Cholesteryl Hemiazelate Present in Cardiovascular Disease Patients Causes Lysosome Dysfunction in Murine Fibroblasts.

There is growing evidence supporting the role of fibroblasts in all stages of atherosclerosis, from the initial phase to fibrous cap and plaque formation. In the arterial wall, as with macrophages and vascular smooth muscle cells, fibroblasts are exposed to a myriad of LDL lipids, including the lipid species formed during the oxidation of their polyunsaturated fatty acids of cholesteryl esters (PUFA-CEs). Recently, our group identified the final oxidation products of the PUFA-CEs, cholesteryl hemiesters (ChE), in tissues from cardiovascular disease patients. Cholesteryl hemiazelate (ChA), the most prevalent lipid of this family, is sufficient to impact lysosome function in macrophages and vascular smooth muscle cells, with consequences for their homeostasis. Here, we show that the lysosomal compartment of ChA-treated fibroblasts also becomes dysfunctional. Indeed, fibroblasts exposed to ChA exhibited a perinuclear accumulation of enlarged lysosomes full of neutral lipids. However, this outcome did not trigger de novo lysosome biogenesis, and only the lysosomal transcription factor E3 (TFE3) was slightly transcriptionally upregulated. As a consequence, autophagy was inhibited, probably via mTORC1 activation, culminating in fibroblasts' apoptosis. Our findings suggest that the impairment of lysosome function and autophagy and the induction of apoptosis in fibroblasts may represent an additional mechanism by which ChA can contribute to the progression of atherosclerosis.

INFLAMMATORY BOWEL DISEASE AND SARCOPENIA: A FOCUS ON MUSCLE STRENGTH - NARRATIVE REVIEW.

•Muscle strength decline is a crucial factor for the course of sarcopenia in inflammatory bowel disease (IBD) patients. •There is a need to discuss the association between IBD and sarcopenia focusing not only on changes of muscle mass, but also on muscle strength. •A narrative review was conducted in order to present the set of factors with impact in both muscle strength and IBD. •Inflammation, reduced nutrient intake and malabsorption, changes in body composition and gut microbiota dysbiosis are most likely the main factors with impact on muscle strength in IBD patients. Inflammation, changes in nutrient absorption and gut dysbiosis are common conditions in patients with inflammatory bowel disease. These factors may lead to variations in macro- and micronutrients and, particularly, to an imbalance of protein metabolism, loss of muscle mass and development of sarcopenia. This narrative review aims to present the set of factors with impact in muscle strength and physical performance that may potentially mediate the relation between inflammatory bowel disease and sarcopenia. Studies that associated changes in muscle strength, sarcopenia and inflammatory bowel disease were selected through a literature search in databases Medline, Pubmed and Scielo using relevant keywords: muscle strength, physical performance, sarcopenia and inflammatory bowel disease. Chronic inflammation is currently reported as a determinant factor in the development of muscle atrophy in inflammatory bowel disease. In addition, strength decline in inflammatory bowel disease patients may be also influenced by changes in body composition and by gut dysbiosis. Measures of muscle strength and physical performance should be considered in the initial identification of sarcopenia, particularly in patients with inflammatory bowel disease, for a timely intervention can be provided. Presence of proinflammatory cytokines, high adiposity, malabsorption and consequent deficits of macro and micronutrients, loss of muscle mass, and gut dysbiosis may be the main factors with impact in muscle strength, that probably mediate the relation between inflammatory bowel disease and sarcopenia.

Doença inflamatória intestinal e sarcopenia: um foco na força muscular - revisão narrativa / Inflammatory bowel disease and sarcopenia: a focus on muscle strength - narrative review

ABSTRACT Inflammation, changes in nutrient absorption and gut dysbiosis are common conditions in patients with inflammatory bowel disease. These factors may lead to variations in macro- and micronutrients and, particularly, to an imbalance of protein metabolism, loss of muscle mass and development of sarcopenia. This narrative review aims to present the set of factors with impact in muscle strength and physical performance that may potentially mediate the relation between inflammatory bowel disease and sarcopenia. Studies that associated changes in muscle strength, sarcopenia and inflammatory bowel disease were selected through a literature search in databases Medline, Pubmed and Scielo using relevant keywords: muscle strength, physical performance, sarcopenia and inflammatory bowel disease. Chronic inflammation is currently reported as a determinant factor in the development of muscle atrophy in inflammatory bowel disease. In addition, strength decline in inflammatory bowel disease patients may be also influenced by changes in body composition and by gut dysbiosis. Measures of muscle strength and physical performance should be considered in the initial identification of sarcopenia, particularly in patients with inflammatory bowel disease, for a timely intervention can be provided. Presence of proinflammatory cytokines, high adiposity, malabsorption and consequent deficits of macro and micronutrients, loss of muscle mass, and gut dysbiosis may be the main factors with impact in muscle strength, that probably mediate the relation between inflammatory bowel disease and sarcopenia.

RESUMO Inflamação, alterações na absorção de nutrientes e a disbiose intestinal são condições comuns em indivíduos com doença inflamatória intestinal. Estes fatores podem levar a variações corporais do teor de macro e micronutrientes e, em particular, a um desequilíbrio no metabolismo de proteínas com perda de massa muscular e desenvolvimento de sarcopenia. Esta revisão narrativa visa apresentar o conjunto de fatores com impacto na força e função muscular que podem potencialmente mediar a relação entre doença inflamatória intestinal e sarcopenia. Estudos que associaram as alterações de força muscular, sarcopenia e doença inflamatória intestinal foram selecionados, através de uma pesquisa bibliográfica nas bases de dados Medline, Pubmed e Scielo, usando palavras-chave relevantes: força muscular, desempenho físico, sarcopenia e doença inflamatória intestinal. A inflamação crónica é atualmente citada como um fator determinante no desenvolvimento de atrofia muscular nos casos de doença inflamatória intestinal. Além disso, o declínio de força em indivíduos com doença inflamatória intestinal, também pode ser influenciado pelas alterações na composição corporal e pela disbiose instestinal. Indicadores de força muscular e de desempenho físico devem ser considerados na identificação inicial de sarcopenia, principalmente em indivíduos com doença inflamatória intestinal, para que uma intervenção precoce possa ocorrer. A presença de citocinas pró-inflamatórias, elevada adiposidade corporal, má absorção intestinal com consequente déficit de macro e micronutrientes, perda de massa muscular e disbiose intestinal poderão ser os principais fatores com impacto na força muscular, que provavelmente medeiam a relação entre doença inflamatória intestinal e sarcopenia.

The Chlamydia trachomatis IncM Protein Interferes with Host Cell Cytokinesis, Centrosome Positioning, and Golgi Distribution and Contributes to the Stability of the Pathogen-Containing Vacuole.

Chlamydia trachomatis is an obligate intracellular bacterial pathogen that causes ocular and urogenital infections in humans. The ability of C. trachomatis to grow intracellularly in a pathogen-containing vacuole (known as an inclusion) depends on chlamydial effector proteins transported into the host cell by a type III secretion system. Among these effectors, several inclusion membrane proteins (Incs) insert in the vacuolar membrane. Here, we show that human cell lines infected by a C. trachomatis strain deficient for Inc CT288/CTL0540 (renamed IncM) displayed less multinucleation than when infected by IncM-producing strains (wild type or complemented). This indicated that IncM is involved in the ability of Chlamydia to inhibit host cell cytokinesis. The capacity of IncM to induce multinucleation in infected cells was shown to be conserved among its chlamydial homologues and appeared to require its two larger regions predicted to be exposed to the host cell cytosol. C. trachomatis-infected cells also displayed IncM-dependent defects in centrosome positioning, Golgi distribution around the inclusion, and morphology and stability of the inclusion. The altered morphology of inclusions containing IncM-deficient C. trachomatis was further affected by depolymerization of host cell microtubules. This was not observed after depolymerization of microfilaments, and inclusions containing wild-type C. trachomatis did not alter their morphology upon depolymerization of microtubules. Overall, these findings suggest that IncM may exert its effector function by acting directly or indirectly on host cell microtubules.

Toward Redox-Free Reverse Electrodialysis with Carbon-Based Slurry Electrodes.

Clean and renewable salinity gradient energy can be harvested using reverse electrodialysis (RED). The electrode system is an essential part to convert ionic current into electrical current. In this study, a typical 0.10 × 0.10 m2 RED stack with a cross-flow configuration was used to test carbon-based slurry electrodes (CSEs) to replace the usual redox solutions, like hexacyanoferrate, to enhance the RED process' sustainability, stability, and economic value. Six different slurry compositions comprising activated carbon, carbon black, and graphite powder were tested. The CSE characteristics were systematically studied by measuring viscosity, electrode compartment pressure drop, maximum current density, stability, and performance of power density and energy efficiency. Using a single membrane configuration, the CSE ran continuously for 17 days with a stable output. The application of CSEs for RED, with artificial seawater and river water, using mixing activated carbon and carbon black at a total concentration of 20 wt %, resulted in the best performance with a net power density of 0.7 W·m-2. Moreover, higher current densities up to 350 A·m-2 were tested for ED and shown to be feasible until 150 A·m-2. CSEs show promising versatility for different application modes.

eHealthResp, a Digital Intervention to Improve Antibiotic Prescribing in Respiratory Infections: A Pilot Study.

The emergence of antibiotic resistance (ABR) is one of the most serious public health threats worldwide. The inappropriate use of antibiotics is considered the main determinant for the increase and development of ABR, contributing to a greater risk of therapeutic ineffectiveness, particularly within primary care context. Therefore, this pilot study aims to raise awareness and promote an adequate antibiotic use among physicians, through the evaluation of the eHealthResp platform, a digital intervention composed by an online course and a mobile application, to aid in the management of respiratory tract infections. The global validation of the eHealthResp platform was carried out by 12 physicians who explored and performed a quantitative and qualitative evaluation of the contents of the online course and mobile app. The global evaluation of the analyzed parameters was very positive, with the highest median scores being attributed to adequacy, correction, format, and trust of the eHealthResp platform. The qualitative feedback enhanced the contents' relevance, clarity, and consolidation, as well as the effectiveness of the educational intervention against ABR. Overall, this study revealed that the eHealthResp may be regarded as an important e-health tool for the management of respiratory tract infections and improvement of antibiotic prescription practices among physicians.

Improving Pharmacists' Awareness of Inadequate Antibiotic Use for URTIs through an Educational Intervention: A Pilot Study.

The inadequate use of antibiotics led to the development of multi-resistant bacteria that are now causing millions of deaths worldwide. Since most antibiotics are prescribed/dispensed to treat respiratory tract infections, it is important to raise awareness among health professionals to optimize antibiotic use, especially within the primary care context. Thus, this pilot study aimed to evaluate pharmacists' feedback about the eHealthResp platform, composed by an online course and a mobile application (app) to help in the management of upper respiratory tract infections (URTIs). Ten community pharmacists were invited to participate in this study, exploring the contents of the eHealthResp platforms and answering a content validation questionnaire composed by eight qualitative and thirty-five quantitative questions about the online course and mobile app. The eHealthResp platform is a comprehensive, consistent, and high-quality e-learning tool. Median scores of 5.00 were attributed to the course contents' and clinical cases' adequacy and correction. Most qualitative feedback was about completeness and objectivity of the course, and its usefulness for clinical practice. This study showed that eHealthResp has great potential as an e-health tool for the management of URTIs' symptoms, which may ultimately aid in reducing inappropriate antibiotic use.

FODMAPs, inflammatory bowel disease and gut microbiota: updated overview on the current evidence.

PURPOSE: Based on the fermentable oligosaccharides, disaccharides, monosaccharides and polyols (FODMAP) hypothesis, the low-FODMAP diet has been suggested as a potential therapeutic approach for inflammatory bowel disease (IBD) with promising results on disease management. However, this diet implies a specific broad food restriction, which potentially increases the risk of nutritional deficiencies and may aggravate gut microbiota dysbiosis of IBD patients. The aim of the present study is to review the effect of individual FODMAPs on the human gut microbiota. In addition, this narrative review provides an updated overview of the use of the low-FODMAP diet in IBD, namely the implementation, advantages, limitations, and the impact on the gut microbiota. METHODS: The literature search strategy was applied to PubMed and Web of Science using relevant keywords, IBD, FODMAPs, Fructose, Lactose, Polyols, FOS, GOS, low-FODMAP diet and gut microbiota. RESULTS: Current data suggest that the low-FODMAP diet may effectively improve clinical outcomes in the management of IBD and ensure better quality of life for IBD patients. However, there is evidence highlighting some issues of concern, particularly the adequacy of the diet and the impact on the gut microbiota. The various FODMAP types differently modulate the gut microbiota. CONCLUSION: IBD management should be achieved with the least possible dietary restriction to avoid detrimental consequences, particularly on nutritional adequacy and gut microbiota. Thus, it is important to individualize and monitor the nutrition intervention. Further studies are required to better characterize the relationship between diet, the gut microbiota, and IBD to support the generalization of this approach for clinical practice in IBD therapy and management.

Lessons learned from urgent computing in Europe: Tackling the COVID-19 pandemic.

PRACE (Partnership for Advanced Computing in Europe), an international not-for-profit association that brings together the five largest European supercomputing centers and involves 26 European countries, has allocated more than half a billion core hours to computer simulations to fight the COVID-19 pandemic. Alongside experiments, these simulations are a pillar of research to assess the risks of different scenarios and investigate mitigation strategies. While the world deals with the subsequent waves of the pandemic, we present a reflection on the use of urgent supercomputing for global societal challenges and crisis management.

Development of an international Core Outcome Set (COS) for best care for the dying person: study protocol.

BACKGROUND: In contrast to typical measures employed to assess outcomes in healthcare such as mortality or recovery rates, it is difficult to define which specific outcomes of care are the most important in caring for dying individuals. Despite a variety of tools employed to assess different dimensions of palliative care, there is no consensus on a set of core outcomes to be measured in the last days of life. In order to optimise decision making in clinical practice and comparability of interventional studies, we aim to identify and propose a set of core outcomes for the care of the dying person. METHODS: Following the COMET initiative approach, the proposed study will proceed through four stages to develop a set of core outcomes: In stage 1, a systematic review of the literature will identify outcomes measured in existing peer reviewed literature, as well as outcomes derived through qualitative studies. Grey literature, will also be included. Stage 2 will allow for the identification and determination of patient and proxy defined outcomes of care at the end of life via quantitative and qualitative methods at an international level. In stage 3, from a list of salient outcomes identified through stages 1 and 2, international experts, family members, patients, and patient advocates will be asked to score the importance of the preselected outcomes through a Delphi process. Stage 4 consists of a face-to-face consensus meeting of international experts and patient/family representatives in order to define, endorse, and propose the final Core Outcomes Set. DISCUSSION: Core Outcome Sets aim at promoting uniform assessment of care outcomes in clinical practice as well as research. If consistently employed, a robust set of core outcomes for the end of life, and specifically for the dying phase, defined by relevant stakeholders, can ultimately be translated into best care for the dying person. Patient care will be improved by allowing clinicians to choose effective and meaningful treatments, and research impact will be improved by employing internationally agreed clinically relevant endpoints and enabling accurate comparison between studies in systematic reviews and/or in meta-analyses.

Quantification and phenotypic characterization of peripheral blood Vδ1 + T cells in chronic lymphocytic leukemia and monoclonal B cell lymphocytosis.

BACKGROUND: Vδ1+ T cells, a subset of γδ T cells, are responsible for innate-like immune responses. Recently, an anti-tumor function mediated by MHC-unrestricted recognition of lipid and stress molecules, has also been described in these cells. This study aimed to quantify and phenotypically characterize circulating Vδ1+ T cells in B cell Chronic Lymphocytic Leukemia (CLL) and Monoclonal B cell lymphocytosis (MBL). METHODS: This study enrolled 58 individuals distributed in five groups: Binet B and C CLL (n = 9), Binet A CLL (n = 26), High count-MBL (n = 10), Low count-MBL (n = 5), and a control group (n = 8). The phenotypic characterization of Vδ1+ T cells, as well as the other T cell subpopulations (CD4+ , CD8+ , CD4+ /CD8+ , and Vδ1- γδT cells), were assessed by flow cytometry, evaluating the frequency of each subset expressing CD27, CD69, and cytoplasmic granzyme B. RESULTS: We observed an increasing percentage of Vδ1+ T cells belonging to CD27- compartment from controls to advanced stages of the disease, which was accompanied by an increasing percentage of these cells expressing granzyme B, a phenotypic pattern that was also observed in the other T cell subpopulations under study since earlier stages of the disease. Moreover, a striking expansion of Vδ1+ T cells in Binet B and C CLL was observed. CONCLUSIONS: These experiment findings point to an expansion of CD27- Vδ1+ T cells with a cytotoxic profile, from controls to advanced stages of the disease, which points to a role of Vδ1+ T cells in the host's anti-tumor responses against clonal B-cells in MBL and CLL. © 2018 Clinical Cytometry Society.

Evidences and perspectives in the utilization of CLNA isomers as bioactive compounds in foods.

Conjugated alpha linolenic acid (CLNA) isomers are promising lipids owing to their similarities with conjugated linoleic acid (CLA) but exerting their bioactivity at lower doses; some isomers also belong to omega 3 family. This review aims to summarize the state of the art about the utilization of CLNA as a functional ingredient. Indeed, in vitro and in vivo studies reported that CLNA exerted anticancer, anti-inflammatory, anti-obese, and antioxidant activities. However, CLNA has not been tested in humans. These compounds are naturally present in meat and milk fat from ruminants but the highest concentrations are found in vegetable oils. Their incorporation in foodstuffs is one of the most effective strategies to elaborate CLNA-enriched products together with the microbiological production. Lactobacilli, propionibacteria, and bifidobacteria strains have been assayed to produce CLNA isomers but at the current moment there are not high CLNA concentration products elaborated using these strains. Furthermore, it is known that CLNA isomers are highly prone to oxidation when compared with linoleic acid and CLA, but the possible effects of elaboration and storage on high CLNA productsare unknown.The utilization of CLNA as a functional compound still remains a challenge and requires more research to address all of its technological and bioactivity aspects.

Initial assessment of patients without cognitive failure admitted to palliative care: a validation study.

OBJECTIVE: Many assessment tools have been developed for palliative care and there are a number of differences between them. Therefore, we felt that there was room for improvement. METHODS: In a previous study, the relevant items were selected by a Delphi process with international experts in palliative care. A 5-point verbal scale was added to the items selected and adapted to the different kind of items. RESULTS: The study included 123 patients, 63 (51%) were female and the median age was 64 (37 to 88). A four-factor structure was found through the principal components analysis, explaining 60.1% of the total variance. The scale presented good reliability, with a Cronbach's alpha of 0.72. It was hypothesized as a validity of construct that as the total symptom burden increased, survival time would decrease. This hypothesis was confirmed by the statistical analysis performed. A hazard ratio of 1.016 (P=0.019) was obtained in the Cox regression model including the final score as an explanatory variable of survival time, which means that for each increment of 1% in the total score, there was an increased risk of death of 1.6%. CONCLUSIONS: This tool is in accordance with the recommended characteristics that an assessment tool should have. It is simple to administer and easy to explain, complete and analyze. It is also a valid tool.

Microbial Contaminants of Cord Blood Units Identified by 16S rRNA Sequencing and by API Test System, and Antibiotic Sensitivity Profiling.

Over a period of ten months a total of 5618 cord blood units (CBU) were screened for microbial contamination under routine conditions. The antibiotic resistance profile for all isolates was also examined using ATB strips. The detection rate for culture positive units was 7.5%, corresponding to 422 samples.16S rRNA sequence analysis and identification with API test system were used to identify the culturable aerobic, microaerophilic and anaerobic bacteria from CBUs. From these samples we recovered 485 isolates (84 operational taxonomic units, OTUs) assigned to the classes Bacteroidia, Actinobacteria, Clostridia, Bacilli, Betaproteobacteria and primarily to the Gammaproteobacteria. Sixty-nine OTUs, corresponding to 447 isolates, showed 16S rRNA sequence similarities above 99.0% with known cultured bacteria. However, 14 OTUs had 16S rRNA sequence similarities between 95 and 99% in support of genus level identification and one OTU with 16S rRNA sequence similarity of 90.3% supporting a family level identification only. The phenotypic identification formed 29 OTUs that could be identified to the species level and 9 OTUs that could be identified to the genus level by API test system. We failed to obtain identification for 14 OTUs, while 32 OTUs comprised organisms producing mixed identifications. Forty-two OTUs covered species not included in the API system databases. The API test system Rapid ID 32 Strep and Rapid ID 32 E showed the highest proportion of identifications to the species level, the lowest ratio of unidentified results and the highest agreement to the results of 16S rRNA assignments. Isolates affiliated to the Bacilli and Bacteroidia showed the highest antibiotic multi-resistance indices and microorganisms of the Clostridia displayed the most antibiotic sensitive phenotypes.

Habitual dietary intake is associated with stool microbiota composition in monozygotic twins.

The impact of diet on the gut microbiota has usually been assessed by subjecting people to the same controlled diet and thereafter following the shifts in the microbiota. In the present study, we used habitual dietary intake, clinical data, quantitative polymerase chain reaction, and denaturing gradient gel electrophoresis (DGGE) to characterize the stool microbiota of Finnish monozygotic twins. The effect of diet on the numbers of bacteria was described through a hierarchical linear mixed model that included the twin individuals, stratified by body mass index, and their families as random effects. The abundance and diversity of the bacterial groups studied did not differ between normal-weight, overweight, and obese individuals with the techniques used. Intakes of energy, monounsaturated fatty acids, n3 polyunsaturated fatty acids (PUFAs), n6 PUFAs, and soluble fiber had significant associations with the stool bacterial numbers (e.g., increased energy intake was associated with reduced numbers of Bacteroides spp.). In addition, co-twins with identical energy intake had more similar numbers and DGGE-profile diversities of Bacteroides spp. than did the co-twins with different intake. Moreover, the co-twins who ingested the same amounts of saturated fatty acids had very similar DGGE profiles of Bacteroides spp., whereas the co-twins with similar consumption of fiber had a very low bifidobacterial DGGE-profile similarity. In conclusion, our findings confirm that the diet plays an important role in the modulation of the stool microbiota, in particular Bacteroides spp. and bifidobacteria.

The currently used commercial DNA-extraction methods give different results of clostridial and actinobacterial populations derived from human fecal samples.

Recently several human health-related microbiota studies have had partly contradictory results. As some differences may be explained by methodologies applied, we evaluated how different storage conditions and commonly used DNA-extraction kits affect bacterial composition, diversity, and numbers of human fecal microbiota. According to our results, the DNA-extraction did not affect the diversity, composition, or quantity of Bacteroides spp., whereas after a week's storage at -20 °C, the numbers of Bacteroides spp. were 1.6-2.5 log units lower (P < 0.05). Furthermore, the numbers of predominant bacteria, Eubacterium rectale (Erec)-group, Clostridium leptum group, bifidobacteria, and Atopobium group were 0.5-4 log units higher (P < 0.05) after mechanical DNA-extraction as detected with qPCR, regardless of storage. Furthermore, the bacterial composition of Erec-group differed significantly after different DNA-extractions; after enzymatic DNA-extraction, the most prevalent genera detected were Roseburia (39% of clones) and Coprococcus (10%), whereas after mechanical DNA-extraction, the most prevalent genera were Blautia (30%), Coprococcus (13%), and Dorea (10%). According to our results, rigorous mechanical lysis enables detection of higher bacterial numbers and diversity from human fecal samples. As it was shown that the results of clostridial and actinobacterial populations are highly dependent on the DNA-extraction methods applied, the use of different DNA-extraction protocols may explain the contradictory results previously obtained.

Comparative study of Curiementor ionization chambers using Monte Carlo simulations.

Ionization chambers (ICs) are the most commonly used instrument for measuring activities of solutions containing radiopharmaceuticals used in nuclear medicine, due to their simplicity of operation and accurate measurement results. As the dose delivered to the patient is determined by these measurements, for optimal results, the activity of these radiopharmaceuticals must be determined as accurately as possible, so that the radioprotection requirements are accomplished and at the same time good therapy or imaging results are achieved. In this work, the response of the ionization chamber Curiementor 2, from PTW-Freiburg, was studied by simulation with Monte Carlo (MC) code MCNPX and compared with experimental results for several radionuclides. The good agreement between experimental and simulation results validates the used methodology. The energy dependent sensitivity function, S (E), was established. Several ionization chambers were studied for (18)F and (99m)Tc, through Monte Carlo simulation method. In particular, some alterations to the original design of the Curiementor 2 were analysed, as well as the geometry of more recent chambers developed by PTW, namely Curiementor 3 and PET Curiementor 4. Calibration coefficients were compared for sources at different positions. Importance volumes were defined and determined for the Curiementors 2 and 3 and its ratio related with the corresponding ratio of sensitivity functions.

Mannosylglycerate is essential for osmotic adjustment in Thermus thermophilus strains HB27 and RQ-1.

We disrupted the mpgS encoding mannosyl-3-phosphoglycerate synthase (MpgS) of Thermus thermophilus strains HB27 and RQ-1, by homologous recombination, to assess the role of the compatible solute mannosylglycerate (MG) in osmoadaptation of the mutants, to examine their ability to grow in NaCl-containing medium and to identify the intracellular organic solutes. Strain HB27 accumulated only MG when grown in defined medium containing 2% NaCl; mutant HB27M9 did not grow in the same medium containing more than 1% NaCl. When trehalose or MG was added, the mutant was able to grow up to 2% of NaCl and accumulated trehalose or MG, respectively, plus amino acids. T. thermophilus RQ-1 grew in medium containing up to 5% NaCl, accumulated trehalose and lower amounts of MG. Mutant RQ-1M1 lost the ability to grow in medium containing more than 3% NaCl and accumulated trehalose and moderate levels of amino acids. Exogenous MG did not improve the ability of the organism to grow above 3% NaCl, but caused a decrease in the levels of amino acids. Our results show that MG serves as a compatible solute primarily during osmoadaptation at low levels of NaCl while trehalose is primarily involved in osmoadaptation during growth at higher NaCl levels.

Organic solutes in Rubrobacter xylanophilus: the first example of di-myo-inositol-phosphate in a thermophile.

The thermophilic and halotolerant nature of Rubrobacter xylanophilus led us to investigate the accumulation of compatible solutes in this member of the deepest lineage of the Phylum Actinobacteria. Trehalose and mannosylglycerate (MG) were the major compounds accumulated under all conditions examined, including those for optimal growth. The addition of NaCl to a complex medium and a defined medium had a slight or negligible effect on the accumulation of these compatible solutes. Glycine betaine, di-myo-inositol-phosphate (DIP), a new phosphodiester compound, identified as di-N-acetyl-glucosamine phosphate and glutamate were also detected but in low or trace levels. DIP was always present, except at the highest salinity examined (5% NaCl) and at the lowest temperature tested (43 degrees C). Nevertheless, the levels of DIP increased with the growth temperature. This is the first report of MG and DIP in an actinobacterium and includes the identification of the new solute di-N-acetyl-glucosamine phosphate.

Truepera radiovictrix gen. nov., sp. nov., a new radiation resistant species and the proposal of Trueperaceae fam. nov.

Two isolates, belonging to a new species of a novel genus of the Phylum "Deinococcus/Thermus ", were recovered from hot spring runoffs on the Island of São Miguel in the Azores. Strains RQ-24(T) and TU-8 are the first cultured representatives of a distinct phylogenetic lineage within this phylum. These strains form orange/red colonies, spherical-shaped cells, have an optimum growth temperature of about 50 degrees C, an optimum pH for growth between about 7.5 and 9.5, and do not grow at pH below 6.5 or above pH 11.2. These organisms grow in complex media without added NaCl, but have a maximum growth rate in media with 1.0% NaCl and grow in media containing up to 6.0% NaCl. The organisms are extremely ionizing radiation resistant; 60% of the cells survive 5.0 kGy. These strains are chemoorganotrophic and aerobic; do not grow in Thermus medium under anaerobic conditions with or without nitrate as electron acceptor and glucose as a source of carbon and energy, but ferment glucose to D-lactate without formation of gas. The organisms assimilate a large variety of sugars, organic acids and amino acids. Fatty acids are predominantly iso- and anteiso-branched; long chain 1,2 diols were also found in low relative proportions; menaquinone 8 (MK-8) is the primary respiratory quinone. Peptidoglycan was not detected. Based on 16S rRNA gene sequence analysis, physiological, biochemical and chemical analysis we describe a new species of one novel genus represented by strain RQ-24(T) (CIP 108686(T)=LMG 22925(T)=DSM 17093(T)) for which we propose the name Truepera radiovictrix. We also propose the family Trueperaceae fam. nov. to accommodate this new genus.