The vast majority of bacteria in the environment have yet to be cultured. Consequently, a major proportion of both genetic diversity within known gene families and an unknown number of novel gene families reside in these uncultured organisms. Isolation of these genes is limited by lack of sequence information. Where such sequence data exist, PCR directed at conserved sequence motifs recovers only partial genes.

Lateral gene transfer has been proposed as a fundamental process underlying bacterial diversity. Transposons, plasmids and phage are widespread and have been shown to significantly contribute to lateral gene transfer. However, the processes by which disparate genes are assembled and integrated into the host regulatory network to yield new phenotypes are poorly known. Recent discoveries about the integron/gene cassette system indicate it has the potential to play a role in this process. Gene cassettes are small mobile elements typically consisting of a promoterless orf and a recombination site. Integrons are capable of acquisition and re-arrangement of gene cassettes and of the expression of their associated genes. The potential of the integron/gene cassette system is thus largely determined by the diversity contained within the cassette pool and the rate at which integrons sample this pool. We show here using a polymerase chain reaction (PCR) approach by which the environmental gene cassette (EGC) metagenome can be directly sampled that this metagenome contains both protein-coding and non-protein coding genes. Environmental gene cassette-associated recombination sites showed greater diversity than previously seen in integron arrays. Class 1 integrons were shown to be capable of accessing this gene pool through tests of recombinational activity with a representative range of EGCs. We propose that gene cassettes represent a vast, prepackaged genetic resource that could be thought of as a metagenomic template for bacterial evolution.

Integrons are genetic elements known for their role in the acquisition and expression of genes conferring antibiotic resistance. Such acquisition is mediated by an integron-encoded integrase, which captures genes that are part of gene cassettes. To test whether integrons occur in environments with no known history of antibiotic exposure, PCR primers were designed to conserved regions of the integrase gene and the gene cassette recombination site. Amplicons generated from four environmental DNA samples contained features typical of the integrons found in antibiotic-resistant and pathogenic bacteria. The sequence diversity of the integrase genes in these clones was sufficient to classify them within three new classes of integron. Since they are derived from environments not associated with antibiotic use, integrons appear to be more prevalent in bacteria than previously observed.

No abstract

Many sequenced strains of Acinetobacter baumannii are established nosocomial pathogens capable of resistance to multiple antimicrobials. Community-acquired A. baumannii in contrast, comprise a minor proportion of all A. baumannii infections and are highly susceptible to antimicrobial treatment. However, these infections also present acute clinical manifestations associated with high reported rates of mortality. We report the complete 3.70 Mbp genome of A. baumannii D1279779, previously isolated from the bacteraemic infection of an Indigenous Australian; this strain represents the first community-acquired A. baumannii to be sequenced. Comparative analysis of currently published A. baumannii genomes identified twenty-four accessory gene clusters present in D1279779. These accessory elements were predicted to encode a range of functions including polysaccharide biosynthesis, type I DNA restriction-modification, and the metabolism of novel carbonaceous and nitrogenous compounds. Conversely, twenty genomic regions present in previously sequenced A. baumannii strains were absent in D1279779, including gene clusters involved in the catabolism of 4-hydroxybenzoate and glucarate, and the A. baumannii antibiotic resistance island, known to bestow resistance to multiple antimicrobials in nosocomial strains. Phenomic analysis utilising the Biolog Phenotype Microarray system indicated that A. baumannii D1279779 can utilise a broader range of carbon and nitrogen sources than international clone I and clone II nosocomial isolates. However, D1279779 was more sensitive to antimicrobial compounds, particularly beta-lactams, tetracyclines and sulphonamides. The combined genomic and phenomic analyses have provided insight into the features distinguishing A. baumannii isolated from community-acquired and nosocomial infections.