Bacterial biofilms are formed by communities that are embedded in a self-produced matrix of extracellular polymeric substances (EPS). Importantly, bacteria in biofilms exhibit a set of 'emergent properties' that differ substantially from free-living bacterial cells. In this Review, we consider the fundamental role of the biofilm matrix in establishing the emergent properties of biofilms, describing how the characteristic features of biofilms - such as social cooperation, resource capture and enhanced survival of exposure to antimicrobials - all rely on the structural and functional properties of the matrix. Finally, we highlight the value of an ecological perspective in the study of the emergent properties of biofilms, which enables an appreciation of the ecological success of biofilms as habitat formers and, more generally, as a bacterial lifestyle.

Biofilms: an emergent form of bacterial life.

SUMMARY Biotic and abiotic surfaces in marine waters are rapidly colonized by microorganisms. Surface colonization and subsequent biofilm formation and development provide numerous advantages to these organisms and support critical ecological and biogeochemical functions in the changing marine environment. Microbial surface association also contributes to deleterious effects such as biofouling, biocorrosion, and the persistence and transmission of harmful or pathogenic microorganisms and their genetic determinants. The processes and mechanisms of colonization as well as key players among the surface-associated microbiota have been studied for several decades. Accumulating evidence indicates that specific cell-surface, cell-cell, and interpopulation interactions shape the composition, structure, spatiotemporal dynamics, and functions of surface-associated microbial communities. Several key microbial processes and mechanisms, including (i) surface, population, and community sensing and signaling, (ii) intraspecies and interspecies communication and interaction, and (iii) the regulatory balance between cooperation and competition, have been identified as critical for the microbial surface association lifestyle. In this review, recent progress in the study of marine microbial surface colonization and biofilm development is synthesized and discussed. Major gaps in our knowledge remain. We pose questions for targeted investigation of surface-specific community-level microbial features, answers to which would advance our understanding of surface-associated microbial community ecology and the biogeochemical functions of these communities at levels from molecular mechanistic details through systems biological integration.

Microbial Surface Colonization and Biofilm Development in Marine Environments

While interactions between roots and microorganisms have been intensively studied, we know little about interactions among root-associated microbes. We used random matrix theory-based network analysis of 16S rRNA genes to identify bacterial networks associated with wild oat (Avena fatua) over two seasons in greenhouse microcosms. Rhizosphere networks were substantially more complex than those in surrounding soils, indicating the rhizosphere has a greater potential for interactions and niche-sharing. Network complexity increased as plants grew, even as diversity decreased, highlighting that community organisation is not captured by univariate diversity. Covariations were predominantly positive (> 80%), suggesting that extensive mutualistic interactions may occur among rhizosphere bacteria; we identified quorum-based signalling as one potential strategy. Putative keystone taxa often had low relative abundances, suggesting low-abundance taxa may significantly contribute to rhizosphere function. Network complexity, a previously undescribed property of the rhizosphere microbiome, appears to be a defining characteristic of this habitat.

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The interconnected rhizosphere: High network complexity dominates rhizosphere assemblages

Disease suppressive soils typically develop after a disease outbreak due to the subsequent assembly of protective microbiota in the rhizosphere. The role of the plant immune system in the assemblage of a protective rhizosphere microbiome is largely unknown. In this study, we demonstrate that Arabidopsis thaliana specifically promotes three bacterial species in the rhizosphere upon foliar defense activation by the downy mildew pathogen Hyaloperonospora arabidopsidis. The promoted bacteria were isolated and found to interact synergistically in biofilm formation in vitro. Although separately these bacteria did not affect the plant significantly, together they induced systemic resistance against downy mildew and promoted growth of the plant. Moreover, we show that the soil-mediated legacy of a primary population of downy mildew infected plants confers enhanced protection against this pathogen in a second population of plants growing in the same soil. Together our results indicate that plants can adjust their root microbiome upon pathogen infection and specifically recruit a group of disease resistance-inducing and growth-promoting beneficial microbes, therewith potentially maximizing the chance of survival of their offspring that will grow in the same soil.

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Disease-induced assemblage of a plant-beneficial bacterial consortium.

https://www.cell.com/trends/microbiology/pdf/S0966-842X(13)00251-5.pdf

Interactions in multispecies biofilms: do they actually matter?

Biofilms that form on roots, litter and soil particles typically contain multiple bacterial species. Currently, little is known about multispecies biofilm interactions and few studies have been based on environmental isolates. Here, the prevalence of synergistic effects in biofilm formation among seven different soil isolates, cocultured in combinations of four species, was investigated. We observed greater biofilm biomass production in 63% of the four-species culture combinations tested than in biofilm formed by single-species cultures, demonstrating a high prevalence of synergism in multispecies biofilm formation. One four-species consortium, composed of Stenotrophomonas rhizophila, Xanthomonas retroflexus, Microbacterium oxydans and Paenibacillus amylolyticus, exhibited strong synergy in biofilm formation and was selected for further study. Of the four strains, X. retroflexus was the only one capable of forming abundant biofilm in isolation, under the in vitro conditions investigated. In accordance, strain-specific quantitative PCR revealed that X. retroflexus was predominant within the four-species consortium (>97% of total biofilm cell number). Despite low relative abundance of all the remaining strains, all were indispensable for the strong synergistic effect to occur within the four-species biofilm. Moreover, absolute individual strain cell numbers were significantly enhanced when compared with those of single-species biofilms, indicating that all the individual strains benefit from inclusion in the multispecies community. Our results show a high prevalence of synergy in biofilm formation in multispecies consortia isolated from a natural bacterial habitat and suggest that interspecific cooperation occurs.

/pdf/high-prevalence-of-biofilm-synergy-among-bacterial-soil-469p253rtg.pdf

High prevalence of biofilm synergy among bacterial soil isolates in cocultures indicates bacterial interspecific cooperation

Multispecies biofilms are predominant in almost all natural environments, where myriads of resident microorganisms interact with each other in both synergistic and antagonistic manners. The interspecies interactions among different bacteria are, despite the ubiquity of these communities, still poorly understood. Here, we report a rapid, reproducible and sensitive approach for quantitative screening of biofilm formation by bacteria when cultivated as mono- and multispecies biofilms, based on the Nunc-TSP lid system and crystal violet staining. The relative proportion of the individual species in a four-species biofilm was assessed using quantitative PCR based on SYBR Green I fluorescence with specific primers. The results indicated strong synergistic interactions in a four-species biofilm model community with a more than 3-fold increase in biofilm formation and demonstrated the strong dominance of two strains, Xanthomonas retroflexus and Paenibacillus amylolyticus. The developed approach can be used as a standard procedure for evaluating interspecies interactions in defined microbial communities. This will be of significant value in the quantitative study of the microbial composition of multispecies biofilms both in natural environments and infectious diseases to increase our understanding of the mechanisms that underlie cooperation, competition and fitness of individual species in mixed-species biofilms.

High-Throughput Screening of Multispecies Biofilm Formation and Quantitative PCR-Based Assessment of Individual Species Proportions, Useful for Exploring Interspecific Bacterial Interactions

It is well known that bacteria often exist in naturally formed multispecies biofilms. Within these biofilms, interspecies interactions seem to have an important role in ecological processes. Little is known about the effects of interspecies interactions on gene expression in these multispecies biofilms. This study presents a comparative gene expression analysis of the Xanthomonas retroflexus transcriptome when grown in a single-species biofilm and in dual- and four-species consortia with Stenotrophomonas rhizophila, Microbacterium oxydans and Paenibacillus amylolyticus. The results revealed complex interdependent interaction patterns in the multispecies biofilms. Many of the regulated functions are related to interactions with the external environment and suggest a high phenotypic plasticity in response to coexistence with other species. Furthermore, the changed expression of genes involved in aromatic and branched-chain amino acid biosynthesis suggests nutrient cross feeding as a contributing factor for the observed synergistic biofilm production when these four species coexists in a biofilm.

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Distinct gene expression profile of Xanthomonas retroflexus engaged in synergistic multispecies biofilm formation

Horizontal gene transfer by conjugation is common among bacterial populations in soil. It is well known that the host range of plasmids depends on several factors, including the identity of the plasmid host cell. In the present study, however, we demonstrate that the composition of the recipient community is also determining for the dissemination of a conjugative plasmid. We isolated 15 different bacterial strains from soil and assessed the conjugation frequencies of the IncP1 plasmid, pKJK10, by flow cytometry, from two different donors, Escherichia coli and Pseudomonas putida, to either 15 different bacterial strains or to the mixed community composed of all the 15 strains. We detected transfer of pKJK10 from P. putida to Stenotrophomonas rhizophila in a diparental mating, but no transfer was observed to the mixed community. In contrast, for E. coli, transfer was observed only to the mixed community, where Ochrobactrum rhizosphaerae was identified as the dominating plasmid recipient. Our results indicate that the presence of a bacterial community impacts the plasmid permissiveness by affecting the ability of strains to receive the conjugative plasmid.

Dawei Ren

Papers

Interactions in multispecies biofilms: do they actually matter?

High prevalence of biofilm synergy among bacterial soil isolates in cocultures indicates bacterial interspecific cooperation

High-Throughput Screening of Multispecies Biofilm Formation and Quantitative PCR-Based Assessment of Individual Species Proportions, Useful for Exploring Interspecific Bacterial Interactions

Distinct gene expression profile of Xanthomonas retroflexus engaged in synergistic multispecies biofilm formation

The ability of soil bacteria to receive the conjugative IncP1 plasmid, pKJK10, is different in a mixed community compared to single strains