Extracellular DNAses Facilitate Antagonism and Coexistence in Bacterial Competitor-Sensing Interference Competition.

Over the last 4 decades, the rate of discovery of novel antibiotics has decreased drastically, ending the era of fortuitous antibiotic discovery. A better understanding of the biology of bacteriogenic toxins potentially helps to prospect for new antibiotics. To initiate this line of research, we quantified antagonists from two different sites at two different depths of soil and found the relative number of antagonists to correlate with the bacterial load and carbon-to-nitrogen (C/N) ratio of the soil. Consecutive studies show the importance of antagonist interactions between soil isolates and the lack of a predicted role for nutrient availability and, therefore, support an in situ role in offense for the production of toxins in environments of high bacterial loads. In addition, the production of extracellular DNAses (exDNases) and the ability to antagonize correlate strongly. Using an in domum-developed probabilistic cellular automaton model, we studied the consequences of exDNase production for both coexistence and diversity within a dynamic equilibrium. Our model demonstrates that exDNase-producing isolates involved in amensal interactions act to stabilize a community, leading to increased coexistence within a competitor-sensing interference competition environment. Our results signify that the environmental and biological cues that control natural-product formation are important for understanding antagonism and community dynamics, structure, and function, permitting the development of directed searches and the use of these insights for drug discovery. IMPORTANCE Ever since the first observation of antagonism by microorganisms by Ernest Duchesne (E. Duchesne, Contribution à l'étude de la concurrence vitale chez les microorganisms. Antagonism entre les moisissures et les microbes, These pour obtenir le grade de docteur en medicine, Lyon, France, 1897), many scientists successfully identified and applied bacteriogenic bioactive compounds from soils to cure infection. Unfortunately, overuse of antibiotics and the emergence of clinical antibiotic resistance, combined with a lack of discovery, have hampered our ability to combat infections. A deeper understanding of the biology of toxins and the cues leading to their production may elevate the success rate of the much-needed discovery of novel antibiotics. We initiated this line of research and discovered that bacterial reciprocal antagonism is associated with exDNase production in isolates from environments with high bacterial loads, while diversity may increase in environments of lower bacterial loads.

Identification and characterization of a NaCl-responsive genetic locus involved in survival during desiccation in Sinorhizobium meliloti.

The Rhizobiaceae are a bacterial family of enormous agricultural importance due to the ability of its members to fix atmospheric nitrogen in an intimate relationship with plants. Their survival as naturally occurring soil bacteria in agricultural soils as well as popular seed inocula is affected directly by drought and salinity. Survival after desiccation in the presence of NaCl is enabled by underlying genetic mechanisms in the model organism Sinorhizobium meliloti 1021. Since salt stress parallels a loss in water activity, the identification of NaCl-responsive loci may identify loci involved in survival during desiccation. This approach enabled identification of the loci asnO and ngg by their reduced ability to grow on increased NaCl concentrations, likely due to their inability to produce the osmoprotectant N-acetylglutaminylglutamine (NAGGN). In addition, the mutant harboring ngg::Tn5luxAB was affected in its ability to survive desiccation and responded to osmotic stress. The desiccation sensitivity may have been due to secondary functions of Ngg (N-acetylglutaminylglutamine synthetase)-like cell wall metabolism as suggested by the presence of a d-alanine-d-alanine ligase (dAla-dAla) domain and by sensitivity of the mutant to ß-lactam antibiotics. asnO::Tn5luxAB is expressed during the stationary phase under normal growth conditions. Amino acid sequence similarity to enzymes producing ß-lactam inhibitors and increased resistance to ß-lactam antibiotics may indicate that asnO is involved in the production of a ß-lactam inhibitor.

Diversity of oligotrichia and choreotrichia ciliates in coastal marine sediments and in overlying plankton.

Elucidating the relationship between ciliate communities in the benthos and the plankton is critical to understanding ciliate diversity in marine systems. Although data for many lineages are sparse, at least some members of the dominant marine ciliate clades Oligotrichia and Choreotrichia can be found in both plankton and benthos, in the latter either as cysts or active forms. In this study, we developed a molecular approach to address the relationship between the diversity of ciliates in the plankton and those of the underlying benthos in the same locations. Samples from plankton and sediments were compared across three sites along the New England coast, and additional subsamples were analyzed to assess reproducibility of methods. We found that sediment and plankton subsamples differed in their robustness to repeated subsampling. Sediment subsamples (i.e., 1-g aliquots from a single approximately 20-g sample) gave variable estimates of diversity, while plankton subsamples produced consistent results. These results indicate the need for additional study to determine the spatial scale over which diversity varies in marine sediments. Clustering of phylogenetic types indicates that benthic assemblages of oligotrichs and choreotrichs appear to be more like those from spatially remote benthic communities than the ciliate communities sampled in the water above them.

Complete Genome sequence of Burkholderia phymatum STM815(T), a broad host range and efficient nitrogen-fixing symbiont of Mimosa species.

Burkholderia phymatum is a soil bacterium able to develop a nitrogen-fixing symbiosis with species of the legume genus Mimosa, and is frequently found associated specifically with Mimosa pudica. The type strain of the species, STM 815(T), was isolated from a root nodule in French Guiana in 2000. The strain is an aerobic, motile, non-spore forming, Gram-negative rod, and is a highly competitive strain for nodulation compared to other Mimosa symbionts, as it also nodulates a broad range of other legume genera and species. The 8,676,562 bp genome is composed of two chromosomes (3,479,187 and 2,697,374 bp), a megaplasmid (1,904,893 bp) and a plasmid hosting the symbiotic functions (595,108 bp).

The evolution of reduced microbial killing.

Bacteria engage in a never-ending arms race in which they compete for limited resources and niche space. The outcome of this intense interaction is the evolution of a powerful arsenal of biological weapons. Perhaps the most studied of these are colicins, plasmid-based toxins produced by and active against Escherichia coli. The present study was designed to explore the molecular responses of a colicin-producing strain during serial transfer evolution. What evolutionary changes occur when colicins are produced with no target present? Can killing ability be maintained in the absence of a target? To address these, and other, questions, colicinogenic strains and a noncolicinogenic ancestor were evolved for 253 generations. Samples were taken throughout the experiment and tested for killing ability. By the 38th transfer, a decreased killing ability and an increase in fitness were observed in the colicin-producing strains. Surprisingly, DNA sequence determination of the colicin plasmids revealed no changes in plasmid sequences. However, a set of chromosomally encoded loci experienced changes in gene expression that were positively associated with the reduction in killing. The most significant expression changes were observed in DNA repair genes (which were downregulated in the evolved strains), Mg ion uptake genes (which were upregulated), and late prophage genes (which were upregulated). These results indicate a fine-tuned response to the evolutionary pressures of colicin production, with far more genes involved than had been anticipated.

The role of SOS boxes in enteric bacteriocin regulation.

Bacteriocins are a large and functionally diverse family of toxins found in all major lineages of Bacteria. Colicins, those bacteriocins produced by Escherichia coli, serve as a model system for investigations of bacteriocin structure-function relationships, genetic organization, and their ecological role and evolutionary history. Colicin expression is often dependent on host regulatory pathways (such as the SOS system), is usually confined to times of stress, and results in death of the producing cells. This study investigates the role of the SOS system in mediating this unique form of toxin expression. A comparison of all the sequenced enteric bacteriocin promoters reveals that over 75 % are regulated by dual, overlapping SOS boxes, which serve to bind two LexA repressor proteins. Furthermore, a highly conserved poly-A motif is present in both of the binding sites examined, indicating enhanced affinity of the LexA protein for the binding site. The use of gene expression analysis and deletion mutations further demonstrates that these unique LexA cooperative binding regions result in a fine tuning of bacteriocin production, limiting it to times of stress. These results suggest that the evolution of dual SOS boxes elegantly accomplishes the task of increasing the amount of toxin produced by a cell while decreasing the rate of uninduced production, effectively reducing the cost of colicin production. This hypothesis may explain why such a promoter motif is present at such high frequencies in natural populations of bacteriocin-producing enteric bacteria.