Crucial Role of the Accessory Genome in the Evolutionary Trajectory of Acinetobacter baumannii Global Clone 1

Abstract

Acinetobacter baumannii is one of the most important nosocomial pathogens able to rapidly develop extensive drug resistance. Here, we study the role of accessory genome in the success of the globally disseminated clone 1 (GC1) with functional and genomic approaches. Comparative genomics was performed with available GC1 genomes (n = 106) against other A. baumannii high-risk and sporadic clones. Genetic traits related to accessory genome were found common and conserved along time as two novel regions of genome plasticity, and a CRISPR-Cas system acquired before clonal diversification located at the same loci as "sedentary" modules. Although identified within hotspot for recombination, other block of accessory genome was also "sedentary" in lineage 1 of GC1 with signs of microevolution as the AbaR0-type genomic island (GI) identified in A144 and in A155 strains which were maintained one month in independent experiments without antimicrobial pressure. The prophage YMC/09/02/B1251_ABA_BP was found to be "mobile" since, although it was shared by all GC1 genomes, it showed high intrinsic microevolution as well as mobility to different insertion sites. Interestingly, a wide variety of Insertion Sequences (IS), probably acquired by the flow of plasmids related to Rep_3 superfamily was found. These IS showed dissimilar genomic location amongst GC1 genomes presumably associated with promptly niche adaptation. On the other hand, a type VI secretion system and three efflux pumps were subjected to deep processes of genomic loss in A. baumannii but not in GC1. As a whole, these findings suggest that preservation of some genetic modules of accessory genome harbored by strains from different continents in combination with great plasticity of IS and varied flow of plasmids, may be central features of the genomic structure of GC1. Competition of A144 and A155 versus A118 (ST 404/ND) without antimicrobial pressure suggested a higher ability of GC1 to grow over a clone with sporadic behavior which explains, from an ecological perspective, the global achievement of this successful pandemic clone in the hospital habitat. Together, these data suggest an essential role of still unknown properties of "mobile" and "sedentary" accessory genome that is preserved over time under different antibiotic or stress conditions.

Keywords: A. baumannii; accessory genome; genomic plasticity; high-risk clone; international clone 1 (GC1).

FIGURE 1

Molecular phylogenetic analysis and antimicrobial resistance determinants of GC1 Group 1 and Outgroup Group 3. The evolutionary history was inferred by using the Maximum Likelihood method based on the General Time Reversible model. The inset legend indicates the genetic determinants highlighted. When required, blastn was used with a cut-off E-value of E^–10.

FIGURE 2

Prediction of elements of the accessory genomes identified in GC1 Group 1 genomes from this study. Genetic and physical map of GC1 chromosomes. The inner black circle belongs to A144 chromosome and the inset legend indicates the remaining GC1 Group1 chromosomes. The outer legend corresponds to RGP, bla genes, the three efflux pumps and CRIPSR/Cas found in the eighteen GC1 Group 1 genomes in the same loci, indicated with black letters and lines is also shown in the outer circle. The hot spots (HS), RGP, and prophages found in A144 are shown in red letters and lines. Black histogram represents CG content of A144 strain. Regions related to some hotspots (HS) disrupted synteny among GC1 chromosomes which also corresponded to RGP in A144 as follows, RGP2/HS8 (JQSF01000083.1: 1-33700 and JQSF01000082.1: 24920-27366), RGP3/HS12 (JQSF01000043.1: 1 -38248 and JQSF01000071.1: 12398-14538), RGP5/HS15 together with RGP6/HS16 (JQSF01000080.1: 153-15902, JQSF01000022.1, JQSF01000055.1, JQSF01000037.1, JQSF01000003.1, QSF01000040.1, JQSF01000042.1, JQSF01000065.1 and JQSF01000041.1: 1-20291), and RGP7/HS18 that corresponded to AbaR GI (JQSF01000053.1: 1-8092, JQSF01000058.1, JQSF01000063.1: 797-1612, JQSF01000054.1: 253-2358, JQSF01000030.1: 448-21672, and JQSF01000084.1: 23-13490).

FIGURE 3

Genetic organization of the AdeFGH, AdeFGH and AdeIJK RND efflux pumps and their regulatory genes. Reference sequences: adeABC and adeRS, NC_010410.1 (coordinates 1883328-1891105); adeFGH and adeL, KR297239.1; adeIJK and adeN, CP000521.1 (coordinates 3171871-3177761 to 2292697-2293350).

FIGURE 4

Representation of the AbaR0-like GI found in Acinetobacter baumannii AYE, A144 and A155 strains. The comM gene is interrupted by the insertion of the AbaR0-like GI. The region of multidrug resistance (MARR) is detailed, in this area we found variations in relation with the AbaR type GI previously reported.

FIGURE 5

Comparison of the attI1 recombination sites found in the GI of A144, A155 and AYE. The variant 1 is the typical attI1 site. Variant 2 shows the insertion of the IS1999 and a 9 bp duplication of a portion of the attI1 site. Variant 3 found in the three genomes, shows a 19 bp duplication. Variant 4 has a deletion of the 3′ end of the attI1 site. The characteristic regions of the attI1 site are marked as follows: Direct Repeat 2 (DR2), broken-line arrow; Direct Repeat 1 (DR1), double-line arrow; simple site, horizontal line; Shine-Dalgarno (SD) sequence identified for the orf-11, stars. The predicted sequence of the orf-11 is shown. The duplications are depicted in boxes. The gene cassette next to each attI1 variant is indicated and the corresponding initial nucleotides are shown in lower case. The sequences of the variants 1 to 4 of the attI1 site found in AYE correspond to the following coordinates in CU459141.1: 3.677.401-3.677.465 bp, 3.661.663-3.663.064 bp, 3.624.336-3.624.419 bp and 3.668.061-3.68.101 bp, respectively.

FIGURE 6

Distribution of hotspots along the core-genome of Acinetobacter baumannii GC1 Group 1 genomes. The gene order of A. baumannii AYE strain was used as a reference (see section “Materials and Methods”). The bars represent the number of different gene families in all the genomes found between two consecutive genes of the core-genome of GC1 genomes.