Osmoadaptation mechanisms in prokaryotes: distribution of compatible solutes

Microorganisms respond to osmotic stress mostly by accumulating compatible solutes, either by uptake from the medium or by de novo synthesis. These osmotically activ molecules preserve the positive turgor pressure required for cell division. The diversity of compatible solutes is large but falls into a few major chemical categories; they are usually small organic molecules such as amino acids or their derivatives, and carbohydrates and their derivatives. Some are widely distributed in nature while others seem to be exclusively present in specific groups of organisms. This review discusses the diversity and distribution of known classes of compatible solutes found in prokaryotes as well as the increasing knowledge of the genes and pathways involved in their synthesis. The alternative roles of some archetypal compatible solutes not subject to osmoregulatory constraints are also discussed

Diversity, distribution and biosynthesis of compatible solutes in prokaryotes

Els microorganismes responen a l'estrès osmòtic acumulant, principalment, soluts compatibles, agafant-los del medi o per síntesi de novo. Aquestes molècules, osmòticament actives, conserven la pressió de turgència positiva necessària per a la divisió cel·lular. La diversitat de soluts compatibles és gran, però es divideix en unes quantes categories químiques essencials; són molècules orgàniques normalment petites, com els aminoàcids, o els seus derivats, i els carbohidrats, o els seus derivats. Alguns són àmpliament distribuïts en la natura, mentre que d'altres sembla que siguin presents exclusivament en grups específics d'organismes. Aquesta revisió parla de la diversitat i la distribució de classes conegudes de soluts compatibles trobats en procariotes, així com del coneixement creixent dels gens i les rutes que s'impliquen en la seva síntesi. També es parla de les rutes alternatives d'alguns soluts compatibles no subjectes a restriccions osmoregulatòries.Microorganisms respond to osmotic stress mostly by accumulating compatible solutes, either by uptake from the medium or by de novo synthesis. These osmotically active molecules preserve the positive turgor pressure required for cell division. The diversity of compatible solutes is large but falls into a few major chemical categories; they are usually small organic molecules such as amino acids or their derivatives, and carbohydrates or their derivatives. Some are widely distributed in nature while others seem to be exclusively present in specific groups of organisms. This review discusses the diversity and distribution of known classes of compatible solutes found in prokaryotes as well as the increasing knowledge of the genes and pathways involved in their synthesis. The alternative roles of some archetypal compatible solutes not subject to osmoregulatory constraints are also discussed

Diversity and biosynthesis of compatible solutes in hyper/thermophiles

The accumulation of compatible solutes, either by uptake from the medium or by de novo synthesis, is a general response of microorganisms to osmotic stress. The diversity of compatible solutes is large but falls into a few major chemical categories, such as carbohydrates or their derivatives and amino acids or their derivatives. This review deals with compatible solutes found in thermophilic or hyperthermophilic bacteria and archaea that have not been commonly identified in microorganisms growing at low and moderate temperatures. The response to NaCl stress of Thermus thermophilus is an example of how a thermophilic bacterium responds to osmotic stress by compatible solute accumulation. Emphasis is made on the pathways leading to the synthesis of mannosylglycerate and glucosylglycerate that have been recently elucidated in several hyper/thermophilic microorganisms. The role of compatible solutes in the thermoprotection of these fascinating microorganisms is also discussed. [Int Microbiol 2006; 9(3):199-206

Autophagy in the fight against tuberculosis.

Tuberculosis (TB), a chronic infectious disease mainly caused by the tubercle bacillus Mycobacterium tuberculosis, is one of the world's deadliest diseases that has afflicted humanity since ancient times. Although the number of people falling ill with TB each year is declining, its incidence in many developing countries is still a major cause of concern. Upon invading host cells by phagocytosis, M. tuberculosis can replicate within infected cells by arresting the maturation of the phagosome whose function is to target the pathogen for elimination. Host cells have mechanisms of controlling this evasion by inducing autophagy, an elaborate cellular process that targets bacteria for progressive elimination, decreasing bacterial loads within infected cells. In addition, autophagy activation also aids in the control of inflammation, contributing to a more efficient innate immune response against M. tuberculosis. Several innovative TB therapies have been envisaged based on autophagy manipulation, with some of them revealing high potential for future clinical trials and eventual implementation in healthcare systems. Thus, this review highlights the recent advances on the innate immune response regulation by autophagy upon M. tuberculosis infection and the promising new autophagy-based therapies for TB.This work was funded by Bill & Melinda Gates Foundation (subcontract on the production of high quality chemical hit series with defined, tractable targets as drug leads for tuberculosis grant awarded to the Foundation for the National Institutes of Health) (OPP1024021), Fundação para a Ciência e a Tecnologia and EU-FEDER-COMPETE for funding (FCOMP-01-0124-FEDER-028359; PTDC/BIAMIC/2779/2012). VM would like to acknowledge Fundação para a Ciência e a Tecnologia for a postdoctoral fellowship (SFRH/BPD/79531/2011).This is the accepted manuscript. The final published version is available from Mary Ann Liebert, Inc., publishers at http://dx.doi.org/10.1089/dna.2014.2745

Structure of Mycobacterium thermoresistibile GlgE defines novel conformational states that contribute to the catalytic mechanism.

GlgE, an enzyme of the pathway that converts trehalose to α-glucans, is essential for Mycobacterium tuberculosis. Inhibition of GlgE, which transfers maltose from a maltose-1-phosphate donor to α-glucan/maltooligosaccharide chain acceptor, leads to a toxic accumulation of maltose-1-phosphate that culminates in cellular death. Here we describe the first high-resolution mycobacterial GlgE structure from Mycobacterium thermoresistibile at 1.96 Å. We show that the structure resembles that of M. tuberculosis and Streptomyces coelicolor GlgEs, reported before, with each protomer in the homodimer comprising five domains. However, in M. thermoresistibile GlgE we observe several conformational states of the S domain and provide evidence that its high flexibility is important for enzyme activity. The structures here reported shed further light on the interactions between the N-terminal domains and the catalytic domains of opposing chains and how they contribute to the catalytic reaction. Importantly this work identifies a useful surrogate system to aid the development of GlgE inhibitors against opportunistic and pathogenic mycobacteria.This is the final version of the article. It was first available from NPG via http://dx.doi.org/10.1038/srep1714

Biochemical characterization of the maltokinase from Mycobacterium bovis BCG

<p>Abstract</p> <p>Background</p> <p>Maltose-1-phosphate was detected in <it>Mycobacterium bovis </it>BCG extracts in the 1960's but a maltose-1-phosphate synthetase (maltokinase, Mak) was only much later purified from <it>Actinoplanes missouriensis</it>, allowing the identification of the <it>mak </it>gene. Recently, this metabolite was proposed to be the intermediate in a pathway linking trehalose with the synthesis of glycogen in <it>M. smegmatis</it>. Although the <it>M. tuberculosis </it>H37Rv <it>mak </it>gene (Rv0127) was considered essential for growth, no mycobacterial Mak has, to date, been characterized.</p> <p>Results</p> <p>The sequence of the Mak from <it>M. bovis </it>BCG was identical to that from <it>M. tuberculosis </it>strains (99-100% amino acid identity). The enzyme was dependent on maltose and ATP, although GTP and UTP could be used to produce maltose-1-phosphate, which we identified by TLC and characterized by NMR. The K_{<it>m </it>}for maltose was 2.52 ± 0.40 mM and 0.74 ± 0.12 mM for ATP; the <it>V</it>_maxwas 21.05 ± 0.89 μmol/min.mg^-1. Divalent cations were required for activity and Mg²⁺was the best activator. The enzyme was a monomer in solution, had maximal activity at 60°C, between pH 7 and 9 (at 37°C) and was unstable on ice and upon freeze/thawing. The addition of 50 mM NaCl markedly enhanced Mak stability.</p> <p>Conclusions</p> <p>The unknown role of maltokinases in mycobacterial metabolism and the lack of biochemical data led us to express the <it>mak </it>gene from <it>M. bovis </it>BCG for biochemical characterization. This is the first mycobacterial Mak to be characterized and its properties represent essential knowledge towards deeper understanding of mycobacterial physiology. Since Mak may be a potential drug target in <it>M. tuberculosis</it>, its high-level production and purification in bioactive form provide important tools for further functional and structural studies.</p

Sustainable starch-based edible films with agrifood residues as potential carriers for the probiotic Lactobacillus rhamnosus

Acknowledgments This work was supported by “Santander Universidades and University of Coimbra” through the project “Projetos Semente Santander UC: Era Pos-COVID-19: ´ uma sociedade mais sustentavel, ´ resiliente e justa” and by Fundaçao ˜ para a Ciˆencia e a Tecnologia (FCT): FCT/MCTEs (UIDB/00102/2020 and UIDP/00102/2020, of CIEPQPF, UIDB/04539/2020, UIDP/04539/2020 and LA/P/0058/2020, of CIBB, and UI/ 05704/2020 of ciTechCare. M. C. Gaspar acknowledges FCT for the financial support through the Institutional Scientific Employment Stimulus (CEECINST/00060/2021).Edible films are promising carriers for probiotics and can be composed by agrifood residues, which are usually rich in polymers and bioactive compounds. In this work, starch-based films were enriched with three types of agrifood residues (quince, potato and orange peels) and the incorporation of the probiotic Lactobacillus rhamnosus was studied, as well as the addition of inulin as a protective prebiotic. The resulting films were characterized in terms of mechanical properties, physicochemical properties, lactobacilli viability and microbiological properties. The mechanical properties of the films generally decreased with the introduction of L. rhamnosus, although this was highly dependent on the film composition. All films exhibited water vapor permeabilities in the typical range of starch-based films and were not greatly affected by the inclusion of probiotics. The loss of probiotic viability during films production was strongly related to the pH of the film-forming solutions. Films with agrifood residues had a slower loss of probiotic viability during storage, when compared to plain starch films, which may be explained by the presence of antioxidant compounds. Inulin was expected to improve viability, but this was not observed. Microbiological analysis showed that agrifood residues powders contained natural contaminant bacteria that were partially eliminated during film formation. Moreover, none of the target foodborne pathogens were detected in the analyzed samples. Overall, the results suggest that edible films containing agrifood residues can be a promising material for the delivery of probiotics and/or as primary packaging for some food products.info:eu-repo/semantics/publishedVersio

The looming tide of nontuberculous mycobacterial infections in Portugal and Brazil

Nontuberculous mycobacteria (NTM) are widely disseminated in the environment and an emerging cause of infectious diseases worldwide. Their remarkable natural resistance to disinfectants and antibiotics and an ability to survive under low-nutrient conditions allows NTM to colonize and persist in man-made environments such as household and hospital water distribution systems. This overlap between human and NTM environments afforded new opportunities for human exposure, and for expression of their often neglected and underestimated pathogenic potential. Some risk factors predisposing to NTM disease have been identified and are mainly associated with immune fragilities of the human host. However, infections in apparently immunocompetent persons are also increasingly reported. The purpose of this review is to bring attention to this emerging health problem in Portugal and Brazil and to emphasize the urgent need for increased surveillance and more comprehensive epidemiological data in both countries, where such information is scarce and seriously thwarts the adoption of proper preventive strategies and therapeutic options.We acknowledge the support of FEDER through COMPETE and of National Funds through FCT - Fundacao para a Ciencia e a Tecnologia grants FCOMP-01-0124-FEDER-028359 [PTDC/BIA-MIC/2779/2012] and UID/NEU/04539/2013.info:eu-repo/semantics/publishedVersio

Improved diabetic wound healing by LFcinB is associated with relevant changes in the skin immune response and microbiota

Bovine lactoferricin (LFcinB) has antimicrobial and immunomodulatory properties; however, the effects on diabetic wound healing remain poorly understood. The wound healing potential of LFcinB was investigated with in vitro, ex vivo, and in vivo models. Cell migration and proliferation were tested on keratinocytes and on porcine ears. A type 1 diabetic mouse model was also used to evaluate wound healing kinetics, bacterial diversity patterns, and the effect of LFcinB on oxidative stress, macrophage phenotype, angiogenesis, and collagen deposition. LFcinB increased keratinocyte migration in vitro (p < 0.05) and ex vivo (p < 0.001) and improved wound healing in diabetic mice (p < 0.05), though not in normoglycemic control mice. In diabetic mouse wounds, LFcinB treatment led to the eradication of Bacillus pumilus, a decrease in Staphylococcus aureus, and an increase in the Staphylococcus xylosus prevalence. LFcinB increased angiogenesis in diabetic mice (p < 0.01), but this was decreased in control mice (p < 0.05). LFcinB improved collagen deposition in both diabetic and control mice (p < 0.05). Both oxidative stress and the M1-to-M2 macrophage ratios were decreased in LFcinB-treated wounds of diabetic animals (p < 0.001 and p < 0.05, respectively) compared with saline, suggesting a downregulation of inflammation in diabetic wounds. In conclusion, LFcinB treatment demonstrated noticeable positive effects on diabetic wound healing

Self-recycling and partially conservative replication of mycobacterial methylmannose polysaccharides

The steep increase in nontuberculous mycobacteria (NTM) infections makes understanding their unique physiology an urgent health priority. NTM synthesize two polysaccharides proposed to modulate fatty acid metabolism: the ubiquitous 6-O-methylglucose lipopolysaccharide, and the 3-O-methylmannose polysaccharide (MMP) so far detected in rapidly growing mycobacteria. The recent identification of a unique MMP methyltransferase implicated the adjacent genes in MMP biosynthesis. We report a wide distribution of this gene cluster in NTM, including slowly growing mycobacteria such as Mycobacterium avium, which we reveal to produce MMP. Using a combination of MMP purification and chemoenzymatic syntheses of intermediates, we identified the biosynthetic mechanism of MMP, relying on two enzymes that we characterized biochemically and structurally: a previously undescribed ?-endomannosidase that hydrolyses MMP into defined-sized mannoligosaccharides that prime the elongation of new daughter MMP chains by a rare ?-(1?4)-mannosyltransferase. Therefore, MMP biogenesis occurs through a partially conservative replication mechanism, whose disruption affected mycobacterial growth rate at low temperature