Genetically engineered microbial biosensors for in situ monitoring of environmental pollution

Microbial biosensors are compact, portable, cost effective, and simple to use, making them seem eminently suitable for the in situ monitoring of environmental pollution. One promising approach for such applications is the fusion of reporter genes with regulatory genes that are dose-dependently responsive to the target chemicals or physiological signals. Their biosensor capabilities, such as target range and sensitivity, could be improved by modification of regulatory genes. Recent uses of such genetically engineered microbial biosensors include the development of portable biosensor kits and high-throughput cell arrays on chips, optic fibers, or other platforms for on-site and on-line monitoring of environmental pollution. This mini-review discusses recent advances in microbial biosensors and their future prospects, with a focus on the development and application of genetically modified microbial biosensors for in situ environmental monitoring.     

Manganese biomining: A review

Biomining comprises of processing and extraction of metal from their ores and concentrates using microbial techniques. Currently this is used by the mining industry to extract copper, uranium and gold from low grade ores but not for low grade manganese ore in industrial scale. The study of microbial genomes, metabolites and regulatory pathways provide novel insights to the metabolism of bioleaching microorganisms and their synergistic action during bioleaching operations. This will promote understanding of the universal regulatory responses that the biomining microbial community uses to adapt to their changing environment leading to high metal recovery. Possibility exists of findings ways to imitate the entire process during industrial manganese biomining endeavor. This paper reviews the current status of manganese biomining research operations around the world, identifies factors that drive the selection of biomining as a processing technology, describes challenges in exploiting these innovations, and concludes with a discussion of Mn biomining’s future.     

Regulation of <Emphasis Type="Italic">Escherichia coli</Emphasis> Biofilm Formation (Review)

The review considers the regulatory mechanisms controlling the formation of biofilms by bacteria Escherichia coli. Under harsh conditions, microbial populations transfer to the structured mode of existence by building biofilms. The regulation of biofilm formation is a complex multistage process. Environmental signals are perceived by two-component signaling systems, which fulfill their transduction to the genome. This switches microbial cells from the planktonic motile lifestyle to the sessile.     

How to evaluate the environmental safety of microbial plant protection products: A proposal

Plant protection products with active micro-organisms are allegedly less hazardous to the environment and wildlife than synthetic chemical pesticides. Nevertheless, they need a proper pre-marketing environmental safety evaluation because of their potential toxicity and pathogenicity. Scientific and technical guidance on such a safety evaluation for regulatory purposes is scarce. Therefore, a risk decision tree is proposed to provide such guidance and to discern the acceptable from the unacceptable environmental risks. The decision tree is based on the risk criteria of the European Union. It takes integrally into account the characterisation, identification and efficacy and also emission, exposure, environmental effects and, finally, the environmental risk assessment. Case by case expert judgement remains necessary in view of limited knowledge of microbial ecology, limited experience with regulatory test protocols and taxonomic difficulties in relation to the indigenousness of active micro-organisms. The decision tree offers regulatory guidance on the environmental safety evaluation of microbial plant protection products.     

Finding A Niche for Soil Microbial Toxicity Tests in Ecological Risk Assessment

Soil microbial toxicity tests are seldom used in ecological risk assessments or in the development of regulatory criteria in the U.S. The primary reason is the lack of an explicit connection between these tests and assessment end-points. Soil microorganisms have three potential roles with respect to ecological assessment endpoints: properties of microbial communities may be end-points; microbial responses may be used to estimate effects on plant production; and microbial responses may be used as surrogates for responses of higher organisms. Rates of microbial processes are important to ecosystem function, and thus should be valued by regulatory agencies. However, the definition of the microbial assessment endpoint is often an impediment to its use in risk assessment. Decreases in rates are not always undesirable. Processes in a nutrient cycle are particularly difficult to define as endpoints, because what constitutes an adverse effect on a process is dependent on the rates of others. Microbial tests may be used as evidence in an assessment of plant production, but the dependence of plants on microbial processes is rarely considered. As assessment endpoints are better defined in the future, microbial ecologists and toxicologists should be provided with more direction for developing appropriate microbial tests.     

Ecological immunogenetics of life-history traits in a model amphibian

Major histocompatibility complex (MHC) genes determine immune repertoires and social preferences of vertebrates. Immunological regulation of microbial assemblages associated with individuals influences their sociality, and should also affect their life-history traits. We exposed Xenopus laevis tadpoles to water conditioned by adult conspecifics. Then, we analysed tadpole growth, development and survivorship as a function of MHC class I and class II peptide-binding region amino acid sequence similarities between tadpoles and frogs that conditioned the water to which they were exposed. Tadpoles approached metamorphosis earlier and suffered greater mortality when exposed to immunogenetically dissimilar frogs. The results suggest that developmental regulatory cues, microbial assemblages or both are specific to MHC genotypes. Tadpoles may associate with conspecifics with which they share microbiota to which their genotypes are well adapted.     

Microbial xylanases and their industrial applications: a review

Despite an increased knowledge of microbial xylanolytic systems in the past few years, further studies are required to achieve a complete understanding of the mechanism of xylan degradation by microorganisms and their enzymes. The enzyme system used by microbes for the metabolism of xylan is the most important tool for investigating the use of the second most abundant polysaccharide (xylan) in nature. Recent studies on microbial xylanolytic systems have generally focussed on induction of enzyme production under different conditions, purification, characterization, molecular cloning and expression, and use of enzyme predominantly for pulp bleaching. Rationale approaches to achieve these goals require a detailed knowledge of the regulatory mechanism governing enzyme production. This review will focus on complex xylan structure and the microbial enzyme complex involved in its complete breakdown, studies on xylanase regulation and production and their potential industrial applications, with special reference to biobleaching.     

Cross talk between signaling pathways in pathogen defense

Plant defense in response to microbial attack is regulated through a complex network of signaling pathways that involve three signaling molecules: salicylic acid (SA), jasmonic acid (JA) and ethylene. The SA and JA signaling pathways are mutually antagonistic. This regulatory cross talk may have evolved to allow plants to fine-tune the induction of their defenses in response to different plant pathogens.     

The role of group 1 and group 2 CD1-restricted T cells in microbial immunity

Group 1 and group 2 CD1 present both self and microbial lipid antigens to T cells. While group 1 CD1-restricted T cells are known for their ability to recognize mycobacterial glycolipid antigens, group 2 CD1-restricted T cells are recognized as regulatory T cells that can influence the outcome of innate and adaptive immune responses. The evidence that these T cells contribute to host defense against infectious diseases is reviewed.     

The emerging role of auxins as bacterial signal molecules: Potential biotechnological applications

Microorganisms are exposed in their natural niches to a wide diversity of signal molecules. Specific detection of these signals results in alterations in microbial metabolism and physiology. Auxins like indole-3-acetic acid are key phytohormones that regulate plant growth and development. Nonetheless, auxin biosynthesis is not restricted to plants but is ubiquitous in all kingdoms of life. This wide phylogenetic distribution of auxins production, together with the diversity of regulated cellular processes, have made auxins key intra- and inter-kingdom signal molecules in life modulating, for example microbial physiology, metabolism and virulence. Despite their increasing importance as global signal molecules, the mechanisms by which auxins perform their regulatory functions in microorganisms are largely unknown. In this article, we outline recent research that has advanced our knowledge of the mechanisms of bacterial auxin perception. We also highlight the potential applications of this research in aspects such as antibiotic production, biosensor design, plant microbiome engineering and antivirulence therapies.     

Media Ion Composition Controls Regulatory and Virulence Response of Salmonella in Spaceflight

The spaceflight environment is relevant to conditions encountered by pathogens during the course of infection and induces novel changes in microbial pathogenesis not observed using conventional methods. It is unclear how microbial cells sense spaceflight-associated changes to their growth environment and orchestrate corresponding changes in molecular and physiological phenotypes relevant to the infection process. Here we report that spaceflight-induced increases in Salmonella virulence are regulated by media ion composition, and that phosphate ion is sufficient to alter related pathogenesis responses in a spaceflight analogue model. Using whole genome microarray and proteomic analyses from two independent Space Shuttle missions, we identified evolutionarily conserved molecular pathways in Salmonella that respond to spaceflight under all media compositions tested. Identification of conserved regulatory paradigms opens new avenues to control microbial responses during the infection process and holds promise to provide an improved understanding of human health and disease on Earth.     

Biodiversity maintenance in food webs with regulatory environmental feedbacks

Although the food web is one of the most fundamental and oldest concepts in ecology, elucidating the strategies and structures by which natural communities of species persist remains a challenge to empirical and theoretical ecologists. We show that simple regulatory feedbacks between autotrophs and their environment when embedded within complex and realistic food-web models enhance biodiversity. The food webs are generated through the niche-model algorithm and coupled with predator-prey dynamics, with and without environmental feedbacks at the autotroph level. With high probability and especially at lower, more realistic connectance levels, regulatory environmental feedbacks result in fewer species extinctions, that is, in increased species persistence. These same feedback couplings, however, also sensitize food webs to environmental stresses leading to abrupt collapses in biodiversity with increased forcing. Feedback interactions between species and their material environments anchor food-web persistence, adding another dimension to biodiversity conservation. We suggest that the regulatory features of two natural systems, deep-sea tubeworms with their microbial consortia and a soil ecosystem manifesting adaptive homeostatic changes, can be embedded within niche-model food-web dynamics.     

Characterising microbial protein test substances and establishing their equivalence with plant-produced proteins for use in risk assessments of transgenic crops

Most commercial transgenic crops are genetically engineered to produce new proteins. Studies to assess the risks to human and animal health, and to the environment, from the use of these crops require grams of the transgenic proteins. It is often extremely difficult to produce sufficient purified transgenic protein from the crop. Nevertheless, ample protein of acceptable purity may be produced by over-expressing the protein in microbes such as Escherichia coli. When using microbial proteins in a study for risk assessment, it is essential that their suitability as surrogates for the plant-produced transgenic proteins is established; that is, the proteins are equivalent for the purposes of the study. Equivalence does not imply that the plant and microbial proteins are identical, but that the microbial protein is sufficiently similar biochemically and functionally to the plant protein such that studies using the microbial protein provide reliable information for risk assessment of the transgenic crop. Equivalence is a judgement based on a weight of evidence from comparisons of relevant properties of the microbial and plant proteins, including activity, molecular weight, amino acid sequence, glycosylation and immuno-reactivity. We describe a typical set of methods used to compare proteins in regulatory risk assessments for transgenic crops, and discuss how risk assessors may use comparisons of proteins to judge equivalence.     

Naturally arising CD25+CD4+ regulatory T cells in maintaining immunologic self-tolerance and preventing autoimmune disease

A large body of evidence indicates that T cell-mediated dominant suppression of self-reactive T cells is indispensable for maintaining immunologic unresponsiveness to self-constituents (i.e., self-tolerance) and preventing autoimmune disease. CD25+CD4+ regulatory T cells naturally present in normal animals, in particular, engage in this function, as their reduction or functional abnormality leads to the development of autoimmune disease in otherwise normal animals. They are at least in part produced by the normal thymus as a functionally mature and distinct subpopulation of T cells. Recent studies have demonstrated that CD25+CD4+ regulatory T cells control not only autoimmune reactions but also other immune responses, including tumor immunity, transplantation tolerance and microbial infection. Thus, this unique population of regulatory T cells can be exploited to control pathological as well as physiological immune responses.     

Regulating biocontrol agents: a historical perspective and a critical examination comparing microbial and macrobial agents

Ever since the inclusion of microbial biocontrol agents (MBCAs) within the regulatory frameworks initially designed for chemical pesticides, there has been awareness that these frameworks are not optimal for assessment and registration of new microbial biocontrol products. It is often claimed that the regulatory situation has contributed to a relatively slow uptake of microbial biocontrol in practice. In contrast to the MBCAs, non-indigenous invertebrate biocontrol agents (IBCAs) are regulated in many countries through quarantine and other biosecurity related legislation for prevention of introduction of alien organisms, whereas use of indigenous IBCAs are generally unregulated. In this study, we investigate what scientific support there is for performing evaluations of these two main groups of biocontrol agents (BCAs) within different frameworks. We compare potential risks of MBCAs and IBCAs, present a retrospective analysis of the development and implementation of the regulatory frameworks, and compare current requirements for MBCAs with those for other applications with microorganisms. One conclusion is that the ecological risks are of similar types between the two groups of BCAs, and that for both groups the environmental safety is most pertinently evaluated according to biological and ecological principles. The main difference between MBCAs and IBCAs with respect to human health is that the former may cause infectious disease. However, we found no evidence that this hazard is more serious for microorganisms for biocontrol than for microbes used in other types of applications, which generally have substantially lower regulatory demands than those for MBCAs. Several international initiatives have produced helpful guidelines and recommendations for simplified assessments and authorisations of BCAs. Still, we conclude that as long as MBCAs are evaluated within systems initially developed for chemicals, the risk for inappropriate emphasis of chemical hazards and therefore unnecessarily complicated assessments will be maintained. Therefore, this study supports the idea that development of new systems for the regulatory oversight of MBCAs, possibly a mutual framework covering all living BCAs, should be considered. Research issues that need to be further explored are to what extent utilisation of MBCAs actually results in increased exposure of non-targets to microorganisms, the biogeography and microbial ecology of representative MBCAs, and finally development of better methodology for determining potential human toxicity and pathogenicity of candidate MBCAs.     

Synthetic Biology Toolkits for Metabolic Engineering of Cyanobacteria

Cyanobacteria are of great importance to Earth's ecology. Due to their capability in photosynthesis and C1 metabolism, they are ideal microbial chassis that can be engineered for direct conversion of carbon dioxide and solar energy into biofuels and biochemicals. Facilitated by the elucidation of the basic biology of the photoautotrophic microbes and rapid advances in synthetic biology, genetic toolkits have been developed to enable implementation of nonnatural functionalities in engineered cyanobacteria. Hence, cyanobacteria are fast becoming an emerging platform in synthetic biology and metabolic engineering. Herein, the progress made in the synthetic biology toolkits for cyanobacteria and their utilization for transforming cyanobacteria into microbial cell factories for sustainable production of biofuels and biochemicals is outlined. Current techniques in heterologous gene expression, strategies in genome editing, and development of programmable regulatory parts and modules for engineering cyanobacteria towards biochemical production are discussed and prospected. As cyanobacteria synthetic biology is still in its infancy, apart from the achievements made, the difficulties and challenges in applying and developing genetic toolkits in cyanobacteria for biochemical production are also evaluated.     

基于微生物共培养的隐性基因簇激活策略

微生物次级代谢产物是药物先导化合物的重要源泉之一。随着测序技术的迅猛发展,越来越多的微生物基因组得以测序完成。伴随着测序技术的进步,生物信息学也得到了快速发展。基因组序列分析发现,链霉菌和丝状真菌等微生物中存在大量的已知的或未知的次级代谢物生物合成基因簇(secondary metabolite-biosynthetic gene clusters,SM-BGCs)。然而,在实验室培养条件下大部分基因簇无法表达或表达量很低,导致难以发现这些基因簇所对应的代谢产物,人们将这类基因簇称为“隐性基因簇”或“沉默基因簇”。通过调节基因簇中特异调控基因或基因簇外全局性调控基因的表达,对代谢途径的定向改造,以及将基因簇导入异源宿主等策略,能够激活部分隐性基因簇的表达。通过激活隐性基因簇的表达,能够发现通过常规实验室培养无法获得的具有独特生物活性的新结构代谢产物,成为创新药物的重要来源之一。然而,这些基因簇激活策略都严重依赖于对特定菌株或宿主的遗传操作。近年来,通过模拟自然混合培养中微生物间相互作用,开发了通过混合特定微生物菌株在厌氧或好氧条件下激活隐性基因簇的方法,称之为共培养激活策略。这种策略不...     

家蚕免疫稳态调控分子的鉴定和表达模式分析

昆虫免疫稳态的维持有赖于准确地激活和有效地抑制Toll或IMD信号通路中的关键转录因子-- Dorsal/Dif或Relish。在果蝇等昆虫中, 已报道了多种降低转录因子稳定性和活性的免疫稳态调控分子, 突变或敲除这类分子导致免疫系统的过度激活。对家蚕Bombyx mori免疫信号通路的研究中, 至今为止尚无对这类分子的探索。本研究通过比较基因组学, 在家蚕基因组中鉴定了多个可能参与免疫稳态调控的分子, 包括Wnt家族成员、 Ubc9、 FAF和POSH等; 并通过检测家蚕被微生物感染后这些分子在多种免疫器官中的诱导表达模式, 发现这些分子的表达水平在微生物感染后普遍呈下降趋势, 虽然在某些组织中表达量有明显的升高（>1.5倍）, 但此高表达水平均不能维持且迅速下降; 而且免疫稳态调控分子和受其调控的信号通路的对应关系在不同组织中表现出差异。本研究是首次对家蚕免疫稳态调控分子的报道, 为深入研究家蚕免疫负调控的分子机制提供了参考。