differences at the sequence level before and after antibiotic treatment. We show that 7 day exposure to cefprozil leads to a statistically significant loss of metagenome sequences. Recovery of gut microbiomes 3 months after antibiotherapy was characterized by the emergence of new genome sequences not observed prior to antibiotic exposure. Participants with low initial gut microbiome diversity had an increased amount of sequences related to antibiotic resistance. Therefore, we suggest that while the taxonomical composition of microbiomes is partially affected by the antibiotic, the genomic content and population structure of bacterial communities is noticeably impacted.

Draft Genome Sequence of an International Clonal Lineage 1 Acinetobacter baumannii Strain from Argentina

ABSTRACT In the last few years Acinetobacter baumannii has emerged worldwide as an important nosocomial pathogen in medical institutions. Here, we present the draft genome sequence of the international clonal lineage 1 (ICL1) A. baumannii strain A144 that was isolated in a hospital in Buenos Aires City in the year 1997. The strain is susceptible to carbapenems and resistant to trimethoprim and gentamicin.