Insight into biofilm-associated microbial life

Microbes cohabit our planet and are engaged in a struggle for survival though on a microscopic scale. This endeavor allows them to develop and devise means for survival and proliferation. One such strategy is the formation of biofilms leading to establishment of a protected community. Such multi-communities may consist of harmful and pathogenic microbes, and they may cause economic problems and threats to human health. Biofilms are formed when microorganisms are typically attached to support surfaces. Biofilm-associated cells are sessile and differentiated from their suspended counterparts by generation of an extracellular polymeric substance matrix, reduced growth rates, and the up- and downregulation of specific genes. Biofilm formation is a complex process regulated by diverse characteristics of the growth medium, substratum, and cell surface. Development of strategies to control or prevent biofilms requires a thorough understanding of the biofilm development process. Biofilm research has witnessed exponential growth, and exciting findings have been reported. This has led us to visualize some previously un-thought-of and fascinating events. This article aims to provide an overview of biofilm research and associated challenges.     

Is biofilm formation intrinsic to the origin of life?

Biofilms are multicellular, often surface-associated, communities of autonomous cells. Their formation is the natural mode of growth of up to 80% of microorganisms living on this planet. Biofilms refractory towards antimicrobial agents and the actions of the immune system due to their tolerance against multiple environmental stresses. But how did biofilm formation arise? Here, I argue that the biofilm lifestyle has its foundation already in the fundamental, surface-triggered chemical reactions and energy preserving mechanisms that enabled the development of life on earth. Subsequently, prototypical biofilm formation has evolved and diversified concomitantly in composition, cell morphology and regulation with the expansion of prokaryotic organisms and their radiation by occupation of diverse ecological niches. This ancient origin of biofilm formation thus mirrors the harnessing environmental conditions that have been the rule rather than the exception in microbial life. The subsequent emergence of the association of microbes, including recent human pathogens, with higher organisms can be considered as the entry into a nutritional and largely stress-protecting heaven. Nevertheless, basic mechanisms of biofilm formation have surprisingly been conserved and refunctionalized to promote sustained survival in new environments.     

Effects of community composition and growth rate on aquifer biofilm bacteria and their susceptibility to betadine disinfection

Biofilm formation and function was studied in mixed culture using 20 bacterial strains isolated from a karst aquifer. When co-cultured in a glucose-limited chemostat, Vogesella indigofera and Pseudomonas putida were the dominant planktonic and biofilm organisms respectively. Biofilm formation and resistance to the iodine disinfectant betadine were then studied with monoculture and binary cultures of V. indigofera and P. putida and a 20-strain community. Biofilm population size [measured as colony-forming units (CFU) cm⁻²] increased with increasing species diversity. Significantly larger populations formed at dilution rates (DRs) of 0.0083 h⁻¹ than at 0.033 h⁻¹. P. putida populations were higher and V. indigofera lower in binary than in monoculture biofilms, suggesting that P. putida outcompeted V. indigofera . In binary biofilms, V. indigofera , a betadine-resistant organism, enhanced the survival of P. putida , a betadine-susceptible organism. In the 20-strain biofilms, this protective effect was not observed because of low concentrations of V. indigofera (< 1% of the total population), suggesting that resistant organisms contribute to overall biofilm disinfectant resistance. Growth at 0.033 h⁻¹ enhanced survival of V. indigofera biofilms against betadine. Although DR did influence survival of the other communities, its effects were neither consistent nor significant. All told, biofilm formation and betadine resistance are complex phenomena, influenced by community composition, growth rate and betadine concentration.     

Biofilm formation by Pneumocystis spp

Pneumocystis spp. can cause a lethal pneumonia in hosts with debilitated immune systems. The manner in which these fungal infections spread throughout the lung, the life cycles of the organisms, and their strategies used for survival within the mammalian host are largely unknown, due in part to the lack of a continuous cultivation method. Biofilm formation is one strategy used by microbes for protection against environmental assaults, for communication and differentiation, and as foci for dissemination. We posited that the attachment and growth of Pneumocystis within the lung alveoli is akin to biofilm formation. An in vitro system comprised of insert wells suspended in multiwell plates containing supplemented RPMI 1640 medium supported biofilm formation by P. carinii (from rat) and P. murina (from mouse). Dramatic morphological changes accompanied the transition to a biofilm. Cyst and trophic forms became highly refractile and produced branching formations that anastomosed into large macroscopic clusters that spread across the insert. Confocal microscopy revealed stacking of viable organisms enmeshed in concanavalin A-staining extracellular matrix. Biofilms matured over a 3-week time period and could be passaged. These passaged organisms were able to cause infection in immunosuppressed rodents. Biofilm formation was inhibited by farnesol, a quorum-sensing molecule in Candida spp., suggesting that a similar communication system may be operational in the Pneumocystis biofilms. Intense staining with a monoclonal antibody to the major surface glycoproteins and an increase in (1,3)-beta-D-glucan content suggest that these components contributed to the refractile properties. Identification of this biofilm process provides a tractable in vitro system that should fundamentally advance the study of Pneumocystis.     

Biofilm and planktonic microbial communities in highly acidic soil (pH < 3) in the Soos National Nature Reserve,Czech Republic

Biofilm formation is a typical life strategy used by microorganisms populating acidic water systems. The same strategy might be used by microbes in highly acidic soils that are, however, neglected in this regard. In the present study, the microbial community in such highly acidic soil in the Soos National Nature Reserve (Czech Republic) has been investigated using high-throughput DNA sequencing and the organisms associated with biofilm life mode and those preferring planktonic life were distinguished using the biofilm trap technique. Our data show the differences between biofilm and planktonic microbiota fraction, although the majority of the organisms were capable of using both life modes. The by far most abundant prokaryotic genus was Acidiphilium and fungi were identified among the most abundant eukaryotic elements in biofilm formations. On the other hand, small flagellates from diverse taxonomical groups predominated in plankton. The application of cellulose amendment as well as the depth of sampling significantly influenced the composition of the detected microbial community.

     

Inositol signaling and plant growth

Living organisms have evolved to contain a wide variety of receptors and signaling pathways that are essential for their survival in a changing environment. Of these, the phosphoinositide pathway is one of the best conserved. The ability of the phosphoinositides to permeate both hydrophobic and hydrophilic environments, and their diverse functions within cells have contributed to their persistence in nature. In eukaryotes, phosphoinositides are essential metabolites as well as labile messengers that regulate cellular physiology while traveling within and between cells. The stereospecificity of the six hydroxyls on the inositol ring provides the basis for the functional diversity of the phosphorylated isomers that, in turn, generate a selective means of intracellular and intercellular communication for coordinating cell growth. Although such complexity presents a difficult challenge for bench scientists, it is ideal for the regulation of cellular functions in living organisms.     

Does Campylobacter jejuni Form Biofilms in Food-Related Environments?

Campylobacter jejuni is one of the most frequent causes of bacterial gastrointestinal food-borne infection worldwide. This species is part of the normal flora of the gastrointestinal tracts of animals used for food production, including poultry, which is regarded as the primary source of human Campylobacter infections. The survival and persistence of C. jejuni in food processing environments, especially in poultry processing plants, represent significant risk factors that contribute to the spread of this pathogen through the food chain. Compared to other food-borne pathogens, C. jejuni is more fastidious in its growth requirements and is very susceptible to various environmental stressors. Biofilm formation is suggested to play a significant role in the survival of C. jejuni in the food production and processing environment. The aims of this minireview were (i) to examine the evidence that C. jejuni forms biofilms and (ii) to establish the extent to which reported and largely laboratory-based studies of C. jejuni biofilms provide evidence for biofilm formation by this pathogen in food processing environments. Overall existing studies do not provide strong evidence for biofilm formation (as usually defined) by most C. jejuni strains in food-related environments under the combined conditions of atmosphere, temperature, and shear that they are likely to encounter. Simple attachment to and survival on surfaces and in existing biofilms of other species are far more likely to contribute to C. jejuni survival in food-related environments based on our current understanding of this species.     

Activation of potassium channels during metabolite detoxification in Escherichia coli

In bacteria the detoxification of compounds as diverse as methylglyoxal and chlorodinitrobenzene proceeds through the formation of a glutathione adduct. In the Gram-negative bacteria, e.g. Escherichia coli, such glutathione adducts activate one, or both, of a pair of potassium efflux systems KefB and KefC. These systems share many of the properties of cation-translocating channels in eukaryotes. The activity of these systems has been found to be present in a range of Gram-negative bacteria, but not in the glutathione-deficient species of Gram-positive organisms. The conservation of the activity of these systems in a diverse range of organisms suggested a physiological role for these systems. Here we demonstrate that in E. coli cells activation of the KefB efflux system is essential for the survival of exposure to methylglyoxal. Methylglyoxal can be added to the growth medium or its synthesis can be stimulated in the cytoplasm. Under both sets of conditions survival is aided by the activity of KefB. Inhibition of KefB activity by the addition of 10 mM potassium to the growth medium stimulates methylglyoxal-induced cell death. This establishes an essential physiological function for the KefB system.     

Ecological determinants of biofilm formation

Biofilm formation is discussed in terms of the ecological processes involved in their formation. It is emphasised that any cell responds to its environment i.e. to the composition of a liquid layer a few microns thick adjacent to the cell surface. The concept of multidimensional habitat domains defining the growth limits to every species is introduced, and put into the context of competition between different organisms. Activity domains of organisms are discussed and a definition for the niche of an organism introduced. The development of a biofilm on a clean surface is described. The constant depth film fermenter is introduced as an appropriate tool for investigating spatial and temporal sequence in biofilm formation. Finally, nutrient concentration is considered and the effects of diffusion limitation on film structure explored with the aid of a cellular automaton model.     

Biofilme — die bevorzugte Lebensform der Bakterien: Flocken,Filme und Schlämme

„Biofilm”︁ describes microbial aggregates such as flocs, films and sludges; most microorganisms on earth prefer the mode of life in aggregates.They all have in common that the organisms are embedded in a matrix of extracellular polymeric substances (EPS) in which they can establish synergistic microconsortia. In this matrix, they are protected against biocides. Furthermore, the matrix acts as a sorbent for nutrients, retains exoenzymes and can be considered as a recycling yard for cellular components as well as for nutrients. Biofilms are ubiquitous and can be found even in extreme environments. In many cases, they are dominated by bacteria and/or algae, but they can also harbor higher trophical levels and represent a nutrient source for protozoa and metazoa. Biofilms are systems characterized by a vast spatial heterogeneity and temporal dynamics. Genes and signals are exchanged intensively. Thus, biofilms bear similarities to multicellular organisms.     

细菌金属离子外排系统及金属稳态调控

铁、铜、锌、锰等金属离子是各类生物体生存和增殖所必需的微量元素,可影响生物体内蛋白酶活性、免疫反应、生理过程和抗感染机制。细菌感染过程中,宿主可通过限制或提高体内环境中金属离子的浓度来抑制细菌增殖,与此同时,细菌进化出各种转运系统以适应宿主体内金属离子水平的变化。由于不同细菌的金属离子外排系统在结构和生化特性上存在变异,它们呈现出独特的金属离子外排模式。本文根据现有文献报道及本团队研究结果,对铁、铜、锌和锰离子的细菌外排系统进行讨论和总结,旨在综述目前对细菌金属离子稳态调控机制研究进展的认识,为深入理解细菌金属稳态调控相关机制提供参考。     

Crenarchaeal biofilm formation under extreme conditions

Background

Biofilm formation has been studied in much detail for a variety of bacterial species, as it plays a major role in the pathogenicity of bacteria. However, only limited information is available for the development of archaeal communities that are frequently found in many natural environments.

Methodology

We have analyzed biofilm formation in three closely related hyperthermophilic crenarchaeotes: Sulfolobus acidocaldarius, S. solfataricus and S. tokodaii. We established a microtitre plate assay adapted to high temperatures to determine how pH and temperature influence biofilm formation in these organisms. Biofilm analysis by confocal laser scanning microscopy demonstrated that the three strains form very different communities ranging from simple carpet-like structures in S. solfataricus to high density tower-like structures in S. acidocaldarius in static systems. Lectin staining indicated that all three strains produced extracellular polysaccharides containing glucose, galactose, mannose and N-acetylglucosamine once biofilm formation was initiated. While flagella mutants had no phenotype in two days old static biofilms of S. solfataricus, a UV-induced pili deletion mutant showed decreased attachment of cells.

Conclusion

The study gives first insights into formation and development of crenarchaeal biofilms in extreme environments.     

Spermidine regulates Vibrio cholerae biofilm formation via transport and signaling pathways

Vibrio cholerae , the causative agent of the devastating diarrheal disease cholera, can form biofilms on diverse biotic and abiotic surfaces. Biofilm formation is important for the survival of this organism both in its natural environment and in the human host. Development of V. cholerae biofilms are regulated by complex regulatory networks that respond to environmental signals. One of these signals, norspermidine, is a polyamine that enhances biofilm formation via the NspS/MbaA signaling system. In this work, we have investigated the role of the polyamine spermidine in regulating biofilm formation in V. cholerae . We show that spermidine import requires PotD1, an ortholog of the periplasmic substrate-binding protein of the spermidine transport system in Escherichia coli . We also show that deletion of the potD1 gene results in a significant increase in biofilm formation. We hypothesize that spermidine imported into the cell hinders biofilm formation. Exogenous spermidine further reduces biofilm formation in a PotD1-independent, but NspS/MbaA-dependent, manner. Our results suggest that polyamines affect biofilm formation in V. cholerae via multiple pathways involving both transport and signaling networks.     

Survival of Escherichia coli with and without ColE1::Tn5 after aerosol dispersal in a laboratory and a farm environment.

The survival of a laboratory strain and a naturally occurring fecal strain of Escherichia coli, with and without a Tn5-containing derivative of ColE1, was examined after aerosol dispersal in a laboratory office and a barn under ambient temperature and humidity conditions. Following the release of paired strains, air and diverse types of surfaces were assayed for the test organisms. In both environments, the number of airborne bacteria declined rapidly within the first 2 h. Longer survival was found on surfaces and varied with surface type: recovery was greatest from wood products. Organisms persisted for 1 day in the office and for up to 20 days in the barn. Survival of the fecal strain was better than that of the laboratory strain in both test environments. In general, plasmid-bearing strains fared similarly to their plasmidless parents, but in several comparisons the ColE1::Tn5-containing strain showed enhanced survival. These studies have implications for the present and proposed release of genetically engineered organisms with and without plasmid vectors.     

Survival of Escherichia coli with and without ColE1::Tn5 after aerosol dispersal in a laboratory and a farm environment

The survival of a laboratory strain and a naturally occurring fecal strain of Escherichia coli, with and without a Tn5-containing derivative of ColE1, was examined after aerosol dispersal in a laboratory office and a barn under ambient temperature and humidity conditions. Following the release of paired strains, air and diverse types of surfaces were assayed for the test organisms. In both environments, the number of airborne bacteria declined rapidly within the first 2 h. Longer survival was found on surfaces and varied with surface type: recovery was greatest from wood products. Organisms persisted for 1 day in the office and for up to 20 days in the barn. Survival of the fecal strain was better than that of the laboratory strain in both test environments. In general, plasmid-bearing strains fared similarly to their plasmidless parents, but in several comparisons the ColE1::Tn5-containing strain showed enhanced survival. These studies have implications for the present and proposed release of genetically engineered organisms with and without plasmid vectors.     

细菌菌膜的成分、调控及其与植物的关系

菌膜是细菌群落发展的一种高度组织化的群体状态。在菌膜形成过程中,细菌胞外物质EPS(Exopolysaccharides)、eDNA(Extracellular DNA)、胞外蛋白等都参与菌膜的形成,它们为菌膜提供机械稳定性,帮助细菌粘附到物体表面,促进菌膜中不同细菌间物质的循环及基因的水平转移。菌膜形成涉及到群体感应、C-di-GMP(Cyclic diguanylate monophosphate)和sRNA等一系列调控机制。土壤环境中栖息着大量的微生物,许多土壤微生物定殖于植物根际,从而与植物发生着密切的相互作用;菌膜的形成是细菌稳定定殖于植物根际的关键因素,有助于植物促生菌或致病菌在根际更好的生存。本文就菌膜的成分、调控及其与植物的关系等三个方面的内容进行综述。     

Streptococcus gordonii biofilm formation: identification of genes that code for biofilm phenotypes

Viridans streptococci, which include Streptococcus gordonii, are pioneer oral bacteria that initiate dental plaque formation. Sessile bacteria in a biofilm exhibit a mode of growth that is distinct from that of planktonic bacteria. Biofilm formation of S. gordonii Challis was characterized using an in vitro biofilm formation assay on polystyrene surfaces. The same assay was used as a nonbiased method to screen isogenic mutants generated by Tn916 transposon mutagenesis for defective biofilm formation. Biofilms formed optimally when bacteria were grown in a minimal medium under anaerobic conditions. Biofilm formation was affected by changes in pH, osmolarity, and carbohydrate content of the growth media. Eighteen biofilm-defective mutants of S. gordonii Challis were identified based on Southern hybridization with a Tn916-based probe and DNA sequences of the Tn916-flanking regions. Molecular analyses of these mutants showed that some of the genes required for biofilm formation are involved in signal transduction, peptidoglycan biosynthesis, and adhesion. These characteristics are associated with quorum sensing, osmoadaptation, and adhesion functions in oral streptococci. Only nine of the biofilm-defective mutants had defects in genes of known function, suggesting that novel aspects of bacterial physiology may play a part in biofilm formation. Further identification and characterization of biofilm-associated genes will provide insight into the molecular mechanisms of biofilm formation of oral streptococci.