Bosi et al. BMC Genomics 2011, 12:403 http://www.biomedcentral.com/1471-2164/12/403 Page 5 of 12 s G A P f o % Figure 3 Taxonomical distribution of PAGs. Relative PAGs abundance and standard deviations across the most represented genera of the dataset. Red and green bars represent statistically PAGs depleted and enriched genera, respectively. Blue bars represent genera for which the content of PAGs was not statistically different from that expected by chance. greater or lower than the observed one. This gives a p- value accounting for the statistical significance of the number of PAGs retrieved within each genus in respect to our data model, i.e. that PAGs were randomly distrib- uted within the plasmids. Overall we found that 71 (out of 106) genera pos- sessed an amount of PAGs that was higher (55 PAGs enriched genera) or lower (16 PAGs depleted genera) than that expected to occur by chance (p-value < 10-4). The two best-scoring genera in terms of PAGs con- tent were Acaryochloris and Shigella (22.5% and 17.9% of all the plasmids encoded proteins, respectively); in both cases PAGs enrichment was shown to be statisti- cally significant. Interestingly, plasmids from A. marina were already shown to possess metabolic capabilitities that were probably acquired HGT transfer [29], thus confirming their mosaic structure assessed by PAGs analysis. Similarly, the mosaicism of Shigella plasmids is a well known issue [30-32] and has been demonstrated to be biologically relevant since it has probably allowed these strains to acquire pathogenic adaptation [31,33]. Both genera resulted PAGs enriched also when lower CI thresholds were applied (respectively 48.5, 74.3 and 103.1 PAGs/plasmid at 90%, 80% and 70% threshold for Acaryochloris and 31.1, 52.4 and 66.8 in the case of Shi- gella representatives) although lowering CI values below 80% resulted in statistical inconsistency, likely due to the inclusion of too many false positives in the dataset. At all CI thresholds analyzed, also PAGs depleted plasmids span over a large taxonomic range, comprising Actinobacteria, Firmicutes, Proteobacteria and Cyanobacteria. Interestingly, we found that a-proteobac- terial plasmids (mainly from Sinorhizobium, Agrobacter- ium and Rhizobium) are higlhy represented within PAGs-depleted plasmids, suggesting that plasmids hosted by representatives of this taxonomic unit might undergo recombination/HGT events less frequently than the others. The finding that these bacteria harbor a lower number of PAGs than that expected by chance, might be accounted for by the fact that these are mainly soil inhabiting microorganisms. Indeed, it has been pro- posed that bacteria inhabiting this ecological niche might represent a less connected component of the overall plasmids-mediated HGT network [34]. Accord- ingly, this might partially explain their lower number of PAGs. Alternatively, since it has been suggested that there is considerable gene flow between replicons in the rhizobiaceae [35,36], it can be surmised that these bac- teria frequently undergo recombination with the chro- mosomes of their hosting cells. However it is noteworthy that, in most cases, compositional features of rhizobiaceae replicons are pretty similar (as, for example, the GC% content (around 60%) in Sinorhizo- bium and Rhizobium representatives along all the repli- cons inhabiting the same cell). Thus, it is absolutely possbile that a fraction of this internal recombination event(s) may remain obscure due to the composition- oriented pipeline developed in this work. Interestingly, for what concerns identified PAGs (for whose identifica- tion another compositional measure was addedd to GC % content, i.e. δ*) we found that in alpha-proteobacteria chromosomal origin PAGs are more represented in