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Bacterial surface components and extracellular compounds such as exopolysaccharides (EPSs) are crucial for interactions between cells, tolerance to different types of stress, and host colonization. Sinorhizobium meliloti produces two EPSs: Succinoglycan (EPS I), which is involved in the establishment of symbiosis with Medicago sativa, and galactoglucan (EPS II), associated with biofilm formation and the promotion of aggregation. Here, we aimed to assess their role in aggregative interactions between cells of the same strain of a given species (auto-aggregation), and between genetically different strains of the same or different species (intra- or intergeneric coaggregation). To do this, we used S. meliloti mutants which are defective in the production of EPS I, EPS II, or both. Macroscopic and microscopic coaggregation tests were performed with combinations or pairs of different bacterial strains. The EPS II-producing strains were more capable of coaggregation than those that cannot produce EPS II. This was true both for coaggregations between different S. meliloti strains, and between S. meliloti and other common rhizobacteria of agricultural relevance, such as Pseudomonas fluorescens and Azospirillum brasilense. The exogenous addition of EPS II strongly promoted coaggregation, thus confirming the polymer's importance for this phenotype. EPS II may therefore be a key factor in events of physiological significance for environmental survival, such as aggregative interactions and biofilm development. Furthermore, it might be a connecting molecule with relevant properties at an ecological, biotechnological, and agricultural level.

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Bacterial surface molecules are crucial for the establishment of a successful rhizobia-legume symbiosis, and, in most bacteria, are also critical for adherence properties, surface colonization, and as a barrier for defense. Rhizobial mutants defective in the production of exopolysaccharides (EPSs), lipopolysaccharides (LPSs), or capsular polysaccharides are usually affected in symbiosis with their plant hosts. In the present study, we evaluated the role of the combined effects of LPS and EPS II in cell-to-cell and cell-to-surface interactions in Sinorhizobium meliloti by studying planktonic cell autoaggregation, biofilm formation, and symbiosis with the host plant Medicago sativa. The lpsB mutant, which has a defective core portion of LPS, exhibited a reduction in biofilm formation on abiotic surfaces as well as altered biofilm architecture compared with the wild-type Rm8530 strain. Atomic force microscopy and confocal laser microscopy revealed an increase in polar cell-to-cell interactions in the lpsB mutant, which might account for the biofilm deficiency. However, a certain level of biofilm development was observed in the lpsB strain compared with the EPS II-defective mutant strains. Autoaggregation experiments carried out with LPS and EPS mutant strains showed that both polysaccharides have an impact on the cell-to-cell adhesive interactions of planktonic bacteria. Although the lpsB mutation and the loss of EPS II production strongly stimulated early attachment to alfalfa roots, the number of nodules induced in M. sativa was not increased. Taken together, this work demonstrates that S. meliloti interactions with biotic and abiotic surfaces depend on the interplay between LPS and EPS II.

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Cáhuil Lagoon in central Chile harbors distinct microbial communities in various solar salterns that are arranged as interconnected ponds with increasing salt concentrations. Here, we report the metagenome of the 3.0- to 0.2-µm fraction of the microbial community present in a crystallizer pond with 34% salinity.

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The role of bacterial surface components in combination with bacterial functional signals in the process of biofilm formation has been increasingly studied in recent years. Plants support a diverse array of bacteria on or in their roots, transport vessels, stems, and leaves. These plant-associated bacteria have important effects on plant health and productivity. Biofilm formation on plants is associated with symbiotic and pathogenic responses, but how plants regulate such associations is unclear. Certain bacteria in biofilm matrices have been found to induce plant growth and to protect plants from phytopathogens (a process termed biocontrol), whereas others are involved in pathogenesis. In this review, we systematically describe the various components and mechanisms involved in bacterial biofilm formation and attachment to plant surfaces and the relationships of these mechanisms to bacterial activity and survival.

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Bacteria of the genus Bradyrhizobium are able to establish a symbiotic relationship with peanut (Arachis hypogaea) root cells and to fix atmospheric nitrogen by converting it to nitrogenous compounds. Quorum sensing (QS) is a cell-cell communication mechanism employed by a variety of bacterial species to coordinate behavior at a community level through regulation of gene expression. The QS process depends on bacterial production of various signaling molecules, among which the N-acylhomoserine lactones (AHLs) are most commonly used by Gram-negative bacteria. Some previous reports have shown the production of QS signaling molecules by various rhizobia, but little is known regarding mechanisms of communication among peanut-nodulating strains. The aims of this study were to identify and characterize QS signals produced by peanut-nodulating bradyrhizobial strains and to evaluate their effects on processes related to cell interaction. Detection of AHLs in 53 rhizobial strains was performed using the biosensor strains Agrobacterium tumefaciens NTL4 (pZLR4) and Chromobacterium violaceum CV026 for AHLs with long and short acyl chains, respectively. None of the strains screened were found to produce AHLs with short acyl chains, but 14 strains produced AHLs with long acyl chains. These 14 AHL-producing strains were further studied by quantification of ß-galactosidase activity levels (AHL-like inducer activity) in NTL4 (pZLR4). Strains displaying moderate to high levels of AHL-like inducer activity were subjected to chemical identification of signaling molecules by high-performance liquid chromatography coupled to mass spectrometry (LC-MS/MS). For each AHL-producing strain, we found at least four different AHLs, corresponding to N-hexanoyl-DL-homoserine lactone (C(6)), N-(3-oxodecanoyl)-L-homoserine lactone (3OC(10)), N-(3-oxododecanoyl)-L-homoserine lactone (3OC(12)), and N-(3-oxotetradecanoyl)-L-homoserine lactone (3OC(14)). Biological roles of 3OC10, 3OC12, and 3OC14 AHLs were evaluated in both AHL-producing and -non-producing peanut-nodulating strains. Bacterial processes related to survival and nodulation, including motility, biofilm formation, and cell aggregation, were affected or modified by the exogenous addition of increasing concentrations of synthetic AHLs. Our results clearly demonstrate the existence of cell communication mechanisms among bradyrhizobial strains symbiotic of peanut. AHLs with long acyl chains appear to be signaling molecules regulating important QS physiological processes in these bacteria.

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Sinorhizobium meliloti is a symbiotic nitrogen-fixing bacterium that elicits nodule formation on roots of alfalfa plants. S. meliloti produces two exopolysaccharides (EPSs), termed EPS I and EPS II, that are both able to promote symbiosis. EPS I and EPS II are secreted in two major fractions that reflect differing degrees of subunit polymerization, designated high- and low-molecular-weight fractions. We reported previously that EPSs are crucial for autoaggregation and biofilm formation in S. meliloti reference strains and isogenic mutants. However, the previous observations were obtained by use of "domesticated" laboratory strains, with mutations resulting from successive passages under unnatural conditions, as has been documented for reference strain Rm1021. In the present study, we analyzed the autoaggregation and biofilm formation abilities of native S. meliloti strains isolated from root nodules of alfalfa plants grown in four regions of Argentina. 16S rRNA gene analysis of all the native isolates revealed a high degree of identity with reference S. meliloti strains. PCR analysis of the expR gene of all the isolates showed that, as in the case of reference strain Rm8530, this gene is not interrupted by an insertion sequence (IS) element. A positive correlation was found between autoaggregation and biofilm formation abilities in these rhizobia, indicating that both processes depend on the same physical adhesive forces. Extracellular complementation experiments using mutants of the native strains showed that autoaggregation was dependent on EPS II production. Our results indicate that a functional EPS II synthetic pathway and its proper regulation are essential for cell-cell interactions and surface attachment of S. meliloti.

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Exopolysaccharide (EPS) production by the rhizobacterium Sinorhizobium meliloti is essential for root nodule formation on its legume host (alfalfa), and for establishment of a nitrogen-fixing symbiosis between the two partners. Production of EPS II (galactoglucan) by certain S. meliloti strains results in a mucoid colony phenotype. Other strains that are unable to produce EPS II display a dry phenotype, due to the presence of an insertion element in the gene expR, a key regulator involved in many important cellular processes, including production of low-molecular-weight EPS II. We describe a series of three programmed undergraduate biochemistry laboratory classes teaching PCR and electrophoresis procedures to detect non-functional expR loci in S. meliloti.

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Planktonic cells of Sinorhizobium meliloti, a Gram-negative symbiotic bacterium, display autoaggregation under static conditions. ExpR is a LuxR-type regulator that controls many functions in S. meliloti, including synthesis of two exopolysaccharides, EPS I (succinoglycan) and EPS II (galactoglucan). Since exopolysaccharides are important for bacterial attachment, we studied the involvement of EPS I and II in autoaggregation of S. meliloti. Presence of an intact copy of the expR locus was shown to be necessary for autoaggregation. A mutant incapable of producing EPS I displayed autoaggregation percentage similar to that of parental strain, whereas autoaggregation was significantly lower for a mutant defective in biosynthesis of EPS II. Our findings clearly indicate that EPS II is the essential component involved in autoaggregation of planktonic S. meliloti cells, and that EPS I plays no role in such aggregation.

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Bacterial surface polysaccharides are crucial for establishment of successful rhizobia-legume symbiosis, and in most bacteria, are also critical for biofilm formation and surface colonization. In Sinorhizobium meliloti, the regulatory protein MucR controls exopolysaccharide production. To clarify the relationship between exopolysaccharide synthesis and biofilm formation, we studied mucR expression under growth conditions that influence attachment to polyvinylchloride, developed a microtiter plate assay to quantify biofilm formation in S. meliloti strain Rm1021 and mutants defective in succinoglycan (EPS I) and/or galactoglucan (EPS II) production, and analyzed expression of EPS I and EPS II genes by quantitative reverse transcriptase-PCR. Consistent with previous studies of planktonic bacteria, we found that disruption of the mucR gene in Rm1021 biofilms increased EPS II, but reduced EPS I gene expression. mucR expression was not affected by environmental conditions that influence biofilm formation on polyvinylchloride, and biofilm formation by Rm1021 was independent of exopolysaccharide synthesis. Other factors on the Rm1021 cell surface, and growth conditions, presumably regulate attachment and/or growth as a biofilm on polyvinylchloride.

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