Showing papers in "Fems Microbiology Letters in 2010"
TL;DR: The possible reasons for 'unculturability' are discussed, advances in the cultivation of previously unculturable bacteria from complex bacterial communities are evaluated and the provision of simulated natural environmental conditions for bacterial culture is evaluated.
Abstract: Molecular ecology methods are now well established for the culture-independent characterization of complex bacterial communities associated with various environmental and animal habitats and are revealing the extent of their diversity. By comparison, it has become clear that only a small minority of microorganisms are readily cultivated in vitro, with the majority of all bacteria remaining 'unculturable' using standard methods. Yet, it is only through the isolation of bacterial species in pure culture that they may be fully characterized, both for their physiological and pathological properties. Hence, the endeavour to devise novel cultivation methods for microorganisms that appear to be inherently resistant to artificial culture is a most important one. This minireview discusses the possible reasons for 'unculturability' and evaluates advances in the cultivation of previously unculturable bacteria from complex bacterial communities. Methods include the use of dilute nutrient media particularly suited for the growth of bacteria adapted to oligotrophic conditions, and the provision of simulated natural environmental conditions for bacterial culture. This has led to the recovery of 'unculturables' from soil and aquatic environments, likely to be due to the inclusion of essential nutrients and/or signalling molecules from the native environment.
623 citations
TL;DR: The results suggest that many samples collected and stored under field conditions without refrigeration may be useful for microbial community analyses, and environmental factors and biases in molecular techniques likely confer greater amounts of variation to microbial communities than do differences in short-term storage conditions.
Abstract: Storage conditions are considered to be a critical component of DNA-based microbial community analysis methods. However, whether differences in short-term sample storage conditions impact the assessment of bacterial community composition and diversity requires systematic and quantitative assessment. Therefore, we used barcoded pyrosequencing of bacterial 16S rRNA genes to survey communities, harvested from a variety of habitats [soil, human gut (feces) and human skin] and subsequently stored at 20, 4, -20 and -80 degrees C for 3 and 14 days. Our results indicate that the phylogenetic structure and diversity of communities in individual samples were not significantly influenced by the storage temperature or the duration of storage. Likewise, the relative abundances of most taxa were largely unaffected by temperature even after 14 days of storage. Our results indicate that environmental factors and biases in molecular techniques likely confer greater amounts of variation to microbial communities than do differences in short-term storage conditions, including storage for up to 2 weeks at room temperature. These results suggest that many samples collected and stored under field conditions without refrigeration may be useful for microbial community analyses.
363 citations
TL;DR: This review presents a basic description of microcalorimetry and examples of microbiological applications of IMC for medical and environmental microbiology, andMiniaturization of isothermal calorimeters provides an even wider range of possibilities.
Abstract: Isothermal calorimetry measures the heat flow of biological processes, which is proportional to the rate at which a given chemical or physical process takes place. Modern isothermal microcalorimeters make measurements of less than a microwatt of heat flow possible. As a result, as few as 10 000-100 000 active bacterial cells in culture are sufficient to produce a real-time signal dynamically related to the number of cells present and their activity. Specimens containing bacteria need little preparation, and isothermal microcalorimetry (IMC) is a nondestructive method. After IMC measurements, the undisturbed samples can be evaluated by any other means desired. In this review, we present a basic description of microcalorimetry and examples of microbiological applications of IMC for medical and environmental microbiology. In both fields, IMC has been used to quantify microbial activity over periods of hours or even days. Finally, the recent development of highly parallel instruments (up to 48 channels) and the constantly decreasing costs of equipment have made IMC increasingly attractive for microbiology. Miniaturization of isothermal calorimeters provides an even wider range of possibilities.
247 citations
TL;DR: In this review, the different EPS from B. subtilis were classified into four main functional categories: structural (neutral polymers), sorptive (charged polymers, surface-active and active polymers) and current information regarding the genetic expression, production and function of the main polymers secreted by B.subilis has been compiled.
Abstract: Bacterial exopolymeric substances (EPS) are molecules released in response to the physiological stress encountered in the natural environment. EPS are structural components of the extracellular matrix in which cells are embedded during biofilm development. The chemical nature and functions of these EPS are dependent on the genetic expression of the cells within each biofilm. Although some bacterial matrices have been characterized, understanding of the function of the EPS is relatively limited, particularly within the Bacillus genus. Similar gaps of knowledge exist with respect to the chemical composition and specific roles of the macromolecules secreted by Bacillus subtilis in its natural environment. In this review, the different EPS from B. subtilis were classified into four main functional categories: structural (neutral polymers), sorptive (charged polymers), surface-active and active polymers. In addition, current information regarding the genetic expression, production and function of the main polymers secreted by B. subtilis strains, particularly those related to biofilm formation and its architecture, has been compiled. Further characterization of these EPS from B. subtilis remains a challenge.
246 citations
TL;DR: The divergent repertoires of the virulence factors responsible for the pathogenesis of the organisms and that ultimately result in the distinct clinical outcomes of infection are detailed.
Abstract: Salmonella enterica represents a major human and animal pathogen. Many S. enterica genomes have been completed and many more genome sequencing projects are underway, constituting an excellent resource for comparative genome analysis studies leading to a better understanding of bacterial evolution and pathogenesis. Salmonella enterica serovar Typhimurium and Typhi are the best-characterized serovars, with the first being involved in localized gastroenteritis in many hosts and the latter causing a systemic human-specific disease. Here, we summarize the major genetic differences between the two different serovars. We detail the divergent repertoires of the virulence factors responsible for the pathogenesis of the organisms and that ultimately result in the distinct clinical outcomes of infection. This comparative genomic overview highlights hypotheses for future investigations on S. enterica pathogenesis and the basis of host specificity.
230 citations
TL;DR: A role for ACCD in the plant root growth-promotion effect by T. asperellum is suggested as well as decreased ability of the mutants to promote root elongation of canola seedlings.
Abstract: 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity was evaluated in the biocontrol and plant growth-promoting fungus Trichoderma asperellum T203. Fungal cultures grown with ACC as the sole nitrogen source showed high enzymatic activity. The enzyme encoding gene (Tas-acdS) was isolated, and an average 3.5-fold induction of the gene by 3 mM ACC was detected by real-time PCR. Escherichia coli bacteria carrying the intron-free cDNA of Tas-acdS cloned into the vector pAlter-EX1 under the control of the tac promoter revealed specific ACC deaminase (ACCD) activity and the ability to promote canola (Brassica napus) root elongation in pouch assays. RNAi silencing of the ACCD gene in T. asperellum showed decreased ability of the mutants to promote root elongation of canola seedlings. These data suggest a role for ACCD in the plant root growth-promotion effect by T. asperellum.
229 citations
TL;DR: Based on the phylogenetic position and the physiological characteristics of strain Sp3(T), this new syntrophic, acetate-oxidizing bacterium is proposed as the new genus and species Syntrophaceticus schinkii, with Sp3 (T) (=JCM 16669(T)) as the type strain.
Abstract: A mesophilic, syntrophic acetate-oxidizing bacterium, designated strain Sp3(T), was isolated from sludge from a mesophilic methanogenic digestor operating at a high ammonium concentration (6.4 g L(-1) NH(4)(+)-N). The strain showed acetate-oxidizing ability in cocultivation with a hydrogen-consuming methanogen. Comparative 16S rRNA gene sequence analysis confirmed that strain Sp3(T) belonged to the Firmicutes-Clostridia class. The most closely related species was Thermacetogenium phaeum (16S rRNA gene sequence identity 92%). Strain Sp3(T) used ethanol, betaine and lactate as carbon and electron sources and showed growth between 25 and 40 degrees C and pH 6.0 and 8.0. Based on the phylogenetic position and the physiological characteristics of strain Sp3(T), this new syntrophic, acetate-oxidizing bacterium is proposed as the new genus and species Syntrophaceticus schinkii, with Sp3(T) (=JCM 16669(T)) as the type strain. An isolate (strain Esp=JCM 16670) with high 16S rRNA gene sequence identity (99%) to syntrophic acetate-oxidizing Clostridium ultunense was also retrieved from the methanogenic digestor.
205 citations
TL;DR: This review will discuss the opportunities of RNA-seq transcriptome sequencing for microorganisms, and also aims to identify challenges and pitfalls of the use of this new technology in microorganisms.
Abstract: Over the past 15 years, microbial functional genomics has been made possible by the combined power of genome sequencing and microarray technology. However, we are now approaching the technical limits of microarray technology, and microarrays are now being superseded by transcriptomics based on high-throughput (next generation) DNA-sequencing technologies. The term RNA-seq has been coined to represent transcriptomics by next-generation sequencing. Although pioneered on eukaryotic organisms due to the relative ease of working with eukaryotic mRNA, the RNA-seq technology is now being ported to microbial systems. This review will discuss the opportunities of RNA-seq transcriptome sequencing for microorganisms, and also aims to identify challenges and pitfalls of the use of this new technology in microorganisms.
181 citations
TL;DR: Investigating how Pseudomonas aeruginosa influences A. fumigatus conidial germination and biofilm formation suggests that small diffusible and heat-stable molecules may be responsible for the competitive inhibition of filamentous fungal growth in polymicrobial environments such as the CF lung.
Abstract: Aspergillus fumigatus is often isolated from the lungs of cystic fibrosis (CF) patients, but unlike in severely immunocompromised individuals, the mortality rates are low. This suggests that competition from bacteria within the CF lung may be inhibitory. The purpose of this study was to investigate how Pseudomonas aeruginosa influences A. fumigatus conidial germination and biofilm formation. Aspergillus fumigatus biofilm formation was inhibited by direct contact with P. aeruginosa, but had no effect on preformed biofilm. A secreted heat-stable soluble factor was also shown to exhibit biofilm inhibition. Coculture of P. aeruginosa quorum-sensing mutants (PAO1:ΔLasI, PAO1:ΔLasR) did not significantly inhibit A. fumigatus biofilms (52.6-58.8%) to the same extent as that of the PA01 wild type (22.9-30.1%), both by direct and by indirect interaction (P<0.001). Planktonic and sessile inhibition assays with a series of short carbon chain molecules (decanol, decanoic acid and dodecanol) demonstrated that these molecules could both inhibit and disrupt biofilms in a concentration-dependent manner. Overall, this suggests that small diffusible and heat-stable molecules may be responsible for the competitive inhibition of filamentous fungal growth in polymicrobial environments such as the CF lung.
180 citations
TL;DR: Phylogenetic analysis of sequences for the D1/D2 domains of the nuclear large subunit rRNA gene placed all sophorolipid-producing species in the S. bombicola subclade of the Starmerella clade.
Abstract: Sophorolipids are carbohydrate-based, amphiphilic biosurfactants that are of increasing interest for use in environmentally benign cleaning agents. Sophorolipid production was tested for 26 strains representing 19 species of the Starmerella yeast clade, including Starmerella bombicola and Candida apicola, which were previously reported to produce sophorolipids. Five of the 19 species tested showed significant production of sophorolipids: S. bombicola, C. apicola, Candida riodocensis, Candida stellata and a new species, Candida sp. NRRL Y-27208. A high-throughput matrix-assisted laser desorption/ionization-time of flight MS assay was developed that showed S. bombicola and C. apicola to produce a lactone form of sophorolipid, whereas C. riodocensis, C. stellata and Candida sp. NRRL Y-27208 produced predominantly free acid sophorolipids. Phylogenetic analysis of sequences for the D1/D2 domains of the nuclear large subunit rRNA gene placed all sophorolipid-producing species in the S. bombicola subclade of the Starmerella clade.
175 citations
TL;DR: The mechanisms involved in bacterial biofilm formation and attachment on plant roots, and the relation of these mechanisms to rhizobial function and survival are reviewed.
Abstract: Biofilms are bacterial communities enclosed within an extracellular matrix of polysaccharides produced by the bacteria, which adhere to a living or an inert macrosurface. In nature, biofilms constitute a protected growth modality allowing bacteria to survive in hostile environments. Studies of environmental isolates have revealed a highly ordered, three-dimensional organization of the extracellular matrix, which has important implications for biofilm physiology. The zone of soil immediately surrounding a plant root where complex biological and ecological processes occur, termed rhizosphere, forms an environment that fulfills the requirements for biofilm formation, including sufficient moisture and supply of nutrients, which are provided by the plant. Biofilm formation on plants appears to be associated with symbiotic and pathogenic responses, but it is unclear how plants regulate the association. Biofilms function as structures resistant against stress factors such as desiccation, UV radiation, predation, and antibiosis, which help create protective niches for rhizobia. However, the role of biofilms in rhizobial–legume symbiosis remains to be clarified. Here, the mechanisms involved in bacterial biofilm formation and attachment on plant roots, and the relation of these mechanisms to rhizobial function and survival are reviewed.
TL;DR: The data suggest that uncultured Prevotella is more abundant than knownPrevotella and that members of this genus appear to have specific metabolic niches.
Abstract: 16S rRNA gene-based analysis of rumen Prevotella was carried out to estimate the diversity and diet specificity of bacteria belonging to this genus. Total DNA was extracted from the rumen digesta of three sheep fed two diets with different hay-to-concentrate ratios (10 : 1 and 1 : 2). Real-time PCR quantification of Prevotella revealed that the relative abundance of this genus in the total rumen bacteria was up to 19.7%, while the representative species Prevotella bryantii and Prevotella ruminicola accounted for only 0.6% and 3.8%, respectively. Denaturing gradient gel electrophoresis analysis for Prevotella revealed shifts in the community composition with the diet. Analysis of 16S rRNA gene clone libraries showed significant differences (P=0.001) between clones detected from the sheep on the diets with different hay-to-concentrate ratios. The majority (87.8%) of Prevotella clones had <97% sequence similarity with known rumen Prevotella. These data suggest that uncultured Prevotella is more abundant than known Prevotella and that members of this genus appear to have specific metabolic niches.
TL;DR: The aim of this research was to identify bacterial isolates having the potential to improve intestinal barrier function and to identify bacteria strains and human oral isolates screened for their ability to enhance tight junction integrity as measured by the transepithelial electrical resistance (TEER) assay.
Abstract: The aim of this research was to identify bacterial isolates having the potential to improve intestinal barrier function. Lactobacillus plantarum strains and human oral isolates were screened for their ability to enhance tight junction integrity as measured by the transepithelial electrical resistance (TEER) assay. Eight commercially used probiotics were compared to determine which had the greatest positive effect on TEER, and the best-performing probiotic strain, Lactobacillus rhamnosus HN001, was used as a benchmark to evaluate the isolates. One isolate, L. plantarum DSM 2648, was selected for further study because it increased TEER 135% more than L. rhamnosus HN001. The ability of L. plantarum DSM 2648 to tolerate gastrointestinal conditions and adhere to intestinal cells was determined, and L. plantarum DSM 2648 performed better than L. rhamnosus HN001 in all the assays. Lactobacillus plantarum DSM 2648 was able to reduce the negative effect of Escherichia coli [enteropathogenic E. coli (EPEC)] O127:H6 (E2348/69) on TEER and adherence by as much as 98.75% and 80.18%, respectively, during simultaneous or prior coculture compared with EPEC incubation alone. As yet, the precise mechanism associated with the positive effects exerted by L. plantarum DSM 2648 are unknown, and may influence its use to improve human health and wellness.
TL;DR: Six fungal endophytes were screened for the suppression of R. solani growth both in vitro and in a greenhouse and improved potato yield significantly and decreased the stem disease severity index of sensitive potato.
Abstract: Rhizoctonia solani is an important soilborne pathogen of potato plants whose control typically depends on chemicals. Here, we screened six fungal endophytes for the suppression of R. solani growth both in vitro and in a greenhouse. These isolates were identified using morphology and internal transcribed spacer regions of rDNA as Alternaria longipes, Epicoccum nigrum, Phomopsis sp., and Trichoderma atroviride. Both T. atroviride and E. nigrum showed significant in vitro inhibition of mycelial growth of R. solani, with the greatest inhibition zone observed for E. nigrum species in dual cultures. The highest inhibition was observed for T. atroviride. The inhibition rate was also significantly correlated with the culture filtrates of these isolates. Confocal microscopy showed that T. atroviride acts as a mycoparasite and competitor. However, E. nigrum and A. longipes produce secondary metabolites, while Phomospsis sp. competes for nutrients and space. Greenhouse experiments confirmed that T. atroviride and E. nigrum improved potato yield significantly and decreased the stem disease severity index of sensitive potato.
TL;DR: Genetic tools for labeling Gram-negative bacteria in vitro and in their natural environment without the necessity of antibiotic pressure for maintenance are developed and it is shown that mCherry in combination with GFP is a suitable marker for studying mixed microbial communities.
Abstract: Live-cell imaging techniques are essential to gain a better understanding of microbial functioning in natural systems, for example in biofilms. Autofluorescent proteins, such as the green fluorescent protein (GFP) and the red fluorescent protein (DsRed), are valuable tools for studying microbial communities in their natural environment. Because of the functional limitations of DsRed such as slow maturation and low photostability, new and improved variants were created such as mCherry. In this study, we developed genetic tools for labeling Gram-negative bacteria in order to visualize them in vitro and in their natural environment without the necessity of antibiotic pressure for maintenance. mcherry was cloned into two broad host-range cloning vectors and a pBK-miniTn7 transposon under the constitutive expression of the tac promoter. The applicability of the different constructs was shown in Escherichia coli, various Pseudomonas spp. and Edwardsiella tarda. The expression of mcherry was qualitatively analyzed by fluorescence microscopy and quantified by fluorometry. The suitability of the constructs for visualizing microbial communities was shown for biofilms formed on glass and tomato roots. In addition, it is shown that mCherry in combination with GFP is a suitable marker for studying mixed microbial communities.
TL;DR: Results suggested that multiple defense pathways in tobacco were involved in Trichokonin-mediated TMV resistance, which sheds light on the potential of peptaibols in plant viral disease control.
Abstract: Trichoderma spp. are well-known biocontrol agents because of their antimicrobial activity against bacterial and fungal phytopathogens. However, the biochemical mechanism of their antiviral activity remains largely unknown. In this study, we found that Trichokonins, antimicrobial peptaibols isolated from Trichoderma pseudokoningii SMF2, could induce defense responses and systemic resistance in tobacco (Nicotiana tabacum var. Samsun NN) against tobacco mosaic virus (TMV) infection. Local Trichokonin (100 nM) treatment led to 54% lesion inhibition, 57% reduction in average lesion diameter and 30% reduction in average lesion area in systemic tissue of tobacco compared with control, indicating that Trichokonins induced resistance in tobacco against TMV infection. Trichokonin treatment increased the production of reactive oxygen species and phenolic compounds in tobacco. Additionally, application of Trichokonins significantly increased activities of pathogenesis-related enzymes PAL and POD, and upregulated the expression of several plant defense genes. These results suggested that multiple defense pathways in tobacco were involved in Trichokonin-mediated TMV resistance. We report on the antivirus mechanism of peptaibols, which sheds light on the potential of peptaibols in plant viral disease control.
TL;DR: Defective energy production through mutations in sucB and ubiF affects persister survival and could serve as new drug targets for persister bacteria.
Abstract: Persisters are a small population of slowly growing or nongrowing bacteria that are phenotypically resistant to antibiotics, but the mechanisms involved are not well understood. The aim of this study is to determine new mechanisms underlying antibiotic-tolerant persisters. The Escherichia coli deletion mutant library was screened to identify mutants that had a defect in persister survival after exposure to ampicillin for 24 h or 5 days. The identified mutants and the parent strain were subjected to minimum inhibitory concentration (MIC) and minimum bactericidal tests and antibiotic or stress conditions in exposure assays. sucB and ubiF mutants deficient in energy production were identified from the mutant screens to have defective persister survival as demonstrated by higher susceptibility to various antibiotics, including ampicillin, norfloxacin, tetracycline and gentamicin, and different stresses such as oxidative stress, acid pH and weak acid compared with the parent strain. In addition, both sucB and ubiF had a twofold lower MIC than the parent strain. The above sucB and ubiF mutant phenotypes could be complemented by their respective functional genes. Defective energy production through mutations in sucB and ubiF affects persister survival and could serve as new drug targets for persister bacteria.
TL;DR: Hydrogen peroxide and lactic acid act co-operatively to kill enteric, vaginosis-associated and uropathogenic pathogens for hydrogen peroxide-producing Lactobacillus strains.
Abstract: The mechanism underlying the killing activity of Lactobacillus strains against bacterial pathogens appears to be multifactorial. Here, we investigate the respective contributions of hydrogen peroxide and lactic acid in killing bacterial pathogens associated with the human vagina, urinary tract or intestine by two hydrogen peroxide-producing strains. In co-culture, the human intestinal strain Lactobacillus johnsonii NCC933 and human vaginal strain Lactobacillus gasseri KS120.1 strains killed enteric Salmonella enterica serovar Typhimurium SL1344, vaginal Gardnerella vaginalis DSM 4944 and urinary tract Escherichia coli CFT073 pathogens. The cell-free culture supernatants (CFCSs) produced the same reduction in SL1344, DSM 4944 and CFT073 viability, whereas isolated bacteria had no effect. The killing activity of CFCSs was heat-stable. In the presence of Dulbecco's modified Eagle's minimum essential medium inhibiting the lactic acid-dependent killing activity, CFCSs were less effective at killing of the pathogens. Catalase-treated CFCSs displayed a strong decreased activity. Tested alone, hydrogen peroxide triggered a concentration-dependent killing activity against all three pathogens. Lactic acid alone developed a killing activity only at concentrations higher than that present in CFCSs. In the presence of lactic acid at a concentration present in Lactobacillus CFCSs, hydrogen peroxide displayed enhanced killing activity. Collectively, these results demonstrate that for hydrogen peroxide-producing Lactobacillus strains, the main metabolites of Lactobacillus, lactic acid and hydrogen peroxide, act co-operatively to kill enteric, vaginosis-associated and uropathogenic pathogens.
TL;DR: This work demonstrates that a group of strains of F. solani are responsible for the symptoms observed on turtle-nesting beaches, and that they represent a risk for the survival of this endangered species.
Abstract: The fungus Fusarium solani (Mart.) Saccardo (1881) was found to be the cause of infections in the eggs of the sea turtle species Caretta caretta in Boavista Island, Cape Verde. Egg shells with early and severe symptoms of infection, as well as diseased embryos were sampled from infected nests. Twenty-five isolates with similar morphological characteristics were obtained. Their ITS rRNA gene sequences were similar to the GenBank sequences corresponding to F. solani and their maximum identity ranged from 95% to 100%. Phylogenetic parsimony and Bayesian analyses of these isolates showed that they belong to a single F. solani clade and that they are distributed in two subclades named A and C (the latter containing 23 out of 25). A representative isolate of subclade C was used in challenge inoculation experiments to test Koch postulates. Mortality rates were c. 83.3% in challenged eggs and 8.3% in the control. Inoculated challenged eggs exhibited the same symptoms as infected eggs found in the field. Thus, this work demonstrates that a group of strains of F. solani are responsible for the symptoms observed on turtle-nesting beaches, and that they represent a risk for the survival of this endangered species.
TL;DR: This discovery extends the paradigm of AHL-mediated QS signalling beyond the Proteobacteria and reinforces its ecological significance in T. maritimum.
Abstract: Tenacibaculum maritimum (formerly Flexibacter maritimus) is a filamentous, biofilm-forming member of the Cytophaga–Flavobacterium–Bacteroides group (or Bacteroidetes), which causes the widely distributed marine fish disease tenacibaculosis. A search for N-acylhomoserine lactones (AHLs) quorum-sensing (QS) signals in the culture media of nine representative strains of this species using different biosensor strains revealed the presence of short-type AHL activity in all of them. N-butyryl-l-homoserine lactone (C4-HSL) was identified in T. maritimum NCIMB2154T by LC-MS. A degradation activity for long-acyl AHLs (C10-HSL) was subsequently demonstrated in T. maritimum NCIMB2154T. The acidification of the culture medium after degradation did not allow the recovery of C10-HSL, which indicates a possible acylase-type degradation activity. Even though the physiological processes under the control of AHL-mediated QS in T. maritimum need to be further characterized, this discovery extends the paradigm of AHL-mediated QS signalling beyond the Proteobacteria and reinforces its ecological significance.
TL;DR: The data suggest that a paucity of F. prausnitzii in the gastrointestinal microbial communities is likely to be a minor aetiological factor in CD: recovery following elemental diet is attributed to lower levels of gut flora.
Abstract: Reports that bacteria within the Firmicutes phylum, especially the species Faecalibacterium prausnitzii, are less abundant in Crohn's disease (CD) patients and supernatants from cultures of this bacterium are anti-inflammatory prompted the investigation of the possible correlations between the abundance of F. prausnitzii and the response to treatment in patients with gut diseases and healthy controls. In a randomized, double-blind trial, faeces were collected from healthy volunteers, and from patients with active CD, ulcerative colitis (UC) and irritable bowel syndrome before and after treatment. The levels of F. prausnitzii DNA in faecal suspensions were determined by PCR. Treatment by an elemental diet was effective, resulting in decreases in both the Harvey and Bradshaw index (P<0.001) and the concentrations of serum C-reactive protein (P<0.05). The total levels of F. prausnitzii in faecal samples from CD patients at presentation were lower than those in the other groups both before and after the treatment. There was no correlation between F. prausnitzii abundance and the severity of CD before treatment. Clinical improvement unexpectedly correlated with a significant decrease in the abundance of F. prausnitzii, especially the A2-165 subgroup (P<0.05). Our data suggest that a paucity of F. prausnitzii in the gastrointestinal microbial communities is likely to be a minor aetiological factor in CD: recovery following elemental diet is attributed to lower levels of gut flora.
TL;DR: An overview of the diverse strategies used in the model yeasts Saccharomyces cerevisiae and Schizosac charomyces pombe, and the pathogenic fungus Candida albicans, to sense and transduce stress signals to their respective SAPKs is presented.
Abstract: The ability of microorganisms to survive and thrive within hostile environments depends on rapid and robust stress responses. Stress-activated protein kinase (SAPK) pathways are important stress-signalling modules found in all eukaryotes, including eukaryotic microorganisms such as fungi. These pathways consist of a SAPK that is activated by phosphorylation through a kinase cascade, and once activated, the SAPK phosphorylates a range of cytoplasmic and nuclear target substrates, which determine the appropriate response. However, despite their conservation in fungi, mechanisms that have evolved to relay stress signals to the SAPK module in different fungi have diverged significantly. Here, we present an overview of the diverse strategies used in the model yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe, and the pathogenic fungus Candida albicans, to sense and transduce stress signals to their respective SAPKs.
TL;DR: It can be concluded that strains DY05(T) and 47666-1 belong to the same novel species of the genus Vibrio, for which the name VIBrio owensii sp.
Abstract: Two bacterial strains (DY05T and 47666-1) were isolated in Queensland, Australia, from diseased cultured crustaceans Panulirus ornatus and Penaeus monodon, respectively. On the basis of 16S rRNA gene sequence identity, the strains were shown to belong to the Harveyi clade of the genus Vibrio. Multilocus sequence analysis using five housekeeping genes (rpoA, pyrH, topA, ftsZ and mreB) showed that the strains form a monophyletic group with 94.4% concatenated sequence identity to the closest species. DNA–DNA hybridization experiments showed that strains DY05T and 47666-1 had 76% DNA similarity to each other, but o70% to their closest neighbours Vibrio harveyi LMG 4044T ( 55%), Vibrio campbellii LMG 11216T ( 52%) and Vibrio rotiferianus LMG 21460T ( 46%). Strains DY05T and 47666-1 could be differentiated from their relatives on the basis of several phenotypic characteristics. The major fatty acids were C15:0 iso 2-OH and/or C16:1 o7, C16:0, C18:1 o7 and C14:0. Based on the polyphasic evidence presented here, it can be concluded that strains DY05T and 47666-1 belong to the same novel species of the genus Vibrio, for which the name Vibrio owensii sp. nov. is proposed. The type strain is DY05T ( = JCM 16517T =ACM 5300T).
TL;DR: It is shown that inverted membrane vesicles from the slow-growing model strain Mycobacterium bovis BCG are active in ATP synthesis, but ATP synthase displays no detectable ATP hydrolysis activity and does not set up a proton-motive force (PMF) using ATP as a substrate.
Abstract: ATP synthase is a validated drug target for the treatment of tuberculosis, and ATP synthase inhibitors are promising candidate drugs for the treatment of infections caused by other slow-growing mycobacteria, such as Mycobacterium leprae and Mycobacterium ulcerans. ATP synthase is an essential enzyme in the energy metabolism of Mycobacterium tuberculosis; however, no biochemical data are available to characterize the role of ATP synthase in slow-growing mycobacterial strains. Here, we show that inverted membrane vesicles from the slow-growing model strain Mycobacterium bovis BCG are active in ATP synthesis, but ATP synthase displays no detectable ATP hydrolysis activity and does not set up a proton-motive force (PMF) using ATP as a substrate. Treatment with methanol as well as PMF activation unmasked the ATP hydrolysis activity, indicating that the intrinsic subunit ɛ and inhibitory ADP are responsible for the suppression of hydrolytic activity. These results suggest that the enzyme is needed for the synthesis of ATP, not for the maintenance of the PMF. For the development of new antimycobacterial drugs acting on ATP synthase, screening for ATP synthesis inhibitors, but not for ATP hydrolysis blockers, can be regarded as a promising strategy.
TL;DR: This study reports, for the first time, the characterization of dextransucrase from Weissella strains using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and in situ polymer production (after incubation with sucrose) from enzymatic fractions harvested from both sucrose and glucose culture media.
Abstract: The study of exopolysaccharide production by heterofermentative sourdough lactic acid bacteria has shown that Weissella strains isolated from sourdoughs produce linear dextrans containing α-(1→6) glucose residues with few α-(1→3) linkages from sucrose. In this study, several dextran-producing strains, Weissella cibaria and Weissella confusa, isolated from sourdough, were characterized according to carbohydrate fermentation, repetitive element-PCR fingerprinting using (GTG)(5) primers and glucansucrase activity (soluble or cell-associated). This study reports, for the first time, the characterization of dextransucrase from Weissella strains using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and in situ polymer production (after incubation with sucrose) from enzymatic fractions harvested from both sucrose and glucose culture media. Results demonstrate that dextransucrase activity was mainly soluble and associated with a constitutive 180-kDa protein. In addition, microsequencing of the active dextransucrase from W. cibaria LBAE-K39 allowed the design of specific primers that could detect the presence of glucansucrase encoding genes similar to GTFKg3 of Lactobacillus fermentum Kg3 and to DSRWC of W. cibaria CMU. This study hence indicates that sourdough Weissella strains synthesize original dextransucrase.
TL;DR: A novel carbohydrate-sensing mechanism is suggested in C. thermocellum, whereby the presence of polysaccharide biomass components is detected extracellularly and the signal is transmitted intracellularly, resulting in the disruption of the interaction between RsgI-like proteins and sigma(I)-like factors, the latter of which serve to activate appropriate genes encoding proteins involved in cellulose utilization.
Abstract: Genome analysis of the Gram-positive cellulolytic bacterium Clostridium thermocellum revealed the presence of multiple negative regulators of alternative σ factors. Nine of the deduced proteins share a strong similarity in their N-terminal sequences to the Bacillus subtilis membrane-associated anti-σI factor RsgI and have an unusual domain organization. In six RsgI-like proteins, the C-terminal sequences contain predicted carbohydrate-binding modules. Three of these modules were overexpressed and shown to bind specifically to cellulose and/or pectin. Bioinformatic analysis of >1200 bacterial genomes revealed that the C. thermocellum RsgI-like proteins are unique to this species and are not present in other cellulolytic clostridial species (e.g. Clostridium cellulolyticum and Clostridium papyrosolvens ). Eight of the nine genes encoding putative C. thermocellum RsgI-like anti-σ factors form predicted bicistronic operons, in which the first gene encodes a putative alternative σ factor, similar to B. subtilis σI, but lacking in one of its domains. These observations suggest a novel carbohydrate-sensing mechanism in C. thermocellum , whereby the presence of polysaccharide biomass components is detected extracellularly and the signal is transmitted intracellularly, resulting in the disruption of the interaction between RsgI-like proteins and σI-like factors, the latter of which serve to activate appropriate genes encoding proteins involved in cellulose utilization.
TL;DR: Within the diverse bacterial communities found in reed roots, endophytic strains might have a strong potential to enhance phytoremediation by reed wetlands, and this study demonstrates that.
Abstract: The community structure and diversity of endophytic bacteria in reed (Phragmites australis) roots growing in the Beijing Cuihu Wetland, China was investigated using the 16S rRNA library technique. Primers 799f and 1492r were used to amplify the specific bacterial 16S rRNA fragments successfully and construct the clone library. In total, 166 individual sequences were verified by colony PCR and used to assess the diversity of endophytic bacteria in reed roots. Phylogenetic analysis revealed that 78.9% of the clones were affiliated with Proteobacteria and included all five classes. Other clones belonged to Firmicutes (9.0%), Cytophaga/Flexibacter/Bacteroids (6.6%), Fusobacteria (2.4%), and nearly 3.0% were unidentified bacteria. In Proteobacteria, the Alpha and Gamma subgroups were the most abundant, accounting for approximately 34.4% and 31.3% of all Proteobacteria, respectively, and the dominant genera included Pleomorphomonas, Azospirillum, and Aeromonas. In addition, nearly 13.6% of the Proteobacteria were very similar to some genera of sulfate-reducing bacteria (SRB) such as Dechloromonas, Desulfovibrio, and Sulfurospirillum. The bacteria in these genera are considered to play important roles in the metabolism of nitrogen, phosphorus, sulfur, and some organic compounds in wetland systems. Hence, this study demonstrates that within the diverse bacterial communities found in reed roots, endophytic strains might have a strong potential to enhance phytoremediation by reed wetlands.
TL;DR: MtrC and MtrF were shown to be potent reductases of chelated ferric iron, birnessite, and a carbon anode in a microbial fuel cell and OmcA-producing cells were unable to catalyze iron and electrode reduction, although the protein was correctly produced and oriented.
Abstract: The formation of outer membrane (OM) cytochromes seems to be a key step in the evolution of dissimilatory iron-reducing bacteria. They are believed to be the endpoints of an extended respiratory chain to the surface of the cell that establishes the connection to insoluble electron acceptors such as iron or manganese oxides. The gammaproteobacterium Shewanella oneidensis MR-1 contains the genetic information for five putative OM cytochromes. In this study, the role and specificity of these proteins were investigated. All experiments were conducted using a markerless deletion mutant in all five OM cytochromes that was complemented via the expression of single, plasmid-encoded genes. MtrC and MtrF were shown to be potent reductases of chelated ferric iron, birnessite, and a carbon anode in a microbial fuel cell. OmcA-producing cells were unable to catalyze iron and electrode reduction, although the protein was correctly produced and oriented. However, OmcA production resulted in a higher birnessite reduction rate compared with the mutant. The presence of the decaheme cytochrome SO_2931 as well as the diheme cytochrome SO_1659 did not rescue the phenotype of the deletion mutant.
TL;DR: It is shown that dietary fibre from almond skins altered the composition of gut bacteria and almond skins resulting from industrial blanching could be used as potential prebiotics.
Abstract: In this study we investigated the potential prebiotic effect of natural (NS) and blanched (BS) almond skins, the latter being a byproduct of the almond-processing industry. A full model of the gastrointestinal tract, including in vitro gastric and duodenal digestion, followed by colonic fermentation using mixed faecal bacterial cultures, was used. Both NS and BS significantly increased the population of bifidobacteria and Clostridium coccoides/Eubacterium rectale group, resulting in a prebiotic index (3.2 for BS and 3.3 for NS) that compared well with the commercial prebiotic fructo-oligosaccharides (4.2) at a 24-h incubation. No significant differences in the proportion of gut bacteria groups and in short-chain fatty acid production were detected between NS and BS, showing that polyphenols present in almond skins did not affect bacterial fermentation. In conclusion, we have shown that dietary fibre from almond skins altered the composition of gut bacteria and almond skins resulting from industrial blanching could be used as potential prebiotics.
TL;DR: This review summarizes the current knowledge on mycobacterial respiratory energy conversion, in particular, during the physiologically dormant state that is associated with latent or persistent tuberculosis infections.
Abstract: Mycobacterium tuberculosis, the causative agent of tuberculosis, poses a global health challenge due to the emergence of drug-resistant strains Recently, bacterial energy metabolism has come into focus as a promising new target pathway for the development of antimycobacterial drugs This review summarizes our current knowledge on mycobacterial respiratory energy conversion, in particular, during the physiologically dormant state that is associated with latent or persistent tuberculosis infections Targeting components of respiratory ATP production, such as type-2 NADH dehydrogenase or ATP synthase, is illustrated as an emerging strategy in the development of novel drugs