Horizontal gene transfer contributes to the diversification and adaptation of micro-organisms. Apart from the core genes that are present in all strains of the same species and are the minimum necessary for survival under optimal growth conditions, bacterial genomes also harbor a variable number of accessory genes that are acquired by horizontal gene transfer, many of which are crucial for bacterial adaptation and survival. The sum of the core genes and accessory genes across the species represent what is regarded as the 'supragenome'. The accessory genes are frequently aggregated into heterogeneous sets of hitherto ill-defined structures called genomic islands, the origins of which are unknown. We recently reported on a set of genomic islands found among Proteobacteria that shared a common ancestor and were coherently structured [1
]. This study suggested for the first time that a family of genomic islands had a deep evolutionary history. However, further evidence indicated they were also capable of propagation by self-directed transfer through conjugation and replication [2
]. The relationships between species-specific subfamilies of this family of genomic islands found among Proteobacteria have not been determined.
In general, genomic islands, although poorly defined, have been regarded as segments of DNA acquired by horizontal gene transfer, with major features that include the following: GC content that is usually different from the rest of the genome; common insertion in tRNA genes; direct repeated DNA sequences at the ends; and the presence of genes such as integrases, transposases, or insertion sequences. Genomic islands often offer selective advantages; thus, according to their gene content, they can be described as pathogenicity, symbiosis, metabolic, fitness, or resistance islands [4
]. Whether all genomic islands will be classified into related families remains to be seen.
The family of genomic islands we previously reported was identified through investigations into the origins of antibiotic resistance that emerged in Haemophilus influenzae
in the early 1970s [5
]. Determining the origins of antibiotic resistance focused on the sequence of an exemplar genomic island named ICEHin1056
was shown to belong to a family of genomic islands with deep evolutionary origins found among Proteobacteria, including Yersinia enterocolitica
, Salmonella enterica
serovar Typhi, Pseudomonas fluorescens
, Ralstonia metallidurans
sp. B13, and Pseudomonas aeruginosa
]. These islands functioned as integrative and conjugative elements (ICEs). Conjugation, the process for self-directed transfer of elements between bacteria, is facilitated in this family of genomic islands by a process involving a novel type IV secretion system (T4SS) [2
]. Furthermore, replication, transfer, and integration of these genomic islands into recipient strains of the host species has been demonstrated [2
]. These limited data indicate a semi-autonomous existence for species-specific subfamilies of these genomic islands; however, further evidence is needed.
Despite an unknown potential for horizontal transfer between species, the islands we have studied [1
] show striking evidence of a phylogeny shaped by descent within the deep evolutionary history of their host bacterial species. In particular, a single common ancestor can be recognized for genomic islands carried by H. influenzae
, Haemophilus ducreyi
, and Haemophilus somnus
. The contrast between evidence for phylogenetic descent on the one hand [1
] and for potential horizontal spread by conjugation on the other [2
] raised questions about the factors both promoting and limiting conjugative spread of these genomic islands. A more thorough understanding of how these genomic islands are evolving and functionally behaving may explain the spread of antibiotic resistance in H. influenzae
. Accordingly, a detailed comparative investigation of multiple examples of the ICEHin1056
subfamily was undertaken.
To address the questions raised above, we report on sequence and functional analyses of seven genomic islands identified in H. influenzae and Haemophilus parainfluenzae. A set of shared core genes predicted to encode proteins involved in replication and T4SS-dependent transfer is described, and we show that these islands also carry a variety of accessory genes that contribute to genetic diversity. The extent and distribution of accessory gene content of this subfamily of genomic islands is similar to that reported for the overall supragenome of the bacterial host H. influenzae. The findings suggest substantial variation in sequence and gene content that is inconsistent with a historically recent acquisition and clonal expansion of these islands. However, the transposons containing antibiotic resistance genes exhibit features indicating that they have more recently been acquired. The conjugative and antibiotic susceptibility phenotypes are compared between islands and the observed variation is attributed to associated background diversity in the bacterial host genomes.