Bacterial biofilms as a natural form of existence of bacteria in the environment and host organism

Advances in microscopic analysis and molecular genetics research methods promoted the acquisition of evidence that natural bacteria populations exist predominately as substrate attached biofilms. Bacteria in biofilms are able to exchange signals and display coordinated activity that is inherent to multicellular organisms. Formation of biofilm communities turned out to be one of the main survival strategies of bacteria in their ecological niche. Bacteria in attached condition in biofilm are protected from the environmental damaging factors and effects of antibacterial substances in the environment and host organism during infection. According to contemporary conception, biofilm is a continuous layer of bacterial cells that are attached to a surface and each other, and contained in a biopolymer matrix. Such bacterial communities may be composed of bacteria of one or several species, and composed of actively functioning cells as well as latent and uncultured forms. Particular attention has recently been paid to the role of biofilms in the environment and host organism. Microorganisms form biofilm on any biotic and abiotic surfaces which creates serious problems in medicine and various areas of economic activity. Currently, it is established that biofilms are one of the pathogenetic factors of chronic inflection process formation. The review presents data on ubiquity of bacteria existence as biofilms, contemporary methods of microbial community analysis, structural-functional features of bacterial biofilms. Particular attention is paid to the role of biofilm in chronic infection process formation, heightened resistance to antibiotics of bacteria in biofilms and possible mechanisms of resistance. Screening approaches for agents against biofilms in chronic infections are discussed.     

基于COMSTAT软件对微生物被膜进行定量分析的方法与意义

目的:微生物被膜是一种具有协调性、功能性和高度结构性的膜状复合物,它可以为微生物提供良好的生存环境,免受外界因素的干扰。研究发现,具有产生微生物被膜能力的细菌治病性明显增强,而现今缺乏一种分析微生物被膜的有效手段。本文探讨利用COMSTAT软件对微生物被膜进行定量分析的方法和意义,为对微生物被膜定性定量分析提供支持手段,从而为研究微生物被膜致病性提供方法理论基础。方法:以葡萄球菌为研究模型,利用激光扫描共聚焦显微镜成像技术,结合COMSTAT微生物被膜分析软件对微生物被膜的单位面积生物量、基质覆盖率、平均厚度、粗糙系数等方面进行定量分析,研究了该葡萄球菌的生物被膜生长变化过程,并考察了抗生素对其生物被膜的抑制作用。结果:在葡萄球菌生物被膜生长过程中,生物量、平均厚度以及平均扩散距离等结构指标数值都有明显增加,而粗糙度和表面积与生物量比值呈现降低趋势,表明了微生物被膜由发生向成熟的转化过程。与此同时,经10μg/mL和100μg/mL的卡那霉素处理得到的葡萄球菌微生物被膜生长受到明显抑制,且随着卡那霉素的浓度增加,抑制效果随之增加。结论:本文运用COMSTAT软件的分析方法首次从生物量、平均厚度等结构指标数值的角度描述了葡萄球菌生物被膜,从而有效评价微生物被膜发生、发展、成熟以及崩解的生长过程。该技术在研究微生物被膜形成的理论机制方面存在潜在价值,可以为研究微生物被膜治病性提供理论基础,具有理论指导意义。     

Mineral‐hosted biofilm communities in the continental deep subsurface,Deep Mine Microbial Observatory,SD, USA

Deep subsurface biofilms are estimated to host the majority of prokaryotic life on Earth, yet fundamental aspects of their ecology remain unknown. An inherent difficulty in studying subsurface biofilms is that of sample acquisition. While samples from marine and terrestrial deep subsurface fluids have revealed abundant and diverse microbial life, limited work has described the corresponding biofilms on rock fracture and pore space surfaces. The recently established Deep Mine Microbial Observatory (DeMMO) is a long‐term monitoring network at which we can explore the ecological role of biofilms in fluid‐filled fractures to depths of 1.5 km. We carried out in situ cultivation experiments with single minerals representative of DeMMO host rock to explore the ecological drivers of biodiversity and biomass in biofilm communities in the continental subsurface. Coupling cell densities to thermodynamic models of putative metabolic reactions with minerals suggests a metabolic relationship between biofilms and the minerals they colonize. Our findings indicate that minerals can significantly enhance biofilm cell densities and promote selective colonization by taxa putatively capable of extracellular electron transfer. In turn, minerals can drive significant differences in biodiversity between fluid and biofilm communities. Given our findings at DeMMO, we suggest that host rock mineralogy is an important ecological driver in deep continental biospheres.     

Should we stay or should we go: mechanisms and ecological consequences for biofilm dispersal

In most environments, bacteria reside primarily in biofilms, which are social consortia of cells that are embedded in an extracellular matrix and undergo developmental programmes resulting in a predictable biofilm 'life cycle'. Recent research on many different bacterial species has now shown that the final stage in this life cycle includes the production and release of differentiated dispersal cells. The formation of these cells and their eventual dispersal is initiated through diverse and remarkably sophisticated mechanisms, suggesting that there are strong evolutionary pressures for dispersal from an otherwise largely sessile biofilm. The evolutionary aspect of biofilm dispersal is now being explored through the integration of molecular microbiology with eukaryotic ecological and evolutionary theory, which provides a broad conceptual framework for the diversity of specific mechanisms underlying biofilm dispersal. Here, we review recent progress in this emerging field and suggest that the merging of detailed molecular mechanisms with ecological theory will significantly advance our understanding of biofilm biology and ecology.     

Interactions between Bdellovibrio and like organisms and bacteria in biofilms: beyond predator–prey dynamics

Bdellovibrio and like organisms (BALOs) prey on Gram-negative bacteria in the planktonic phase as well as in biofilms, with the ability to reduce prey populations by orders of magnitude. During the last few years, evidence has mounted for a significant ecological role for BALOs, with important implications for our understanding of microbial community dynamics as well as for applications against pathogens, including drug-resistant pathogens, in medicine, agriculture and aquaculture, and in industrial settings for various uses. However, our understanding of biofilm predation by BALOs is still very fragmentary, including gaps in their effect on biofilm structure, on prey resistance, and on evolutionary outcomes of both predators and prey. Furthermore, their impact on biofilms has been shown to reach beyond predation, as they are reported to reduce biofilm structures of non-prey cells (including Gram-positive bacteria). Here, we review the available literature on BALOs in biofilms, extending known aspects to potential mechanisms employed by the predators to grow in biofilms. Within that context, we discuss the potential ecological significance and potential future utilization of the predatory and enzymatic possibilities offered by BALOs in medical, agricultural and environmental applications.     

Evolved Biofilm: Review on the Experimental Evolution Studies of Bacillus subtilis Pellicles

For several decades, laboratory evolution has served as a powerful method to manipulate microorganisms and to explore long-term dynamics in microbial populations. Next to canonical Escherichia coli planktonic cultures, experimental evolution has expanded into alternative cultivation methods and species, opening the doors to new research questions. Bacillus subtilis, the spore-forming and root-colonizing bacterium, can easily develop in the laboratory as a liquid–air interface colonizing pellicle biofilm. Here, we summarize recent findings derived from this tractable experimental model. Clonal pellicle biofilms of B. subtilis can rapidly undergo morphological and genetic diversification creating new ecological interactions, for example, exploitation by biofilm non-producers. Moreover, long-term exposure to such matrix non-producers can modulate cooperation in biofilms, leading to different phenotypic heterogeneity pattern of matrix production with larger subpopulation of “ON” cells. Alternatively, complementary variants of biofilm non-producers, each lacking a distinct matrix component, can engage in a genetic division of labor, resulting in superior biofilm productivity compared to the “generalist” wild type. Nevertheless, inter-genetic cooperation appears to be evanescent and rapidly vanquished by individual biofilm formation strategies altering the amount or the properties of the remaining matrix component. Finally, fast-evolving mobile genetic elements can unpredictably shift intra-species interactions in B. subtilis biofilms. Understanding evolution in clonal biofilm populations will facilitate future studies in complex multispecies biofilms that are more representative of nature.     

Model parameter uncertainties in a dual-species biofilm competition model affect ecological output parameters much stronger than morphological ones

Bacterial biofilms are complex microbial depositions on immersed interfaces that form wherever the environmental conditions sustain microbial growth. Despite their name, biofilms can develop in highly irregular structures. Recently several mathematical concepts have been introduced to model these spatially structured microbial populations. Regardless of the type of model, they all have, even for microbially relatively simple systems, many parameters which generally are known at most approximately. We investigate the effect of uncertainties in model parameters on four morphological and four ecological output parameters using a nonlinear diffusion model for a biofilm in which two species compete for a shared nutrient. To this end we conduct an extensive computer simulation experiment for two different levels of data uncertainty, three different hydrodynamic conditions, and two different scenarios of bulk substrate availability. Our results indicate that input model parameter uncertainties have a much larger effect on ecological than on morphological output parameters.     

New Methods for Analysis of Spatial Distribution and Coaggregation of Microbial Populations in Complex Biofilms

In biofilms, microbial activities form gradients of substrates and electron acceptors, creating a complex landscape of microhabitats, often resulting in structured localization of the microbial populations present. To understand the dynamic interplay between and within these populations, quantitative measurements and statistical analysis of their localization patterns within the biofilms are necessary, and adequate automated tools for such analyses are needed. We have designed and applied new methods for fluorescence in situ hybridization (FISH) and digital image analysis of directionally dependent (anisotropic) multispecies biofilms. A sequential-FISH approach allowed multiple populations to be detected in a biofilm sample. This was combined with an automated tool for vertical-distribution analysis by generating in silico biofilm slices and the recently developed Inflate algorithm for coaggregation analysis of microbial populations in anisotropic biofilms. As a proof of principle, we show distinct stratification patterns of the ammonia oxidizers Nitrosomonas oligotropha subclusters I and II and the nitrite oxidizer Nitrospira sublineage I in three different types of wastewater biofilms, suggesting niche differentiation between the N. oligotropha subclusters, which could explain their coexistence in the same biofilms. Coaggregation analysis showed that N. oligotropha subcluster II aggregated closer to Nitrospira than did N. oligotropha subcluster I in a pilot plant nitrifying trickling filter (NTF) and a moving-bed biofilm reactor (MBBR), but not in a full-scale NTF, indicating important ecophysiological differences between these phylogenetically closely related subclusters. By using high-resolution quantitative methods applicable to any multispecies biofilm in general, the ecological interactions of these complex ecosystems can be understood in more detail.     

Management of biofilm-associated infections: what can we expect from recent research on biofilm lifestyles?

Biofilms are surface-associated microbial communities present in all environments. Although biofilms play important ecological roles, they also lead to negative or deleterious effects in industrial and medical settings. In the latter, high levels of antibiotic tolerance of bacterial biofilms developing on medical devices and during chronic infections determine the physiopathology of many healthcare-associated infections. Original approaches have been developed to avoid bacterial adhesion or biofilm development targetting specific mechanisms or pathways. We herein review recent data about biofilm lifestyle understanding and ways to fight against related infections.     

Mini-review: Microbial coaggregation: ubiquity and implications for biofilm development

Coaggregation is the specific recognition and adherence of genetically distinct microorganisms. Because most biofilms are polymicrobial communities, there is potential for coaggregation to play an integral role in spatiotemporal biofilm development and the moderation of biofilm community composition. However, understanding of the mechanisms contributing to coaggregation and the relevance of coaggregation to biofilm ecology is at a very early stage. The purpose of this review is to highlight recent advances in the understanding of microbial coaggregation within different environments and to describe the possible ecological ramifications of such interactions. Bacteria that coaggregate with many partner species within different environments will be highlighted, including oral streptococci and oral bridging organisms such as fusobacteria, as well as the freshwater sphingomonads and acinetobacters. Irrespective of environment, it is proposed that coaggregation is essential for the orchestrated development of multi-species biofilms.     

Controlling of microbial biofilms formation: Anti- and probiofilm agents

Controlling the formation and reconstruction of microbial biofilms is of ever increasing importance for the ecological, medical, and biotechnological aspects of biofilm studies. The goal of this review was to provide systematization and analysis of the results obtained in recent years on the modes and mechanisms of the stimulatory or inhibitory effect of extreme factors and biocidal agents on biofilm formation. Special attention is paid to controlling the formation of medically (infective diseases, implant biofouling) and technologically or biotechnologically important biofilms (bioremediation, biocorrosion, and biosynthesis of biologically active compounds).     

Biofilm—“City of microbes” or an analogue of multicellular organisms?

The definition of the term “biofilm” and the validity of the analogy between these structured microbial communities and multicellular organisms are discussed in the review. The mechanisms of biofilm formation, the types of interrelations of the components of biofilms, and the reasons for biofilm resistance to biocides and stress factors are considered in detail. The role of biofilms in microbial ecology and in biotechnology is discussed.     

Biofilm--"City of microbes" or an analogue of multicellular organisms?

The definition of the term "biofilm" and the validity of the analogy between these structured microbial communities and multicellular organisms are discussed in the review. The mechanisms of biofilm formation, the types of interrelations of the components of biofilms, and the reasons for biofilm resistance to biocides and stress factors are considered in detail. The role of biofilms in microbial ecology and in biotechnology is discussed.     

Observations of binary population biofilms

Biofilm research has focused on studies of undefined mixed microbial populations and, more recently, on investigations of monopopulation biofilms. In the first case, the biofilm is considered a homogeneous mass, ignoring the properties of individual species. The second case concentrates on the properties and processes of one microbial species in the biofilm. This article describes biofilm experiments conducted with monopopulations of Klebsiella pneumoniae and Pseudomonas aeruginosa and with binary populations of K. pneumoniae and P. aeruginosa. Process rates and stoichiometric coefficients were determined for the monopopulation and for the binary population biofilms and evaluated in light of the species distribution in the latter. Results indicate that neither the specific cellular product formation rate nor the glucose-oxygen stoichiometric ratio of K. pneumoniae or P. aeruginosa in the binary biofilm is affected by the presence of the other species. Consequently, species interaction was not observed. Although the specific cellular growth rate of K. pneumoniae is five times that of P. aeruginosa, the former species did not dominate the microbial population in the biofilm. Possible reasons for this unexpected behavior are discussed.     

Marine Biofilms as Mediators of Colonization by Marine Macroorganisms: Implications for Antifouling and Aquaculture

In the marine environment, biofilms on submerged surfaces can promote or discourage the settlement of invertebrate larvae and macroalgal spores. The settlement-mediating effects of biofilms are believed to involve a variety of biofilm attributes including surface chemistry, micro-topography, and a wide range of microbial products from small-molecule metabolites to high-molecular weight extracellular polymers. The settled organisms in turn can modify microbial species composition of biofilms and thus change the biofilm properties and dynamics. A better understanding of biofilm dynamics and chemical signals released and/or stored by biofilms will facilitate the development of antifouling and mariculture technologies. This review provides a brief account of 1) existing knowledge of marine biofilms that are relevant to settlement mediation, 2) biotechnological application of biofilms with respect to developing non-toxic antifouling technologies and improving the operation of aquaculture facilities, and 3) challenges and future directions for advancing our understanding of settlement-mediating functions of biofilms and for applying this knowledge to real-life situations.     

Interspecies bacterial interactions in biofilms

Interactions among bacterial populations can have a profound influence on the structure and physiology of microbial communities. Interspecies microbial interactions begin to influence a biofilm during the initial stages of formation, bacterial attachment and surface colonization, and continue to influence the structure and physiology of the biofilm as it develops. Although the majority of research on bacterial interactions has utilized planktonic communities, the characteristics of biofilm growth (cell positions that are relatively stable and local areas of hindered diffusion) suggest that interspecies interactions may be more significant in biofilms.     

基于微流控的生物被膜研究进展及口腔微生态系统应用展望

口腔是人体最重要的微生物储藏库之一,这些微生物通常以生物被膜的形式稳定地黏附于口腔内粘膜及牙齿表面,在病理条件下则可以深入髓腔及牙槽骨内,成为龋病、根尖周炎、牙周病等常见口腔疾病的主要病因,甚至与许多全身性疾病密切相关.在人类口腔微生态系统中,微生物种类繁多,生物被膜的构成及相互作用关系复杂,目前对其认识还十分有限;此...     

Optical Sensing of Microbial Life on Surfaces

The label-free detection of microbial cells attached to a surface is an active field of research. The field is driven by the need to understand and control the growth of biofilms in a number of applications, including basic research in natural environments, industrial facilities, and clinical devices, to name a few. Despite significant progress in the ability to monitor the growth of biofilms and related living cells, the sensitivity and selectivity of such sensors are still a challenge. We believe that among the many different technologies available for monitoring biofilm growth, optical techniques are the most promising, as they afford direct imaging and offer high sensitivity and specificity. Furthermore, as each technique offers different insights into the biofilm growth mechanism, our analysis allows us to provide an overview of the biological processes at play. In addition, we use a set of key parameters to compare state-of-the-art techniques in the field, including a critical assessment of each method, to identify the most promising types of sensors. We highlight the challenges that need to be overcome to improve the characteristics of current biofilm sensor technologies and indicate where further developments are required. In addition, we provide guidelines for selecting a suitable sensor for detecting microbial cells on a surface.