Classical medical research is disease focused and still defines health as absence of disease. Languages, however, associate a positive concept of wholeness with health as does the WHO health definition. Newer medical health definitions emphasize the capacity to adapt to changing external and internal circumstances. The results of the 2010 Global Burden of Disease study provides keys for a quantifiable health metrics by developing statistical tools calculating healthy life expectancy. Of central social and economic importance is the question whether healthy ageing can be achieved. This concept hinges on theories on the biological basis of lifespan determination and whether negligible senescence and the compression of morbidity can be achieved in human societies. Since the health impact of the human gut microbiome is currently a topical research area, microbiologists should be aware of the problems in defining health.

Radionuclides in the environment are a major human and environmental health concern. Like the Chernobyl disaster of 1986, the Fukushima Daiichi nuclear disaster in 2011 is once again causing damage to the environment: a large quantity of radioactive waste is being generated and dumped into the environment, and if the general population is exposed to it, may cause serious life-threatening disorders. Bioremediation has been viewed as the ecologically responsible alternative to environmentally destructive physical remediation. Microorganisms carry endogenous genetic, biochemical and physiological properties that make them ideal agents for pollutant remediation in soil and groundwater. Attempts have been made to develop native or genetically engineered (GE) microbes for the remediation of environmental contaminants including radionuclides. Microorganism-mediated bioremediation can affect the solubility, bioavailability and mobility of radionuclides. Therefore, we aim to unveil the microbial-mediated mechanisms for biotransformation of radionuclides under various environmental conditions as developing strategies for waste management of radionuclides. A discussion follows of ‘-omics’-integrated genomics and proteomics technologies, which can be used to trace the genes and proteins of interest in a given microorganism towards a cell-free bioremediation strategy.

Bioremediation has been viewed as the ecologically responsible alternative to environmentally destructive physical remediation. Microorganism-mediated bioremediation can affect the solubility, bioavailability, and mobility of radionuclides. This article aims to unveil the microbial-mediated mechanisms for biotransformation of radionuclides under various environmental conditions as developing strategies for waste management of radionuclides.

l-Histidine biosynthesis is an ancient metabolic pathway present in bacteria, archaea, lower eukaryotes, and plants. For decades l-histidine biosynthesis has been studied mainly in Escherichia coli and Salmonella typhimurium, revealing fundamental regulatory processes in bacteria. Furthermore, in the last 15 years this pathway has been also investigated intensively in the industrial amino acid-producing bacterium Corynebacterium glutamicum, revealing similarities to E. coli and S. typhimurium, as well as differences. This review summarizes the current knowledge of l-histidine biosynthesis in C. glutamicum. The genes involved and corresponding enzymes are described, in particular focusing on the imidazoleglycerol-phosphate synthase (HisFH) and the histidinol-phosphate phosphatase (HisN). The transcriptional organization of his genes in C. glutamicum is also reported, including the four histidine operons and their promoters. Knowledge of transcriptional regulation during stringent response and by histidine itself is summarized and a translational regulation mechanism is discussed, as well as clues about a histidine transport system. Finally, we discuss the potential of using this knowledge to create or improve C. glutamicum strains for the industrial l-histidine production.

This minireview gives the state-of-the-art on histidine biosynthesis, its regulation and biotechnological application in the industrially important amino acid producer Corynebacterium glutamicum.

Cultivable bacterial strains associated with field-grown Brassica napus L. (soil, rhizosphere and roots) from a trace elements (Cd, Zn and Pb) contaminated field and a non-contaminated control field were characterized genotypically and phenotypically. Correspondence analysis of the genotypic data revealed a correlation between soil and rhizosphere communities isolated from the same field, indicating that local conditions play a more important role in influencing the composition of (rhizosphere) soil bacterial communities than root exudates. In contrast, endophytic communities of roots showed a correlation between fields, suggesting that plants on the two fields contain similar obligate endophytes derived from a common seed endophytic community and/or can select bacteria from the rhizosphere. The latter seemed not very likely since, despite the presence of several potential endophytic taxa in the rhizosphere, no significant correlation was found between root and rhizosphere communities. The majority of Cd/Zn tolerant strains capable of phosphorus solubilization, nitrogen fixation, indole-3-acetic acid production and showing 1-aminocyclopropane-1-carboxylate deaminase capacity were found in the rhizosphere and roots of plants growing on the contaminated field.

Cultivable bacterial strains associated with field-grown Brassica napus L. (soil, rhizosphere and roots) from a trace elements (Cd, Zn and Pb) contaminated field and a non-contaminated control field were characterised genotypically and phenotypically. Local conditions play a more important role in influencing the composition of (rhizosphere) soil bacterial communities than root exudates. Endophytic communities of roots showed a correlation between fields, suggesting that plants on the two fields contain similar obligate endophytes derived from a common seed endophytic community.

Accumulating evidence demonstrates the intimate association between human hosts and the gut microbiome. Starting at birth, the sterile gut of the newborn acquires a diverse spectrum of microbes, needed for immunological priming. However, current practices (caesarean sections, use of formula milk) deprive newborns from being exposed to this broad spectrum of microbes. Unnecessary use of antibiotics and excessive hygienic precautions (e.g. natural versus chlorinated drinking water) together with the Western diet further contribute to a decreased microbial diversity in the adult gut. This has been correlated with recurrent Clostridium difficile infection, inflammatory bowel diseases and obesity, among others. A healthy gut microbiome is thus characterized by a diverse network of metabolically interacting microbial members. In this context, we review several existing and novel approaches to manage the gut microbiome. First, prebiotic compounds should be re-defined in the sense that they should enhance the ecological biodiversity rather than stimulating single species. Recent studies highlight that structurally different polysaccharides require specific primary degraders but also enhance a similar network of secondary degraders that benefit from cross-feeding. A faecal transplantation is a second approach to restore biodiversity when the microbiota is severely dysbiosed, with promising results regarding C. difficile-associated disease and obesity-related metabolic syndromes. A final strategy is the introduction of key microbial network units, i.e. pre-organized microbial associations, which strengthen the overall microbial network of the gut microbiome that supports human health.

Current society practices contribute to a decreased microbial diversity of the human gut microbiome, thus disturbing the intimate association between humans and the gut microbes. In this context, we review existing (prebiotic) and novel (fecal transplants, key microbial network units) approaches to manage the human gut microbiome.

The bacterial community composition of activated sludge from a wastewater treatment plant (Almería, Spain) with the particularity of using seawater was investigated by applying 454-pyrosequencing. The results showed that Deinococcus-Thermus, Proteobacteria, Chloroflexi and Bacteroidetes were the most abundant retrieved sequences, while other groups, such as Actinobacteria, Chlorobi, Deferribacteres, Firmicutes, Planctomycetes, Spirochaetes and Verrumicrobia were reported at lower proportions. Rarefaction analysis showed that very likely the diversity is higher than what could be described despite most of the unknown microorganisms probably correspond to rare diversity. Furthermore, the majority of taxa could not be classified at the genus level and likely represent novel members of these groups. Additionally, the nitrifiers in the sludge were characterized by pyrosequencing the amoA gene. In contrast, the nitrifying bacterial community, dominated by the genera Nitrosomonas, showed a low diversity and rarefaction curves exhibited saturation. These results suggest that only a few populations of low abundant but specialized bacteria are responsible for removal of ammonia in these saline wastewater systems.

The bacterial community composition of activated sludge from a seawater-processing WWTP (Almería, Spain) was investigated by 454-pyrosequencing. Members of Deinococcus-Thermus, Proteobacteria, Chloroflexi and Bacteroidetes were the most abundant, and rarefaction curves showed that diversity was higher than what could be described despite most of the unknown microorganisms probably correspond to rare diversity. Additionally, the nitrifying bacterial community was dominated by the genera Nitrosomonas.

While practised for over thousand years, there is presently a renaissance in the interest of using of faecal transplantations to modify the intestinal microbiota of patients. This clinical practice consists of delivering large amounts of bowel microbes in various forms into the intestinal tract of the recipient that usually has been cleared previously. The major reason for the popularity of faecal transplantations is their effectiveness in treating a variety of diseases. Hence, there is a need to develop this procedure to the next level. While there are various developments to select, standardize and store the donor microbiota, it is more challenging to understand the intestinal microbial communities and develop ways to deliver these via robust biotechnological processes. The various approaches that have been followed to do so are discussed in this contribution that is also addressing the concept of the minimal microbiome as well as the production of the synthetic communities that can be instrumental in new therapeutic avenues to modify the intestinal microbiota.

Composition of human fecal material. The distribution of dry weight constituents (left) and the differentiation of the microbiota into viable, dead and damaged cells (right). The latter piechart reveals that approximately half of the microbiota is dead or damaged in the fecal material.

A collection of marine bacteria isolated from a temperate coastal zone has been screened in a programme of biodiscovery. A total of 34 enzymes with biotechnological potential were screened in 374 isolates of marine bacteria. Only two enzymes were found in all isolates while the majority of enzyme activities were present in a smaller proportion of the isolates. A cluster analysis demonstrated no significant correlation between taxonomy and enzyme function. However, there was evidence of co-occurrence of some enzyme activity in the same isolate. In this study marine Proteobacteria had a higher complement of enzymes with biodiscovery potential than Actinobacteria; this contrasts with the terrestrial environment where the Actinobacteria phylum is a proven source of enzymes with important industrial applications. In addition, a number of novel enzyme functions were more abundant in this marine culture collection than would be expected on the basis of knowledge from terrestrial bacteria. There is a strong case for future investigation of marine bacteria as a source for biodiscovery.

Cluster analysis of the co-occurrence of enzymatic activities among 374 bacterial isolates from the marine environment. The 34 enzymes are divided into two main clusters (I, II), which closely resemble the definition of ‘core’ (cluster I) and ‘specialist’ (cluster II) enzymes. Enzymes not meeting this definition are indicated in italics and are underlined.

Hundred years ago Metchnikoff associated human health and particularly healthy ageing with a specific type of gut microbiota. Classical culture methods associated a decrease in bifidobacteria and an increase in enterobacteria with ageing. Modern molecular methods blurred this simple picture and documented a substantial inter-individual variability for the gut microbiome even when stratifying the elderly subjects according to health status. Nutritional interventions with resistant starch showed consistent gut microbiota changes across studies from different geographical areas and prebiotic supplementation induced a 10-fold increase in gut bifidobacteria. However, in the ELDERMET study, microbiota changes do not precede, but follow the changes in health status of elderly subjects possibly as a consequence of diet changes.

The ¹³C isotope tracer method was used to investigate the glucose metabolic flux distribution and regulation in Lactobacillus amylophilus to improve lactic acid production using kitchen waste saccharified solution (KWSS). The results demonstrate that L. amylophilus is a homofermentative bacterium. In synthetic medium, 60.6% of the glucose entered the Embden–Meyerhof–Parnas (EMP) to produce lactic acid, whereas 36.4% of the glucose entered the pentose phosphate metabolic pathway (HMP). After solid–liquid separation of the KWSS, the addition of Fe³⁺ during fermentation enhanced the NADPH production efficiency and increased the NADH content. The flux to the EMP was also effectively increased. Compared with the control (60.6% flux to EMP without Fe³⁺ addition), the flux to the EMP with the addition of Fe³⁺ (74.3%) increased by 23.8%. In the subsequent pyruvate metabolism, Fe³⁺ also increased lactate dehydrogenase activity, and inhibited alcohol dehydrogenase, pyruvate dehydrogenase and pyruvate carboxylase, thereby increasing the lactic acid production to 9.03 g l⁻¹, an increase of 8% compared with the control. All other organic acid by-products were lower than in the control. However, the addition of Zn²⁺ showed an opposite effect, decreasing the lactic acid production. In conclusion it is feasible and effective means using GC-MS, isotope experiment and MATLAB software to integrate research the metabolic flux distribution of lactic acid bacteria, and the results provide the theoretical foundation for similar metabolic flux distribution.

The 13C isotope tracer method was used to investigate the glucose metabolic flux distribution and regulation in Lactobacillus amylophilus to improve lactic acid production using kitchen waste saccharified solution (KWSS); The addition of Fe3+ during fermentation enhanced the NADPH production efficiency and increased the NADH content. The flux to the EMP was also effectively increased.

We have the technology and capability to develop an all-in-one microarray that can provide complete information on a microbial community, including algae, protozoa, bacteria, archaea, fungi, viruses, antimicrobial resistance, biotoxins and functional activity. With lab-on-a-chip, nanotechnology integrating a variety of the latest methods for a large number of sample types (water, sediment, waste water, food, blood, etc.) it is possible to make a desktop instrument that would have universal applications. There are two major thrusts to this grand challenge that will allow us to take advantage of the latest biotechnological breakthroughs in real time. The first is a bioengineering thrust that will take advantage of the large multidisciplinary laboratories in developing key technologies. Miniaturization will reduce reagent costs and increase sensitivity and reaction kinetics for rapid turnaround time. New and evolving technologies will allow us to port the designs for state-of-the-art microarrays today to completely new nanotechnology inspired platforms as they mature. The second thrust is in bioinformatics to use our existing expertise to take advantage of the rapidly evolving landscape of bioinformatics data. This increasing capacity of the data set will allow us to resolve microbial species to greatly improved levels and identify functional genes beyond the hypothetical protein level. A cheap and portable assay would impact countless areas, including clean water technologies, emerging diseases, bioenergy, infectious disease diagnosis, climate change, food safety, environmental clean-up and bioterrorism. In my opinion it is possible but it will require a very large group of multidiscplenary scientists from multiple institutions crossing many international boundaries and funding over a 5-year period of more than $100 million. Given the impact that this SuperChip could have it is well worth the price!!!

Environmental microbes oscillate between feast and famine and need to carefully manage utilization, storage and conversion of reserve products to exploitable sources of carbon and energy. Polyhydroxyalkanoates (PHAs) are storage polymers that serve bacteria as sources of food materials under physiological conditions of carbon demand. In order to obtain insights into the role of PHA depolymerase (PhaZ) and its relationship to a PHA polymerase (PhaC2) in the carbon management activity of Pseudomonas putida strain U, we created a polymerase hyperexpression strain and a depolymerase knockout mutant of this strain, and examined their synthesis of PHA and expression of their PHA genes. This study revealed that hyperexpression of PhaC2 led to the accumulation of higher amounts of PHA (44%wt) than in the wild-type strain (24%wt) after 24 h of cultivation, which then returned to wild-type levels by 48 h, as a result of elevated depolymerization. The phaZ mutant, however, accumulated higher levels of PHA than the parental strain (62%wt), which were maintained for at least 96 h. Transcriptional analysis of the pha cluster by RT-PCR revealed that PHA operon proteins, including depolymerase, are expressed from the beginning of the growth phase. Hyperexpression of the PhaC2 polymerase was accompanied by an increase in the expression of the PhaZ depolymerase and a decrease in expression of another PHA polymerase, PhaC1. This suggests tight regulatory coupling of PHA polymerase and depolymerase activities that act in synergy, and in concert with other PHA proteins, to provide dynamic PHA granule synthesis and remodelling that rapidly and sensitively respond to changes in availability of carbon and the physiological-metabolic needs of the cell, to ensure optimal carbon resource management.

In order to obtain insights into the role of polyhydroxyalkanoate (PHA) depolymerase (PhaZ) and its relationship to a PHA polymerase (PhaC2) in the carbon management activity of Pseudomonas putida strain U, PHA polymerase hyperexpression and PHA depolymerase knockout mutants were created and their phenotypes analyzed. Hyperexpression of PhaC2 led to accumulation of higher amounts of PHA (44 %wt) than in the wild type strain (24 %wt) after 24 h of cultivation, which then returned to wild-type levels by 48 h, as a result of elevated depolymerization. The phaZ mutant, however, accumulated higher levels of PHA than the parental strain (62 %wt), which were maintained for at least 96 h. Transcriptional analysis of the pha cluster by RT-PCR revealed regulatory coupling of PHA polymerase and depolymerase activities that act in synergy, and in concert with other PHA proteins, to provide dynamic PHA granule synthesis and remodeling that rapidly and sensitively respond to changes in availability of carbon and the physiological-metabolic needs of the cell, to ensure optimal carbon resource management.

The lantibiotic lacticin 3147 consists of two ribosomally synthesized and post-translationally modified antimicrobial peptides, Ltnα and Ltnβ, which act synergistically against a wide range of Gram-positive microorganisms. We performed saturation mutagenesis of specific residues of Ltnα to determine their functional importance. The results establish that Ltnα is more tolerant to change than previously suggested by alanine scanning mutagenesis. One substitution, LtnαH23S, was identified which improved the specific activity of lacticin 3147 against one pathogenic strain, Staphylococcus aureus NCDO1499. This represents the first occasion upon which the activity of a two peptide lantibiotic has been enhanced through bioengineering.

Saturation mutagenesis of specific residues of Ltnα established that it is more tolerant to change than previously suggested by alanine scanning mutagenesis. One substitution LtnαH23S, was identified which improved the specific activity of lacticin 3147 against one pathogenic strain, representing the first occasion upon which the activity of a two peptide lantibiotic has been enhanced through bioengineering.

In this study, perstractive fermentation of intracellular Monascus pigments in nonionic surfactant micelle aqueous solution had been studied. The permeability of cell membrane modified by nonionic surfactant might have influence on the rate of export of intracellular pigments into its extracellular broth while nearly no effect on the final extracellular pigment concentration. However, the solubilization of pigments in nonionic surfactant micelles strongly affected the final extracellular pigment concentration. The solubilization capacity of micelles depended on the kind of nonionic surfactant, the super-molecule assembly structure of nonionic surfactant in an aqueous solution, and the nonionic surfactant concentration. Elimination of pigment degradation by export of intracellular Monascus pigments and solubilizing them into nonionic surfactant micelles was also confirmed experimentally. Thus, nonionic surfactant micelle aqueous solution is potential for replacement of organic solvent for perstractive fermentation of intracellular product.

Cell membrane modified by nonionic surfactant increases the rate of export of intracellular pigments while has no effect on final extracellular pigment concentration. The presence of nonionic surfactant in the aqueous solution intensifies the export of intracellular pigments into its broth.

With the advent of high-resolution mass spectrometry together with sophisticated data analysis and interpretation algorithms, determination of protein synthesis and degradation rates (i.e. protein turnover) on a proteome-wide scale by employing stable isotope-labelled amino acids has become feasible. These dynamic data provide a deeper understanding of protein homeostasis and stress response mechanisms in microorganisms than well-established ‘steady state’ proteomics approaches. In this article, we summarize the technological challenges and solutions both on the biochemistry/mass spectrometry and bioinformatics level for turnover proteomics with a focus on chromatographic techniques. Although the number of available case studies for Corynebacterium glutamicum and related actinobacteria is still very limited, our review illustrates the potential of protein turnover studies for an improved understanding of questions in the area of biotechnology and biomedicine. Here, new insights from investigations of growth phase transition and different stress dynamics including iron, acid and heat stress for pathogenic but also for industrial actinobacteria are presented. Finally, we will comment on the advantages of integrated software solutions for biologists and briefly discuss the remaining technical challenges and upcoming possibilities for protein turnover analysis.

Proteome turnover in bacteria is reviewed for Corynebacterium glutamicum and related bacteria. The technical solutions are covered on the mass spectrometry and bioinformatics level. Case studies are presented that demonstrate the potential of this approach to understand bacterial physiology.

Competition between microbial species is a product of, yet can lead to a reduction in, the microbial diversity of specific habitats. Microbial habitats can resemble ecological battlefields where microbial cells struggle to dominate and/or annihilate each other and we explore the hypothesis that (like plant weeds) some microbes are genetically hard-wired to behave in a vigorous and ecologically aggressive manner. These ‘microbial weeds’ are able to dominate the communities that develop in fertile but uncolonized – or at least partially vacant – habitats via traits enabling them to out-grow competitors; robust tolerances to habitat-relevant stress parameters and highly efficient energy-generation systems; avoidance of or resistance to viral infection, predation and grazers; potent antimicrobial systems; and exceptional abilities to sequester and store resources. In addition, those associated with nutritionally complex habitats are extraordinarily versatile in their utilization of diverse substrates. Weed species typically deploy multiple types of antimicrobial including toxins; volatile organic compounds that act as either hydrophobic or highly chaotropic stressors; biosurfactants; organic acids; and moderately chaotropic solutes that are produced in bulk quantities (e.g. acetone, ethanol). Whereas ability to dominate communities is habitat-specific we suggest that some microbial species are archetypal weeds including generalists such as: Pichia anomala, Acinetobacter spp. and Pseudomonas putida; specialists such as Dunaliella salina, Saccharomyces cerevisiae, Lactobacillus spp. and other lactic acid bacteria; freshwater autotrophs Gonyostomum semen and Microcystis aeruginosa; obligate anaerobes such as Clostridium acetobutylicum; facultative pathogens such as Rhodotorula mucilaginosa, Pantoea ananatis and Pseudomonas aeruginosa; and other extremotolerant and extremophilic microbes such as Aspergillus spp., Salinibacter ruber and Haloquadratum walsbyi. Some microbes, such as Escherichia coli, Mycobacterium smegmatis and Pseudoxylaria spp., exhibit characteristics of both weed and non-weed species. We propose that the concept of nonweeds represents a ‘dustbin’ group that includes species such as Synodropsis spp., Polypaecilum pisce, Metschnikowia orientalis, Salmonella spp., and Caulobacter crescentus. We show that microbial weeds are conceptually distinct from plant weeds, microbial copiotrophs, r-strategists, and other ecophysiological groups of microorganism. Microbial weed species are unlikely to emerge from stationary-phase or other types of closed communities; it is open habitats that select for weed phenotypes. Specific characteristics that are common to diverse types of open habitat are identified, and implications of weed biology and open-habitat ecology are discussed in the context of further studies needed in the fields of environmental and applied microbiology.

This article explores the ecology, and cellular and molecular traits, of a select number of microbial species that consistently dominate their respective habitats. We suggest that these ‘microbial weeds’ have key features in common with plant weeds, and are conceptually distinct from other groupings of microorganisms such as generalists, specialists, copiotrophs, oligotrophs, r-strategists, K-strategists, etc. Microbial weed species are most likely to emerge in open habitats; we describe and give examples of such habitats and discuss environmental and applied implications of weed biology and open-habitat ecology as well as further work needed in these areas.

In a continuous culture under phosphate limitation the metabolism of Clostridium acetobutylicum depends on the external pH level. By comparing seven steady-state conditions between pH 5.7 and pH 4.5 we show that the switch from acidogenesis to solventogenesis occurs between pH 5.3 and pH 5.0 with an intermediate state at pH 5.1. Here, an integrative study is presented investigating how a changing external pH level affects the clostridial acetone–butanol–ethanol (ABE) fermentation pathway. This is of particular interest as the biotechnological production of n-butanol as biofuel has recently returned into the focus of industrial applications. One prerequisite is the furthering of the knowledge of the factors determining the solvent production and their integrative regulations. We have mathematically analysed the influence of pH-dependent specific enzyme activities of branch points of the metabolism on the product formation. This kinetic regulation was compared with transcriptomic regulation regarding gene transcription and the proteomic profile. Furthermore, both regulatory mechanisms were combined yielding a detailed projection of their individual and joint effects on the product formation. The resulting model represents an important platform for future developments of industrial butanol production based on C. acetobutylicum.

The industrial-relevant bacterium Clostridium acetobutylicum, grown in a phosphate-limited continuous culture, shifts its metabolism in response to changing pH levels. This results in a transition from the formation of the acids acetate and butyrate to the solvents acetone and butanol.

In this study, we investigate how pH-dependent kinetic and transcriptional regulatory mechanisms trigger this pH-induced metabolic switch.

We demonstrate that these two forms of regulation can either independently or jointly affect the observed metabolic phase transition.

Whereas previous studies have elucidated pH-induced changes on the transcriptomic and the proteomic level, our investigation shows that information about pH-dependent kinetic enzyme properties, in particular specific catalytic activities, is required for an improved understanding of the pH-induced alterations to the metabolic flux through the network.

To assess the applicability of latex cell coatings as an ‘off-the-shelf’ biocatalyst, the effect of osmoprotectants, temperature, humidity and O₂ on preservation of H₂ production in Rhodopseudomonas palustris coatings was evaluated. Immediately following latex coating coalescence (24 h) and for up to 2 weeks of dry storage, rehydrated coatings containing different osmoprotectants displayed similar rates of H₂ production. Beyond 2 weeks of storage, sorbitol-treated coatings lost all H₂ production activity, whereas considerable H₂ production was still detected in sucrose- and trehalose-stabilized coatings. The relative humidity level at which the coatings were stored had a significant impact on the recovery and subsequent rates of H₂ production. After 4 weeks storage under air at 60% humidity, coatings produced only trace amounts of H₂ (0–0.1% headspace accumulation), whereas those stored at < 5% humidity retained 27–53% of their H₂ production activity after 8 weeks of storage. When stored in argon at < 5% humidity and room temperature, R. palustris coatings retained full H₂ production activity for 3 months, implicating oxidative damage as a key factor limiting coating storage. Overall, the results demonstrate that biocatalytic latex coatings are an attractive cell immobilization platform for preservation of bioactivity in the dry state.

An important technical hurdle that must be addressed before biocatalytic latex coatings can be used as “off-the-shelf” catalysts for hydrogen production, or other applications, is that of maintaining bioactivity stability as a function of long-term storage. In this study, latex-embedded cells of R. palustris were stored in a dry state at room temperature for up to 3 months while maintaining their original hydrogen production activity. Successful preservation of cell activity required the addition of select osmotic stabilizers, i.e. sucrose or trehalose, to the coating mixture, low relative humidity (<5%) and anaerobic conditions during storage.

Escherichia coli-based bioreporters for arsenic detection are typically based on the natural feedback loop that controls ars operon transcription. Feedback loops are known to show a wide range linear response to the detriment of the overall amplification of the incoming signal. While being a favourable feature in controlling arsenic detoxification for the cell, a feedback loop is not necessarily the most optimal for obtaining highest sensitivity and response in a designed cellular reporter for arsenic detection. Here we systematically explore the effects of uncoupling the topology of arsenic sensing circuitry on the developed reporter signal as a function of arsenite concentration input. A model was developed to describe relative ArsR and GFP levels in feedback and uncoupled circuitry, which was used to explore new ArsR-based synthetic circuits. The expression of arsR was then placed under the control of a series of constitutive promoters, which differed in promoter strength, and which could be further modulated by TetR repression. Expression of the reporter gene was maintained under the ArsR-controlled P_ars promoter. ArsR expression in the systems was measured by using ArsR–mCherry fusion proteins. We find that stronger constitutive ArsR production decreases arsenite-dependent EGFP output from P_ars and vice versa. This leads to a tunable series of arsenite-dependent EGFP outputs in a variety of systematically characterized circuitries. The higher expression levels and sensitivities of the response curves in the uncoupled circuits may be useful for improving field-test assays using arsenic bioreporters.

Design of gene reporter circuits for assaying environmental toxicants is often based on an exploitation of naturally existing regulatory loops. Here we show how uncoupling the ArsR-controlled feedback loop benefits tunable reporter gene expression for arsenite bioreporters.

The current knowledge of trehalose biosynthesis under stress conditions is incomplete and needs further research. Since trehalose finds industrial and pharmaceutical applications, enhanced accumulation of trehalose in bacteria seems advantageous for commercial production. Moreover, physiological role of trehalose is a key to generate stress resistant bacteria by metabolic engineering. Although trehalose biosynthesis requires few metabolites and enzyme reactions, it appears to have a more complex metabolic regulation. Trehalose biosynthesis in bacteria is known through three pathways – OtsAB, TreYZ and TreS. The interconnections of in vivo synthesis of trehalose, glycogen or maltose were most interesting to investigate in recent years. Further, enzymes at different nodes (glucose-6-P, glucose-1-P and NDP-glucose) of metabolic pathways influence enhancement of trehalose accumulation. Most of the study of trehalose biosynthesis was explored in medically significant Mycobacterium, research model Escherichia coli, industrially applicable Corynebacterium and food and probiotic interest Propionibacterium freudenreichii. Therefore, the present review dealt with the trehalose metabolism in these bacteria. In addition, an effort was made to recognize how enzymes at different nodes of metabolic pathway can influence trehalose accumulation.

Trehalose is a non-reducing disaccharide which has several industrial and pharmaceutical applications. In addition, trehalose is accumulated as compatible solute in bacteria under stress conditions. Henceforth, a global picture of trehalose metabolism is prerequisite to develop novel processes for trehalose production and stress resistant bacterial strains (based on trehalose accumulation). Therefore, this review highlights how dissimilar bacteria have developed metabolic adaptation for enhanced trehalose synthesis.

A microbial fuel cell (MFC) was operated with a pure culture of Cupriavidus basilensis bacterial cells growing in the anode compartment in a defined medium containing acetate or phenol. Operating this mediator-less MFC under a constant external resistor of 1 kΩ with acetate or phenol led to current generation of 902 and 310 mA m⁻² respectively. In the MFC which was operated using acetate or phenol, the current density measured from the plankton bacterial cells with a fresh electrode was 125 and 109 mA m⁻², respectively, whereas the current obtained with biofilm-covered electrodes in sterile medium was 541 and 228 mA m⁻² respectively. After 72 h in the MFC, 86% of the initial phenol concentration was removed, while only 64% was removed after the same time in the control MFC which was held at an open circuit potential (OCP). Furthermore, SEM and confocal microscopy analyses demonstrated a developed biofilm with a live C. basilensis population. In conclusion, in this study we demonstrated, for the first time, use of C. basilensis facultative aerobe bacterial cells in a MFC using acetate or phenol as the sole carbon source which led to electricity generation.

A microbial fuel cell (MFC) was operated with a pure culture of Cupriavidus basilensis bacterial cells growing in the anode compartment in a defined medium containing acetate or phenol. Operating this mediator-less MFC under a constant external resistor of 1 kΩ with acetate or phenol led to current generation of 902 and 310 mA m^-2, respectively. After 72 h in the MFC, 86% of the initial phenol concentration was removed, while only 64% was removed after the same time in the control MFC which was held at an open circuit potential (OCP).

Anaerobic digestion is an environmental key technology in the future bio-based economy. To achieve functional stability, a minimal microbial community diversity is required. This microbial community should also have a certain ‘elasticity’, i.e. the ability to rapidly adapt to suboptimal conditions or stress. In this study it was evaluated whether a higher degree of functional stability could be achieved by changing the feeding pattern, which can change the evenness, dynamics and richness of the bacterial community. The first reactor (CSTR_stable) was fed on daily basis, whereas the second reactor (CSTR_dynamic) was fed every 2 days. Average biogas production was 0.30 l CH₄ l⁻¹ day⁻¹ in both reactors, although daily variation was up to four times higher in the CSTR_dynamic compared with the CSTR_stable during the first 50 days. Bacterial analysis revealed that this CSTR_dynamic had a two times higher degree of bacterial community dynamics. The CSTR_dynamic also appeared to be more tolerant to an organic shock load of 8 g COD l⁻¹ and ammonium levels up to 8000 mg TAN l⁻¹. These results suggest that the regular application of a limited pulse of organic material and/or a variation in the substrate composition might promote higher functional stability in anaerobic digestion.

The application of a pulse feeding pattern in anaerobic digestion leads to a higher degree of operational stability, i.e. a higher tolerance to stress. The higher stability was correlated to a very dynamic bacterial community and stable methanogenic community. These results call for the regular application of a limited pulse of organic material and/or a variation in the substrate to obtain a higher degree of functional stability in anaerobic digestion.

Polystyrene (PS) nanoparticle (NP) copolymerized with acrylic acid (AA) and coloured monomer, i.e. 2,3,6,7-tetra(2,2′-bithiophene)-1,4,5,8-naphthalenetetracarboxylic-N,N′-di(2-methylallyl)-bisimide (ALN8T), was synthesized via the miniemulsion polymerization. Before applying for malaria antigen detection, the blue NP was conjugated with human polyclonal malaria IgG antibody (Ab) specific to Plasmodium falciparum. For the conjugation, three methods, i.e. physical adsorption, covalent coupling and affinity binding via streptavidin (SA) and biotin interaction, were employed. The optimum ratio of Ab to NPs used in each immobilization procedure and the latex agglutination test based on the reaction between Ab conjugated NPs and malaria patient plasma were investigated. All Ab–latex conjugates provided the high sensitivity for the detection of P. falciparum malaria plasma. The highest specificity to P. falciparum was obtained from using Ab–NPs conjugated via the SA–biotin interaction.

Polystyrene (PS) nanoparticle copolymerized with acrylic acid (AA) and colored monomer (ALN8T) (PS/AA-ALN8T) was synthesized and used as a solid support of antibody specific to Plasmodium falciparum for malaria detection. Antibody immobilization was performed by three processes to compare the sensitivity and specificity. High sensitivity of 100% was obtained in all kinds of antibody coated PS/AA-ALN8T nanoparticles when tested with P. falciparum, while the antibody coated nanoparticles prepared from streptavidin-biotin interaction exhibited the highest specificity to P. falciparum.

Biosurfactants are produced by hydrocarbon-degrading marine bacteria in response to the presence of water-insoluble hydrocarbons. This is believed to facilitate the uptake of hydrocarbons by bacteria. However, these diffusible amphiphilic surface-active molecules are involved in several other biological functions such as microbial competition and intra- or inter-species communication. We report the isolation and characterization of a marine bacterial strain identified as Cobetia sp. MM1IDA2H-1, which can grow using the sulfur-containing heterocyclic aromatic hydrocarbon dibenzothiophene (DBT). As with DBT, when the isolated strain is grown in the presence of a microbial competitor, it produces a biosurfactant. Because the obtained biosurfactant was formed by hydroxy fatty acids and extracellular lipidic structures were observed during bacterial growth, we investigated whether the biosurfactant at its critical micelle concentration can interfere with bacterial communication systems such as quorum sensing. We focused on Aeromonas salmonicida subsp. salmonicida, a fish pathogen whose virulence relies on quorum sensing signals. Using biosensors for quorum sensing based on Chromobacterium violaceum and Vibrio anguillarum, we showed that when the purified biosurfactant was mixed with N-acyl homoserine lactones produced by A. salmonicida, quorum sensing was inhibited, although bacterial growth was not affected. In addition, the transcriptional activities of A. salmonicida virulence genes that are controlled by quorum sensing were repressed by both the purified biosurfactant and the growth in the presence of Cobetia sp. MM1IDA2H-1. We propose that the biosurfactant, or the lipid structures interact with the N-acyl homoserine lactones, inhibiting their function. This could be used as a strategy to interfere with the quorum sensing systems of bacterial fish pathogens, which represents an attractive alternative to classical antimicrobial therapies in fish aquaculture.

The marine bacterial strain identified as Cobetia sp. MM1IDA2H-1 when grown both, using the sulphur-containing heterocyclic aromatic hydrocarbon dibenzothiophene (DBT) or in the presence of Aeromonas salmonicida subsp. salmonicida, produces a biosurfactant that plays a role in microbial competition. At the critical micelle concentration, the biosurfactant forms lipid structures that interacts with N-Acyl homoserine lactones, inhibiting their function in quorum sensing of bacterial fish pathogens. The finding represents an attractive alternative to classical antimicrobial therapies in fish aquaculture.

The growth kinetics and fermentation performance of Williopsis saturnus and Saccharomyces cerevisiae at ratios of 10:1, 1:1 and 1:10 (W.:S.) were studied in papaya juice with initial 7-day fermentation by W. saturnus, followed by S. cerevisiae. The growth kinetics of W. saturnus were similar at all ratios, but its maximum cell count decreased as the proportion of S. cerevisiae was increased. Conversely, there was an early death of S. cerevisiae at the ratio of 10:1. Williopsis saturnus was the dominant yeast at 10:1 ratio that produced papaya wine with elevated concentrations of acetate esters. On the other hand, 1:1 and 1:10 ratios allowed the coexistence of both yeasts which enabled the flavour-enhancing potential of W. saturnus as well as the ethyl ester and alcohol-producing abilities of S. cerevisiae. In particular, 1:1 and 1:10 ratios resulted in production of more ethyl esters, alcohols and 2-phenylethyl acetate. However, the persistence of both yeasts at 1:1 and 1:10 ratios led to formation of high levels of acetic acid. The findings suggest that yeast ratio is a critical factor for sequential fermentation of papaya wine by W. saturnus and S. cerevisiae as a strategy to modulate papaya wine flavour.

Growth kinetics and fermentation performance of Williopsis saturnus and Saccharomyces cerevisiae at ratios of 10:1, 1:1 and 1:10 (W.:S.) were evaluated in papaya juice with initial fermentation by W. saturnus, followed by S. cerevisiae. The 1:1 and 1:10 ratios enabled co-existence of both yeasts and enhanced production of desirable volatile compounds.

With expansion of our understanding of pathogen effector strategies and the multiplicity of their host targets, it is becoming evident that novel approaches to engineering broad-spectrum resistance need to be deployed. The increasing availability of high temporal gene expression data of a range of plant–microbe interactions enables the judicious choices of promoters to fine-tune timing and magnitude of expression under specified stress conditions. We can therefore contemplate engineering a range of transgenic lines designed to interfere with pathogen virulence strategies that target plant hormone signalling or deploy specific disease resistance genes. An advantage of such an approach is that hormonal signalling is generic so if this strategy is effective, it can be easily implemented in a range of crop species. Additionally, multiple re-wired lines can be crossed to develop more effective responses to pathogens.

Increasing numbers of infectious crop diseases that are caused by fungi and oomycetes urge the need to develop alternative strategies for resistance breeding. As an alternative for the use of resistance (R) genes, the application of mutant susceptibility (S) genes has been proposed as a potentially more durable type of resistance. Identification of S genes is hampered by their recessive nature. Here we explore the use of pathogen-derived effectors as molecular probes to identify S genes. Effectors manipulate specific host processes thereby contributing to disease. Effector targets might therefore represent S genes. Indeed, the Pseudomonas syringae effector HopZ2 was found to target MLO2, an Arabidopsis thaliana homologue of the barley S gene Mlo. Unfortunately, most effector targets identified so far are not applicable as S genes due to detrimental effects they have on other traits. However, some effector targets such as Mlo are successfully used, and with the increase in numbers of effector targets being identified, the numbers of S genes that can be used in resistance breeding will rise as well.

Bacterial phytopathogens utilize a myriad of virulence factors to modulate their plant hosts in order to promote successful pathogenesis. One potent virulence strategy is to inject these virulence proteins into plant cells via the type III secretion system. Characterizing the host targets and the molecular mechanisms of type III secreted proteins, known as effectors, has illuminated our understanding of eukaryotic cell biology. As a result, these effectors can serve as molecular probes to aid in our understanding of plant cellular processes, such as immune signalling, vesicle trafficking, cytoskeleton stability and transcriptional regulation. Furthermore, given that effectors directly and specifically interact with their targets within plant cells, these virulence proteins have enormous biotechnological potential for manipulating eukaryotic systems.

Bacterial phytopathogens utilize a myriad of virulence factors to modulate their plant hosts in order to promote successful pathogenesis. These virulence proteins represent invaluable tools to effectively manipulate eukaryotic systems for pure and applied research.

Mushrooms are able to secrete lignin peroxidase (LiP) and manganese peroxidase (MnP), and able to use the cellulose as sources of carbon. This article focuses on the relation between peroxidase-secreting capacity and cultivation period of mushrooms with non-laccase activity. Methylene blue and methyl catechol qualitative assay and spectrophotometry quantitative assay show LiP secreting unvaryingly accompanies the MnP secreting in mushroom strains. The growth rates of hyphae are detected by detecting the dry hyphal mass. We link the peroxidase activities to growth rate of mushrooms and then probe into the relationship between them. The results show that there are close relationships between LiP- and/or MnP-secretory capacities and the cultivation periods of mushrooms. The strains with high LiP and MnP activities have short cultivation periods. However, those strains have long cultivation periods because of the low levels of secreted LiP and/or MnP, even no detectable LiP and/or MnP activity. This study provides the first evidence on the imitate relation between the level of secreted LiP and MnP activities and cultivation periods of mushrooms with non-laccase activity. Our study has significantly increased the understanding of the role of LiP and MnP in the growth and development of mushrooms with non-laccase activity.

This article focuses on the relation between peroxidase-secreting capacity and cultivation period of mushrooms with non-laccase activity. The strains with high LiP and MnP activities have short cultivation periods. However, those strains have long cultivation periods because of the low levels of secreted LiP and/or MnP, even no detectable LiP and/or MnP activity.

Fungal degradation of wood is mainly restricted to basidiomycetes, these organisms having developed complex oxidative and hydrolytic enzymatic systems. Besides these systems, wood-decaying fungi possess intracellular networks allowing them to deal with the myriad of potential toxic compounds resulting at least in part from wood degradation but also more generally from recalcitrant organic matter degradation. The members of the detoxification pathways constitute the xenome. Generally, they belong to multigenic families such as the cytochrome P450 monooxygenases and the glutathione transferases. Taking advantage of the recent release of numerous genomes of basidiomycetes, we show here that these multigenic families are extended and functionally related in wood-decaying fungi. Furthermore, we postulate that these rapidly evolving multigenic families could reflect the adaptation of these fungi to the diversity of their substrate and provide keys to understand their ecology. This is of particular importance for white biotechnology, this xenome being a putative target for improving degradation properties of these fungi in biomass valorization purposes.

Cytochrome P450 monooxygenases and glutathione transferases are part of the xenome, responsible for degradation of complex molecules. In wood decaying fungi, these multigenic families are functionally related and evolve rapidly both at the genomic and functional level, reflecting adaptation of fungi to the substrate diversity and provide keys to understand their ecology.

Biological control of plant diseases has gained acceptance in recent years. Bacillus subtilis UMAF6639 is an antagonistic strain specifically selected for the efficient control of the cucurbit powdery mildew fungus Podosphaera fusca, which is a major threat to cucurbits worldwide. The antagonistic activity relies on the production of the antifungal compounds iturin and fengycin. In a previous study, we found that UMAF6639 was able to induce systemic resistance (ISR) in melon and provide additional protection against powdery mildew. In the present work, we further investigated in detail this second mechanism of biocontrol by UMAF6639. First, we examined the signalling pathways elicited by UMAF6639 in melon plants, as well as the defence mechanisms activated in response to P. fusca. Second, we analysed the role of the lipopeptides produced by UMAF6639 as potential determinants for ISR activation. Our results demonstrated that UMAF6639 confers protection against cucurbit powdery mildew by activation of jasmonate- and salicylic acid-dependent defence responses, which include the production of reactive oxygen species and cell wall reinforcement. We also showed that surfactin lipopeptide is a major determinant for stimulation of the immune response. These results reinforce the biotechnological potential of UMAF6639 as a biological control agent.

In the present work we investigated in detail the mechanisms by which the antagonistic strain Bacillus subtilis UMAF6639 is also able to induce systemic resistance in melon and provide protection against powdery mildew. Our results demonstrated that UMAF6639 combats disease by activation of jasmonate- and salicylic acid-dependent defence responses, which include the production of reactive oxygen species and cell wall reinforcement. We also showed that surfactin lipopeptide is a major determinant for stimulation of the plant immune response. These results reinforce the biotechnological potential of UMAF6639 as a biological control agent.

Olive knot disease, caused by Pseudomonas savastanoi pv. savastanoi, is one of the most important biotic constraints for olive cultivation. Pseudomonas fluorescens PICF7, a natural colonizer of olive roots and effective biological control agent (BCA) against Verticillium wilt of olive, was examined as potential BCA against olive knot disease. Bioassays using in vitro-propagated olive plants were carried out to assess whether strain PICF7 controlled knot development either when co-inoculated with the pathogen in stems or when the BCA (in roots) and the pathogen (in stems) were spatially separated. Results showed that PICF7 was able to establish and persist in stem tissues upon artificial inoculation. While PICF7 was not able to suppress disease development, its presence transiently decreased pathogen population size, produced less necrotic tumours, and sharply altered the localization of the pathogen in the hyperplasic tissue, which may pose epidemiological consequences. Confocal laser scanning microscopy combined with fluorescent tagging of bacteria revealed that when PICF7 was absent the pathogen tended to be localized at the knot surface. However, presence of the BCA seemed to confine P. savastanoi at inner regions of the tumours. This approach has also enabled to prove that the pathogen can moved systemically beyond the hypertrophied tissue.

Pseudomonas fluorescens PICF7, a natural colonizer of olive roots and effective biological control agent (BCA) against Verticillium wilt of olive, was examined as potential BCA against olive knot disease caused by Pseudomonas savastanoi pv. savastanoi. While PICF7 was not able to suppress disease development, its presence transiently decreased pathogen population size, produced less necrotic tumors, and sharply altered the localization of the pathogen in the hyperplasic tissue. Confocal laser scanning microscopy revealed that when PICF7 was absent the pathogen tended to be localized at the knot surface. However, presence of the BCA seemed to confine P. savastanoi at inner regions of the tumors. This approach has also enabled to prove that the pathogen can moved systemically beyond the hypertrophied tissue.

Phytoextraction has been reported as an economically and ecologically sound alternative for the remediation of metal-contaminated soils. Willow is a metal phytoextractor of interest because it allows to combine a gradual contaminant removal with production of biomass that can be valorized in different ways. In this work two willow clones growing on a metal-contaminated site were selected: ‘Belgisch Rood’ (BR) with a moderate metal extraction capacity and ‘Tora’ (TO) with a twice as high metal accumulation. All cultivable bacteria associated with both willow clones were isolated and identified using 16SrDNA ARDRA analysis followed by 16SrDNA sequencing. Further all isolated bacteria were investigated for characteristics that might promote plant growth (production of siderophores, organic acids and indol acetic acid) and for their metal resistance. The genotypic and phenotypic characterization of the isolated bacteria showed that the TO endophytic bacterial population is more diverse and contains a higher percentage of metal-resistant plant growth promoting bacteria than the endophytic population associated with BR. We hypothesize that the difference in the metal accumulation capacity between BR and TO clones might be at least partly related to differences in characteristics of their associated bacterial population.

In this work, the bacterial population associated with the willow clone ‘Belgisch Rood’ (BR), with a moderate metal extraction capacity, was compared with the bacteria associated with ‘Tora’ (TO), a willow clone with a twice as high metal accumulation. The genotypic and phenotypic bacterial characterization showed that the TO-associated endophytes are more diverse and contain a higher percentage of metal-resistant plant growth promoting bacteria than the endophytic population associated with BR.

Fungal, ligninolytic enzymes have attracted a great attention for their bioremediation capabilities. A deficient knowledge of regulation of enzyme production, however, hinders the use of ligninolytic fungi in bioremediation applications. In this work, a transcriptional analyses of laccase and manganese peroxidase (MnP) production by two white rots was combined with determination of pI of the enzymes and the evaluation of 17α-ethinyloestradiol (EE2) degradation to study regulation mechanisms used by fungi during EE2 degradation. In the cultures of Trametes versicolor the addition of EE2 caused an increase in laccase activity with a maximum of 34.2 ± 6.7 U g⁻¹ of dry mycelia that was observed after 2 days of cultivation. It corresponded to a 4.9 times higher transcription levels of a laccase-encoding gene (lacB) that were detected in the cultures at the same time. Simultaneously, pI values of the fungal laccases were altered in response to the EE2 treatment. Like T. versicolor, Irpex lacteus was also able to remove 10 mg l⁻¹ EE2 within 3 days of cultivation. While an increase to I. lacteus MnP activity and MnP gene transcription levels was observed at the later phase of the cultivation. It suggests another metabolic role of MnP but EE2 degradation.

A deficient knowledge of regulation of enzyme production hinders the use of ligninolytic fungi in bioremediation applications. In the cultures of Trametes versicolor the addition of 17α- ethinylestradiol (EE2) caused an increase in laccase activity with a maximum of 34.2 ± 6.7 Ug−¹ of dry mycelia that corresponded to a 4.9 times higher transcription levels of a laccase encoding gene lacB. Simultaneously, pI values of the fungal laccases were altered in response to the EE2 treatment. While an increase to MnP activity and MnP gene transcription levels was observed in Irpex lacteus first after the removal of EE2 from the cultures. It suggests another metabolic role of MnP but EE2 degradation.

Pseudomonas putida KT2440 has the ability to colonize the rhizosphere of a wide range of plants and can reach cell densities in the range of 10⁵–10⁶ cfu g soil⁻¹. Using the IVET technology we investigated which KT2440 genes were expressed in the rhizosphere of four different plants: pine, cypress, evergreen oak and rosemary. We identified 39 different transcriptional fusions containing the promoters of annotated genes that were preferentially expressed in the rhizosphere. Six of them were expressed in the rhizosphere of all the plant types tested, 11 were expressed in more than one plant and the remaining 22 fusions were found to be expressed in only one type of plant. Another 40 fusions were found to correspond to likely promoters that encode antisense RNAs of unknown function, some of which were isolated as fusions from the bacteria recovered in the rhizosphere from all of the plants, while others were specific to one or several of the plants. The results obtained in this study suggest that plant-specific signals are sensed by KT2440 in the rhizosphere and that the signals and consequent gene expression are related to the bacteria's successful establishment in this niche.

Pseudomonas putida KT2440 efficiently colonizes the rhizosphere of a wide range of plants; genes specifically activated in the rhizosphere of four different plants (pine, cypress, evergreen oak and rosemary) were investigated using the IVET technology. The results revealed that Pseudomonas responds specifically to plant signals through the activation of common genes and some specific ones for a given plant. This seems to be undoubtedly part of the ability of KT2440 success in the colonization of the rhizosphere of different plants.