The control of virulence gene expression in the human pathogen Staphylococcus aureus is under the partial control of the two-component quorum-sensing system encoded by genes of the agr locus. The product of the agrA gene has been shown by amino acid sequence similarity to be the putative response regulator; however, binding of AgrA to promoters under its control has not yet been demonstrated. In this study, we isolated and purified soluble AgrA by expression under osmotic shock conditions and ion-exchange chromatography. Purified AgrA showed high-affinity binding to the RNAIII-agr intergenic region by electrophoretic mobility shift assays. Binding was localized by DNase I protection assays to a pair of direct repeats in the P2 and P3 promoter regions of the agr locus. We found that this binding was enhanced by the addition of the small phosphoryl donor, acetyl phosphate. The difference in binding affinity between these two promoters was found to result from a 2-bp difference between the downstream direct repeats of the P2 and P3 sites. Mutation of these base pairs in the P3 site to match those found in the P2 site increased the affinity of AgrA for the P3 site relative to that for the P2 site. These results are consistent with the function of AgrA as a response regulator with recognition sites in the promoter regions of RNAIII and the agr locus.

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Low‐temperature pyrolysis of biomass produces a product known as biochar The incorporation of this material into the soil has been advocated as a C sequestration method. Biochar also has the potential to influence the soil N cycle by altering nitrification rates and by adsorbing or NH3 Biochar can be incorporated into the soil during renovation of intensively managed pasture soils. These managed pastures are a significant source of N2O, a greenhouse gas, produced in ruminant urine patches. We hypothesized that biochar effects on the N cycle could reduce the soil inorganic‐N pool available for N2O‐producing mechanisms. A laboratory study was performed to examine the effect of biochar incorporation into soil (20 Mg ha−1) on N2O‐N and NH3–N fluxes, and inorganic‐N transformations, following the application of bovine urine (760 kg N ha−1). Treatments included controls (soil only and soil plus biochar), and two urine treatments (soil plus urine and soil plus biochar plus urine). Fluxes of N2O from the biochar plus urine treatment were generally higher than from urine alone during the first 30 d, but after 50 d there was no significant difference (P = 0.11) in terms of cumulative N2O‐N emitted as a percentage of the urine N applied during the 53‐d period; however, NH3–N fluxes were enhanced by approximately 3% of the N applied in the biochar plus urine treatment compared with the urine‐only treatment after 17 d. Soil inorganic‐N pools differed between treatments, with higher concentrations in the presence of biochar, indicative of lower rates of nitrification. The inorganic‐N pool available for N2O‐producing mechanisms was not reduced, however, by adding biochar.

MutS proteins are ubiquitous in cellular organisms and have important roles in DNA mismatch repair or recombination. In the virus world, the amoeba-infecting Mimivirus, as well as the recently sequenced Cafeteria roenbergensis virus are known to encode a MutS related to the homologs found in octocorals and e-proteobacteria. To explore the presence of MutS proteins in other viral genomes, we performed a genomic survey of four giant viruses ('giruses') (Pyramimonas orientalis virus (PoV), Phaeocystis pouchetii virus (PpV), Chrysochromulina ericina virus (CeV) and Heterocapsa circularisquama DNA virus (HcDNAV)) that infect unicellular marine algae. Our analysis revealed the presence of a close homolog of Mimivirus MutS in all the analyzed giruses. These viral homologs possess a specific domain structure, including a C-terminal HNH-endonuclease domain, defining the new MutS7 subfamily. We confirmed the presence of conserved mismatch recognition residues in all members of the MutS7 subfamily, suggesting their role in DNA mismatch repair rather than DNA recombination. PoV and PpV were found to contain an additional type of MutS, which we propose to call MutS8. The MutS8 proteins in PoV and PpV were found to be closely related to homologs from 'Candidatus Amoebophilus asiaticus', an obligate intracellular amoeba-symbiont belonging to the Bacteroidetes. Furthermore, our analysis revealed that MutS7 and MutS8 are abundant in marine microbial metagenomes and that a vast majority of these environmental sequences are likely of girus origin. Giruses thus seem to represent a major source of the underexplored diversity of the MutS family in the microbial world.

a b s t r a c tSoil bacterial communities mediate a wide range of ecosystem processes. In agricultural systems these processes determine soil health through bacterial roles in residue decomposition, nutrient cycling and their associations with other organisms. In this study, we investigated the abundance and composition of native soil bacterial communities under different long-term fertilization practices using quantitative real-time polymerase chain reaction (qPCR) and denaturing gradient gel electrophoresis (DGGE). Soil samples were taken from a 20-year fertilization trial at the Plastic-film Mulching Experiment Station in Northeast China, which including four treatments of control (CK), inorganic-N fertilizer amendment (N), organic manure amendment (M), and half fertilizer-N plus half organic manure (M + N), to identify the difference of soil bacterial community under inorganic fertilizer and manure amendment. The size of the general bacterial population in the N treatment was significantly smaller than those in the CK, M and M + N treatments. Soil bacterial 16S rRNA gene copy numbers were positively correlated with soil pH and total organic carbon (TOC) content. The DGGE band patterns in CK, M and M + N treatments demonstrated more similarly to each other than to those of soil with only N amendment, while canonical correspondence analysis (CCA) revealed that the bacterial community structure in the N treatment was significantly different from the treatments of CK and M bacterial community profiles. Phylogenetic analysis of bands excised from the DGGE gels identified seven major phyla (Acidobacteria, Bacteroidetes, Gemmatimonadetes, Proteobacteria, Actinobacteria, Gammaproteobacteria and Firmicutes) present in all the treatments except the N treatment, in which no Gammaproteobacteria was observed. Results from this study suggest that agricultural fertilization practices have a great impact on soil bacterial community size and structure.