Subscribe: pubmed: 0099-2240
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pubmed: 0099-2240



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Differences in temperature and water chemistry shape distinct diversity patterns in thermophilic microbial communities.

Differences in temperature and water chemistry shape distinct diversity patterns in thermophilic microbial communities.

Appl Environ Microbiol. 2017 Aug 18;:

Authors: Chiriac CM, Szekeres E, Rudi K, Baricz A, Hegedus A, Dragoş N, Coman C

Abstract
This study presents the biodiversity and ecology of microbial mats developed in thermal gradients (20-65 °C) in the surroundings of three drillings (CH, CI, MB) tapping a hyperthermal aquifer from Romania. Using a metabarcoding approach, 16S rRNA genes were sequenced from both DNA and RNA transcripts (cDNA) and compared. The relationships between the microbial diversity and the physico-chemical factors were explored. Additionally, the cDNA data was used for in silico functionality predictions, bringing new insights into the functional potential and dynamics of these communities. The results showed that each hot spring determined the formation of distinct microbial communities. In the CH mats (40-53 °C), the abundance of Cyanobacteria decreased with temperature, opposite to that of Chloroflexi and Proteobacteria. Ectothiorhodospira, Oscillatoria and methanogenic archaea dominated the CI communities (20-65 °C), while the MB microbial mats (53-65 °C) were mainly composed of Chloroflexi, Hydrogenophilus, Thermi and Aquificae. Alpha diversity was negatively correlated with the increase in water temperature, while beta diversity was shaped in each hot spring by the unique combination of physico-chemical parameters, regardless of the type of nucleic acid analyzed (DNA vs. cDNA). The rank correlation analysis revealed a unique model that associated environmental data with community composition, consisting in the combined effect of Na(+), K(+), HCO3(-), PO4(3-) concentrations, together with temperature and electrical conductivity. These factors seem to determine the grouping of samples according to location, rather than with the similarities in thermal regimes, showing that other parameters beside temperature alone are significant drivers of biodiversity.Importance Hot spring microbial mats are remarkable manifestation of life on Earth, and were intensively studied for decades. Moreover, as hot spring areas are isolated and have a limited exchange of organisms, nutrients and energy with the surrounding environments, hot spring microbial communities can be used as model studies to elucidate the colonizing potential within extreme settings. Thus, they are of great importance in evolutionary biology, microbial ecology and exobiology. With all the efforts that have been made, the current understanding of the influence of temperature and water chemistry on the microbial community composition, diversity and abundance in microbial mats is limited. In this study, the composition and diversity of microbial communities developed in thermal gradients in the vicinity of three hot springs from Romania were investigated, each geothermal water having particular physico-chemical characteristics. Our results expose new factors that could determine the formation of these ecosystems, expanding the current knowledge in this regard.

PMID: 28821552 [PubMed - as supplied by publisher]




Coordinated Regulation of the EII(Man) and fruRKI Operons of Streptococcus mutans by Global and Fructose-Specific Pathways.

Coordinated Regulation of the EII(Man) and fruRKI Operons of Streptococcus mutans by Global and Fructose-Specific Pathways.

Appl Environ Microbiol. 2017 Aug 18;:

Authors: Zeng L, Chakraborty B, Farivar T, Burne RA

Abstract
The glucose/mannose-PTS permease EII(Man) encoded by manLMN in the dental caries pathogen Streptococcus mutans has a dominant influence on sugar-specific, CcpA-independent catabolite repression (CR). Mutations in manL affect energy metabolism and virulence-associated traits, including biofilm formation, acid tolerance and competence. Using promoter:reporter fusions, expression of the manLMN and the fruRKI operons, encoding a transcriptional regulator, a fructose-1-P kinase and a fructose-PTS permease EII(Fru), respectively, was monitored in response to carbohydrate source and in mutants lacking CcpA, FruR, and components of EII(Man) Expression of genes for EII(Man) and EII(Fru) was directly regulated by CcpA and CR, as evinced by in vivo and in vitro methods. Unexpectedly, not only was the fruRKI operon negatively regulated by FruR, but so was manLMN Carbohydrate transport by EII(Man) had a negative influence on expression of manLMN, but not fruRKI In agreement with the proposed role of FruR in regulating these PTS operons, loss of fruR or fruK substantially altered growth on a number of carbohydrates, including fructose. RNA deep sequencing revealed profound changes in gene regulation caused by deletion of fruK or fruR Collectively, these findings demonstrate intimate interconnection of the regulation of two major PTS permeases in S. mutans and reveal novel and important contributions of fructose metabolism to global regulation of gene expression.IMPORTANCE. The ability of Streptococcus mutans and other streptococcal pathogens to survive and cause human diseases is directly dependent upon their capacity to metabolize a variety of carbohydrates, including glucose and fructose. Our research reveals that metabolism of fructose has broad influences on the regulation of utilization of glucose and other sugars, and mutants in certain genes involved in fructose metabolism display profoundly different abilities to grow and express virulence-related traits. Mutants lacking the FruR regulator or a particular phosphofructokinase, FruK, display changes in expression of a large number of genes encoding transcriptional regulators; enzymes required for energy metabolism, biofilm development, biosynthetic and degradative processes; and tolerance of a spectrum of environmental stressors. Since fructose is a major component of the modern human diet, the results have substantial significance in the context of oral health and the development of dental caries.

PMID: 28821551 [PubMed - as supplied by publisher]




Crystal structure of the TetR family repressor, AlkX, from Dietzia sp. DQ12-45-1b implicated in biodegradation of n-alkanes.

Crystal structure of the TetR family repressor, AlkX, from Dietzia sp. DQ12-45-1b implicated in biodegradation of n-alkanes.

Appl Environ Microbiol. 2017 Aug 18;:

Authors: Liang JL, Gao Y, He Z, Nie Y, Wang M, JiangYang JH, Zhang XC, Shu WS, Wu XL

Abstract
n-Alkanes are ubiquitous in nature and are widely used by microorganisms as carbon sources. Alkane hydroxylation by alkane monooxygenases is a critical step in the aerobic biodegradation of n-alkanes, which plays important roles in natural alkane attenuation and is used in industrial and environmental applications. The alkane oxidation operon, alkW1-alkX, in an alkane-degrading strain Dietzia sp. DQ12-45-1b, is negatively auto-regulated by the TetR family repressor, AlkX, via a product positive feedback mechanism. To predict the gene regulation mechanism, we determined the 3.1 Å crystal structure of an AlkX homodimer in a non-DNA-bound state. The structure showed traceable long electron density deep inside a hydrophobic cavity of each monomer along the long axis of the helix bundle, and further gas chromatography-mass spectrometry analysis of AlkX revealed that it contained the E.coli-derived long-chain fatty acid molecules as ligand. Moreover, an unusual structural feature of AlkX is an extra helix, α6' , forming a lid-like structure with α6 covering the inducer-binding pocket and occupying the space between the two symmetrical DNA-binding motifs in one dimer, indicating a distinct conformational transition mode in modulating DNA binding. Sequence alignment of AlkX homologs from Dietzia strains showed that the residues involved in DNA and inducer binding are highly conserved, suggesting that the regulation mechanisms of n-alkane hydroxylation is possibly a common characteristic of Dietzia strains.Importance With ubiquitous n-alkanes in nature, many bacteria from terrestrial and aquatic environments have evolved n-alkane oxidation functions. Alkane hydroxylation by alkane monooxygenases is a critical step in the aerobic biodegradation of n-alkanes, which plays important roles in natural alkane attenuation and petroleum-contaminating environment bioremediation. The gene regulation of the most common alkane hydroxylase AlkB has been studied widely in Gram-negative bacteria, but less explored in Gram-positive bacteria. Our previous study showed that the TetR family regulator (TFR), AlkX, negatively auto-regulated the alkane oxidation operon, alkW1-alkX, in a Gram-positive strain Dietzia sp. DQ12-45-1b. As one of the most common transcriptional regulator family in bacteria, the TFR involving in n-alkane metabolism has been reported only recently. In this study, we determined the crystal structure of AlkX, which implies a distinct DNA/ligand binding mode. Our results shed light upon the regulation mechanism of common alkane degradation process in nature.

PMID: 28821550 [PubMed - as supplied by publisher]




Phosphate-Catalyzed Hydrogen Peroxide Formation from Agar, Gellan, and κ-Carrageenan and Recovery of Microorganisms Cultivability by Catalase and Pyruvate.

Phosphate-Catalyzed Hydrogen Peroxide Formation from Agar, Gellan, and κ-Carrageenan and Recovery of Microorganisms Cultivability by Catalase and Pyruvate.

Appl Environ Microbiol. 2017 Aug 18;:

Authors: Kawasaki K, Kamagata Y

Abstract
Previously, we reported that when agar is autoclaved with phosphate buffer, H2O2 is formed in the resulting medium (PT) and the colony count on the medium inoculated with environmental samples becomes much lower than that on a medium in which agar and phosphate are autoclaved separately (PS) (Appl Environ Microbiol 80:7659-7666). However, the physicochemical mechanisms underlying this observation remain largely unknown. Here, we determined the factors affecting H2O2 formation in agar. The H2O2 formation was pH-dependent: H2O2 was formed at high concentrations in alkaline or neutral phosphate buffer, but not in acidic one. Ammonium ions enhanced H2O2 formation, implying the involvement of the Maillard reaction catalyzed by phosphate. We found that other gelling agents (e.g., gellan and κ-carrageenan) also produced H2O2 after being autoclaved with phosphate. We then examined cultivability of microorganisms from a fresh water sample to test whether catalase and pyruvate known as H2O2 scavenger are effective in yielding high colony counts. The colony count on PT was only 5.7% of that on PS. Catalase treatment effectively restored the colony count of PT (to 106% of that on PS). By contrast, pyruvate was not as much effective as catalase: the colony count on sodium pyruvate-supplemented PT was 58% of that on PS. Given that both catalase and pyruvate can remove H2O2 from PT, these observations indicate that although H2O2 is the main cause of reduced colony count on PT, other unknown growth-inhibiting substances that cannot be removed by pyruvate (but can be by catalase) may also be involved.IMPORTANCE The majority of bacteria in natural environments are recalcitrant to laboratory culture techniques. Previously, we demonstrated that one reason for this is the formation of high H2O2 in the medium prepared by autoclaving agar and phosphate buffer together (PT medium). In this study we investigated the factors affecting H2O2 formation from agar. H2O2 formation is pH-dependent and ammonium ions promote this phosphate-catalyzed H2O2 formation. Amendment of catalase or pyruvate, a well-known H2O2-scavenging agent, effectively eliminated H2O2 Yet results suggest that growth-inhibiting factor(s) that cannot be eliminated by pyruvate (but can be by catalase) are present in PT medium.

PMID: 28821549 [PubMed - as supplied by publisher]




The transcriptional regulator PhlH modulates 2,4-diacetylphloroglucinol biosynthesis in response to the biosynthetic intermediate and end product.

The transcriptional regulator PhlH modulates 2,4-diacetylphloroglucinol biosynthesis in response to the biosynthetic intermediate and end product.

Appl Environ Microbiol. 2017 Aug 18;:

Authors: Yan X, Yang R, Zhao RX, Han JT, Jia WJ, Li DY, Wang Y, Zhang N, Wu Y, Zhang LQ, He YX

Abstract
Certain strains of biocontrol Pseudomonas fluorescens produce the secondary metabolite 2,4-diacetylphloroglucinol (2,4-DAPG) to antagonize the soil-borne phytopathogens in the rhizosphere. The gene cluster responsible for the biosynthesis of 2,4-DAPG is named phlACBDEFGH and it is still unclear how the pathway-specific regulator phlH within this gene cluster regulates the metabolism of 2,4-DAPG. Here, we found that PhlH in Pseudomonas fluorescens strain 2P24 represses the expression of the phlG gene encoding the 2,4-DAPG hydrolase by binding to a sequence motif overlapping with the -35 site recognized by σ(70) factors. Through biochemical screening of PhlH ligands we identified the end product 2,4-DAPG and its biosynthetic intermediate monoacetylphloroglucinol (MAPG) can act as signaling molecules to modulate the binding of PhlH to the target sequence and activate the expression of phlG Comparison of 2,4-DAPG production between the ΔphlH, ΔphlG and ΔphlHG mutants confirmed that phlH and phlG impose negative feedback regulation over 2,4-DAPG biosynthesis. It was further demonstrated that the 2,4-DAPG degradation catalyzed by PhlG plays an insignificant role in 2,4-DAPG tolerance, but contributes to bacterial growth advantages in carbon/nitrogen starvation conditions. Taken together, our data suggests that by monitoring and down-tuning in situ levels of 2,4-DAPG, the phlHG genes could dynamically modulate the metabolic loads attributed to 2,4-DAPG production and potentially contribute to the rhizosphere adaptation.Importance 2,4-DAPG synthesized by biocontrol pseudomonads is a broad-spectrum antibiotic against bacteria, fungi, oomycetes and nematodes and plays an important role in suppressing the soil-borne plant pathogens. Although most of the genes in the 2,4-DAPG biosynthetic gene cluster (phl) were characterized, it is still not clear how the pathway-specific regulator phlH is involved in 2,4-DAPG metabolism. This work revealed the role of PhlH in modulating 2,4-DAPG levels by controlling the expression of 2,4-DAPG hydrolase PhlG in response to 2,4-DAPG and MAPG. Since 2,4-DAPG biosynthesis imposes a metabolic burden to biocontrol pseudomonads, it is expected that the fine regulation of phlG by PhlH offers a way to dynamically modulate the metabolic loads attributed to 2,4-DAPG production.

PMID: 28821548 [PubMed - as supplied by publisher]




Cultivation in space flight produces minimal alterations in the susceptibility of Bacillus subtilis cells to 72 different antibiotics and growth-inhibiting compounds.

Cultivation in space flight produces minimal alterations in the susceptibility of Bacillus subtilis cells to 72 different antibiotics and growth-inhibiting compounds.

Appl Environ Microbiol. 2017 Aug 18;:

Authors: Morrison MD, Fajardo-Cavazos P, Nicholson WL

Abstract
Past results have suggested that bacterial antibiotic susceptibility is altered during space flight. To test this notion, Bacillus subtilis cells were cultivated in matched hardware, medium, and environmental conditions either in spaceflight microgravity on the International Space Station, termed Flight (FL) samples, or at Earth-normal gravity, termed Ground Control (GC) samples. Susceptibility of FL and GC samples was compared to 72 antibiotics and growth-inhibitory compounds using the Omnilog Phenotype Microarray (PM) system. Only 9 compounds were identified by PM screening as exhibiting significant differences (P < 0.05, Student's t-test) in FL vs. GC samples: 6-mercaptopurine, cesium chloride, enoxacin, lomefloxacin, manganese (II) chloride, nalidixic acid, penimepicycline, rolitetracycline, and trifluoperazine. Testing of the same compounds by standard broth dilution assay did not reveal statistically significant differences in the IC50 values between FL and GC samples. The results indicate that the susceptibility of B. subtilis cells to a wide range of antibiotics and growth inhibitors is not dramatically altered by space flight.Importance: This study addresses a major concern of mission planners for human spaceflight, that bacteria accompanying astronauts on long-duration missions might develop a higher level of resistance to antibiotics due to exposure to the spaceflight environment. The results of this study do not support that notion.

PMID: 28821547 [PubMed - as supplied by publisher]




A supervised statistical learning approach for accurate Legionella pneumophila source attribution during outbreaks.

A supervised statistical learning approach for accurate Legionella pneumophila source attribution during outbreaks.

Appl Environ Microbiol. 2017 Aug 18;:

Authors: Buultjens AH, Chua KYL, Baines SL, Kwong J, Gao W, Cutcher Z, Adcock S, Ballard S, Schultz MB, Tomita T, Subasinghe N, Carter GP, Pidot SJ, Franklin L, Seemann T, Gonçalves Da Silva A, Howden BP, Stinear TP

Abstract
Public health agencies are increasingly relying on genomics during Legionnaires' disease investigations. However, the causative bacterium (Legionella pneumophila) has an unusual population structure with extreme temporal and spatial genome sequence conservation. Furthermore, Legionnaires' disease outbreaks can be caused by multiple L. pneumophila genotypes in a single source. These factors can confound cluster identification using standard phylogenomic methods. Here, we show that a statistical learning approach based onL. pneumophila core genome single nucleotide polymorphism (SNP) comparisons eliminates ambiguity for defining outbreak clusters and accurately predicts exposure sources for clinical cases. We illustrate the performance of our method by genome comparisons of 234 L. pneumophila isolates obtained from patients and cooling towers in Melbourne, Australia between 1994 and 2014. This collection included one of the largest reported Legionnaires' disease outbreaks, involving 125 cases at an aquarium. Using only sequence data from L. pneumophila cooling tower isolates and including all core genome variation, we built a multivariate model using discriminant analysis of principal components (DAPC) to find cooling tower-specific genomic signatures, and then used it to predict the origin of clinical isolates. Model assignments were 93% congruent with epidemiological data, including the aquarium Legionnaires' outbreak and three other unrelated outbreak investigations. We applied the same approach to a recently described investigation of Legionnaires' disease within a UK hospital and observed model predictive ability of 86%. We have developed a promising means to breach L. pneumophila genetic diversity extremes and provide objective source attribution data for outbreak investigations.Importance Microbial outbreak investigations are moving to a paradigm where whole genome sequencing and phylogenetic trees are used to support epidemiological investigations. It's critical that outbreak source predictions are accurate, particularly for pathogens like Legionella pneumophila, which can spread widely and rapidly via cooling system aerosols causing Legionnaires' disease. Here, by studying hundreds of Legionella pneumophila genomes collected over 21 years around a major Australian city, we uncovered limitations with the phylogenetic approach that could lead to misidentification of outbreak sources. We implement instead a statistical learning technique that eliminates the ambiguity of inferring disease transmission from phylogenies. Our approach takes geolocation information and core genome variation from environmental L. pneumophila isolates to build statistical models that predict with high confidence the environmental source of clinical L. pneumophila during disease outbreaks. We show the versatility of the technique by applying it to unrelated Legionnaires' disease outbreaks in Australia and the UK.

PMID: 28821546 [PubMed - as supplied by publisher]




Heterologous production of thermophile Thermochromatium tepidum photosynthetic reaction centre and light harvesting 1 complexes in the mesophile Rhodobacter sphaeroides and thermal stabilities of a hybrid core complex.

Heterologous production of thermophile Thermochromatium tepidum photosynthetic reaction centre and light harvesting 1 complexes in the mesophile Rhodobacter sphaeroides and thermal stabilities of a hybrid core complex.

Appl Environ Microbiol. 2017 Aug 18;:

Authors: Jun D, Huang V, Beatty JT

Abstract
The photosynthetic complexes of the thermophile Thermochromatium tepidum are of considerable interest in biohybrid solar cell applications because of the capability of thermophilic proteins to tolerate elevated temperatures. Synthetic operons encoding reaction centre (RC) and light harvesting 1 (LH1) pigment-protein complexes of T. tepidum were expressed in the mesophile Rhodobacter sphaeroides The T. tepidum RC (TRC) was assembled, and functional with the addition of menadione to populate the QA pocket. The production of the T. tepidum LH1 (TLH1) was increased by selection of a phototrophy-capable mutant after UV irradiation mutagenesis, which yielded a hybrid RC-TLH1 core complex consisting of the R. sphaeroides RC and T. tepidum TLH1, confirmed by the absorbance peak of TLH1 at 915 nm. Affinity chromatography partial purification and subsequent sucrose gradient analysis of the hybrid RC-TLH1 core complex indicated that this core complex assembled as a monomer. Furthermore, the RC-TLH1 hybrid core complex was more tolerant of 70 °C temperature than the R. sphaeroides RC-LH1 core complexes in both the dimeric and monomeric forms: after 1 h, the hybrid complex retained 58% of the initial starting value, compared to 11 and 53% for the R. sphaeroides RC-LH1 dimer and monomer forms, respectively.Importance This work is important because it is a new approach to bio-engineering of photosynthesis proteins for potential use in bio-photovoltaic solar energy capture. The work establishes proof-of-principle for future biohybrid solar cell applications.

PMID: 28821545 [PubMed - as supplied by publisher]