Exploring the evolutionary response of Bacillus simplex strains to the slope-specific habitats of 'Evolution Canyon' I and II, Israel, we report here on presumably adaptive differences in fatty acid (FA) content that correlate with one particular feature of the habitats, temperature difference. These two canyons represent similar ecological sites, separated by 40 km, in which the orientation of the sun yields a strong sun-exposed and hot 'African' south-facing slope versus a rather cooler and mesic-lush 'European' north-facing slope within a distance of only 50-400 m. Among 131 strains, which are identical in their 16S sequences, those assigned genetically to the 'African' ecotypes express phenotypically generally more high-temperature-tolerance-providing iso-branched FAs than strains assigned to the 'European' ecotypes when grown at 20 degrees C, 28 degrees C and 40 degrees C. Conversely, 'European' lineages express larger amounts of low-temperature-tolerance-providing anteiso-branched and non-saturated FAs when grown at the same temperatures. Moreover, 'African' ecotypes show a stronger adjustment of their high- and low-temperature-tolerance-providing FAs in response to low temperatures, which suggests that, as a result of temperature adaptation, 'African' and 'European' ecotypes have evolved different reaction norms within their phenotypic plasticity response. Thus, bacterial adaptive microevolution may include such multigenic and highly complex organs as the bacterial cell membrane. The results contribute to our understanding of the speciation process among the 'Evolution Canyon' B. simplex ecotypes.

The central questions of bacterial ecology and evolution require a method to consistently demarcate, from the vast and diverse set of bacterial cells within a natural community, the groups playing ecologically distinct roles (ecotypes). Because of a lack of theory-based guidelines, current methods in bacterial systematics fail to divide the bacterial domain of life into meaningful units of ecology and evolution. We introduce a sequence-based approach ("ecotype simulation") to model the evolutionary dynamics of bacterial populations and to identify ecotypes within a natural community, focusing here on two Bacillus clades surveyed from the "Evolution Canyons" of Israel. This approach has identified multiple ecotypes within traditional species, with each predicted to be an ecologically distinct lineage; many such ecotypes were confirmed to be ecologically distinct, with specialization to different canyon slopes with different solar exposures. Ecotype simulation provides a long-needed natural foundation for microbial ecology and systematics.

The presence and in vitro expression of homologues to 22 bacterial human virulence determinants amongst culturable soil bacteria were investigated. About 25% of the bacterial isolates contained virulence gene homologues representing toxin (hblA, cytK2), adhesin (fimH), regulator (phoQ) and resistance (yfbI) determinants in pathogenic bacteria. The homologues of the toxin genes were found in Actinobacteria and Firmicutes (hblA), and in Firmicutes and Alpha- and Gammaproteobacteria (cytK2). The homologues to the type 1 fimbrial adhesin gene, fimH, and the L-Ara4N transferase gene, yfbI, were observed in Actinobacteria, Firmicutes and Gammaproteobacteria. The regulator gene, phoQ, was only found in Gammaproteobacteria. The presence of cytK2 in Alpha- and Gammaproteobacteria, fimH in Actinobacteria and Firmicutes, and hblA in Actinobacteria has not previously been described. A close sequence similarity (84-100%) was observed between the genes of environmental and clinical isolates, and expression assays suggested that the genes in some cases were expressed in vitro. The presence of functional virulence gene homologues underpins their importance for the survival of environmental bacteria. Furthermore, the high degree of sequence conservation to clinical sequences indicates that natural environments may be 'evolutionary cribs' of emerging pathogens.

A hallmark of the Gram-positive bacteria, such as the soil-dwelling bacterium Bacillus subtilis, is their cell wall. Here, we report that d-leucine and flavomycin, biofilm inhibitors targeting the cell wall, activate the �]-lactamase PenP. This �]-lactamase contributes to ampicillin resistance in B. subtilis under all conditions tested. In contrast, both Spo0A, a master regulator of nutritional stress, and the general cell wall stress response, differentially contribute to �]-lactam resistance under different conditions. To test whether �]-lactam resistance and �]-lactamase genes are widespread in other Bacilli, we isolated Bacillus species from undisturbed soils, and found that their genomes can encode up to five �]-lactamases with differentiated activity spectra. Surprisingly, the activity of environmental �]-lactamases and PenP, as well as the general stress response, resulted in a similarly reduced lag phase of the culture in the presence of �]-lactam antibiotics, with little or no impact on the logarithmic growth rate. The length of the lag phase may determine the outcome of the competition between �]-lactams and �]-lactamases producers. Overall, our work suggests that antibiotic resistance genes in B. subtilis and related species are ancient and widespread, and could be selected by interspecies competition in undisturbed soils.

We sequenced a naturally competent bacterial isolate, WY10, cultured from a Wyoming soil sample. Sequence analysis revealed that WY10 is a novel strain of Bacillus simplex To our knowledge, WY10 is the first B. simplex strain to be characterized as naturally competent for DNA uptake by transformation.