RESUMO
Systems characterization of immune landscapes in health, disease and clinical intervention cases is a priority in modern medicine. High-throughput transcriptomes accumulated from gene-knockout (KO) experiments are crucial for deciphering target KO signaling pathways that are impaired by KO genes at the systems-level. There is a demand for integrative platforms. This article describes the PathwayKO platform, which has integrated state-of-the-art methods of pathway enrichment analysis, statistics analysis, and visualizing analysis to conduct cutting-edge integrative pathway analysis in a pipeline fashion and decipher target KO signaling pathways at the systems-level. We focus on describing the methodology, principles and application features of PathwayKO. First, we demonstrate that the PathwayKO platform can be utilized to comprehensively analyze real-world mouse KO transcriptomes (GSE22873 and GSE24327), which reveal systemic mechanisms underlying the innate immune responses triggered by non-infectious extensive hepatectomy (2 hours after 85% liver resection surgery) and infectious CASP-model sepsis (12 hours after CASP-model surgery). Strikingly, our results indicate that both cases hit the same core set of 21 KO MyD88-associated signaling pathways, including the Toll-like receptor signaling pathway, the NFκB signaling pathway, the MAPK signaling pathway, and the PD-L1 expression and PD-1 checkpoint pathway in cancer, alongside the pathways of bacterial, viral and parasitic infections. These findings suggest common fundamental mechanisms between these immune responses and offer informative cues that warrant future experimental validation. Such mechanisms in mice may serve as models for humans and ultimately guide formulating the research paradigms and composite strategies to reduce the high mortality rates of patients in intensive care units who have undergone successful traumatic surgical treatments. Second, we demonstrate that the PathwayKO platform model-based assessments can effectively evaluate the performance difference of pathway analysis methods when benchmarked with a collection of proper transcriptomes. Together, such advances in methods for deciphering biological insights at the systems-level may benefit the fields of bioinformatics, systems immunology and beyond.
Assuntos
Fígado , Transdução de Sinais , Humanos , Camundongos , Animais , Transdução de Sinais/genética , Fígado/metabolismo , NF-kappa B/metabolismo , Imunidade Inata , TranscriptomaRESUMO
Bacteriophages are the most abundant entities in the biosphere, and many genomes of rare and novel bacteriophages have been sequenced to date. However, bacteriophage functional genomics has been limited by a lack of effective research methods. Clustered regularly interspaced short palindromic repeat/CRISPR-associated gene (CRISPR-Cas) systems provide bacteriophages with a new mechanism for attacking host bacteria as well as new tools for study bacteriophage functional genomics. It has been reported that bacteriophages are not only the driving elements of the evolution of prokaryote CRISPR arrays but also the targets of CRISPR-Cas systems. In this study, a phage genome editing platform based on the heterologous CRISPR-Cas9 system was theoretically designed, and a Vibrio natriegens phage TT4P2 genome editing experiment was carried out in vivo in the host bacterium Vibrio natriegens TT4 to achieve phage gene deletion and replacement. The construction of this phage genome editing platform is expected to solve the problem of insufficient research on phage gene diversity, promote the development of phage synthetic biology and nanotechnology, and even accelerate the discovery of new molecular biology tools. IMPORTANCE Bacteriophages are the most numerous organisms on earth and are known for their diverse lifestyles. Since the discovery of bacteriophages, our knowledge of the wider biological world has undergone immense and unforeseen changes. A variety of V. natriegens phages have been detected, but few have been well characterized. CRISPR was first documented in Escherichia coli in 1987. It has been reported that the CRISPR-Cas system can target and cleave invaders, including bacteriophages, in a sequence-specific manner. Here, we show that the construction of a phage genome editing platform based on the heterologous CRISPR-Cas9 system can achieve V. natriegens phage TT4P2 gene editing and can also improve the efficiency and accuracy of phage TT4P2 gene editing.
Assuntos
Bacteriófagos , Edição de Genes , Bacteriófagos/genética , Sistemas CRISPR-Cas , Escherichia coli/genética , Edição de Genes/métodos , VibrioRESUMO
Colon ascendens stent peritonitis (CASP) surgery induces a leakage of intestinal contents which may cause polymicrobial sepsis related to post-operative failure of remote multi-organs (including kidney, liver, lung and heart) and possible death from systemic syndromes. Mechanisms underlying such phenomena remain unclear. This article aims to elucidate the mechanisms underlying the CASP-model sepsis by analyzing real-world GEO data (GSE24327_A, B and C) generated from mice spleen 12 hours after a CASP-surgery in septic MyD88-deficient and wildtype mice, compared with untreated wildtype mice. Firstly, we identify and characterize 21 KO MyD88-associated signaling pathways, on which true key regulators (including ligands, receptors, adaptors, transducers, transcriptional factors and cytokines) are marked, which were coordinately, significantly, and differentially expressed at the systems-level, thus providing massive potential biomarkers that warrant experimental validations in the future. Secondly, we observe the full range of polymicrobial (viral, bacterial, and parasitic) sepsis triggered by the CASP-surgery by comparing the coordinated up- or down-regulations of true regulators among the experimental treatments born by the three data under study. Finally, we discuss the observed phenomena of "systemic syndrome", "cytokine storm" and "KO MyD88 attenuation", as well as the proposed hypothesis of "spleen-mediated immune-cell infiltration". Together, our results provide novel insights into a better understanding of innate immune responses triggered by the CASP-model sepsis in both wildtype and MyD88-deficient mice at the systems-level in a broader vision. This may serve as a model for humans and ultimately guide formulating the research paradigms and composite strategies for the early diagnosis and prevention of sepsis.
Assuntos
Peritonite , Sepse , Animais , Imunidade Inata , Camundongos , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/metabolismo , Peritonite/etiologia , Transdução de Sinais , Stents/efeitos adversosRESUMO
Identifying cancer-related miRNAs (or microRNAs) that precisely target mRNAs is important for diagnosis and treatment of cancer. Creating novel methods to identify candidate miRNAs becomes an imminent Frontier of researches in the field. One major obstacle lies in the integration of the state-of-the-art databases. Here, we introduce a novel method, MIMRDA, which incorporates the miRNA and mRNA expression profiles for predicting miRNA-disease associations to identify key miRNAs. As a proof-of-principle study, we use the MIMRDA method to analyze TCGA datasets of 20 types (BLCA, BRCA, CESE, CHOL, COAD, ESCA, HNSC, KICH, KIRC, KIRP, LIHC, LUAD, LUSC, PAAD, PRAD, READ, SKCM, STAD, THCA and UCEC) of cancer, which identified hundreds of top-ranked miRNAs. Some (as Category 1) of them are endorsed by public databases including TCGA, miRTarBase, miR2Disease, HMDD, MISIM, ncDR and mTD; others (as Category 2) are supported by literature evidences. miR-21 (representing Category 1) and miR-1258 (representing Category 2) display the excellent characteristics of biomarkers in multi-dimensional assessments focusing on the function similarity analysis, overall survival analysis, and anti-cancer drugs' sensitivity or resistance analysis. We compare the performance of the MIMRDA method over the Limma and SPIA packages, and estimate the accuracy of the MIMRDA method in classifying top-ranked miRNAs via the Random Forest simulation test. Our results indicate the superiority and effectiveness of the MIMRDA method, and recommend some top-ranked key miRNAs be potential biomarkers that warrant experimental validations.
RESUMO
To simplify industrial mushroom cultivation, we introduced a bacterial Pseudomonas sp. UW4 acdS gene, encoding 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase (AcdS), into fungus Agaricus bisporus. Transformant A. bisporus-acdS14 cased with sterilized-vermiculite generated primordia 5 days sooner than wild-type strain, confirming the specific role of the AcdS enzyme. Being consistent with the AcdS enzyme activity increased by 84%, the mycelium growth rate was increased by 25%; but, the ACC and ethylene concentrations were reduced by 71% and 36%, respectively, in the A. bisporus-acdS14 transformant. And the bacterium P. sp. UW4 attachment on the mycelium of the A. bisporus-acdS14 transformant was drastically reduced. We conclude that the heterogeneously expressed bacterial acdS gene degrades ACC and reduces ethylene-synthesis, eliminating ethylene inhibition on the mycelium growth and primordium formation in A. bisporus. Our results provide new insights into the mechanism underlying casing soil bacterium, and help formulate a casing-less cultivation for the next-generation mushroom industry.
Assuntos
Agaricus/crescimento & desenvolvimento , Agaricus/genética , Carpóforos/crescimento & desenvolvimento , Pseudomonas/enzimologia , Pseudomonas/genética , Aminoácidos Cíclicos/metabolismo , Carbono-Carbono Liases/genética , Carbono-Carbono Liases/metabolismo , Clonagem Molecular , Etilenos/metabolismo , Regulação Fúngica da Expressão Gênica , Micélio/crescimento & desenvolvimento , Solo , Transformação GenéticaRESUMO
Protease activity of allergens has been suggested to be involved in the pathogenesis of allergic diseases. The major allergen Der f 3 from Dermatophagoides farinae harbors serine protease activity, but its immunopathogenesis remains unclear. This study aims to explore the effect of Der f 3 on the airway epithelial barrier and on the molecular pathways by which Der f 3 induces inflammation. RNA-seq was performed to identify differentially expressed genes in bronchial airway epithelial cells (AEC) between native Der f 3 and heat-inactivated (H) Der f 3, coupled with real-time PCR (RT-PCR) and ELISA for validation. Unlike other protease allergens such as that induce Th2-promoting alarmins (IL-25, IL-33, TSLP) in AECs, Der f 3 induced pro-inflammatory cytokines and chemokines including IL-6, IL-8 and GM-CSF, which are known to promote Th17 response. These pro-inflammatory mediators were induced by Der f 3 via the MAPK and NF-κB pathways as well as the store-operated calcium signaling. Gene silencing with small interfering RNA in A549 and BEAS-2B cells indicated that activation of AECs by Der f 3 was mainly dependent on protease-activated receptor 2 (PAR-2), while PAR-1 was also required for the full activation of AECs. Double knock-down of PAR-1 and PAR-2 largely impaired Der f 3-inducecd IL-8 production and subsequent signaling pathways. Our data suggest that Der f 3 induces pro-inflammatory mediators in human epithelial cell lines via the PARs-MAPK-NF-κB axis. Our results provide a molecular mechanism by which Der f 3 may trigger the Th17-skewed allergic response toward house dust mites.
Assuntos
Alérgenos/imunologia , Proteínas de Artrópodes/imunologia , Células Epiteliais/imunologia , Pyroglyphidae/imunologia , Receptor PAR-1/imunologia , Receptor PAR-2/imunologia , Mucosa Respiratória/imunologia , Serina Endopeptidases/imunologia , Células A549 , Alérgenos/farmacologia , Animais , Proteínas de Artrópodes/farmacologia , Sinalização do Cálcio/efeitos dos fármacos , Sinalização do Cálcio/genética , Sinalização do Cálcio/imunologia , Citocinas/genética , Citocinas/imunologia , Células Epiteliais/patologia , MAP Quinases Reguladas por Sinal Extracelular/genética , MAP Quinases Reguladas por Sinal Extracelular/imunologia , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/imunologia , Técnicas de Silenciamento de Genes , Humanos , Inflamação/induzido quimicamente , Inflamação/genética , Inflamação/imunologia , Inflamação/patologia , Mediadores da Inflamação/imunologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/genética , Sistema de Sinalização das MAP Quinases/imunologia , NF-kappa B/genética , NF-kappa B/imunologia , Receptor PAR-1/genética , Receptor PAR-2/genética , Serina Endopeptidases/farmacologia , Células Th17/imunologia , Células Th17/patologia , Células Th2/imunologia , Células Th2/patologiaRESUMO
BACKGROUND: Mal f 1, the first allergen cloned from Malassezia furfur, has positive IgE reactivity in sera from atopic dermatitis (AD) patients. The mechanism by which Mal f 1 induces the maturation of human dendritic cells (DCs) and maintains the symptoms of AD is not well understood. OBJECTIVE: The present study aims to explore the activation profile of THP-1 derived dendritic cells (TDDCs) stimulated by recombinant Mal f 1, as well as to explore the IgE-binding ability of rMal f 1 and its correlation with IgE-binding activity of complete allergens of M. furfur. METHODS: rMal f 1 was produced by expression in E. coli and purification with affinity chromatography. The ability of rMal f 1 and ImmunoCAP complete allergens of M. furfur to bind to serum specific IgE was assayed in parallel by ELISA and immunoblotting. Immature TDDCs were stimulated with rMal f 1 or an enzyme-digested product of rMal f 1. The expression levels of markers, CD83, CD80, CD86, and HLA-DR, were investigated by flow cytometry. The levels of interleukin (IL)-6, IL-10, IL-12p70 and tumor necrosis factor (TNF)-α in culture supernatants were determined by ELISA. RESULTS: Eighteen patient sera were identified that reacted positively to the complete allergens of M. furfur as determined by ImmunoCAP and also showed positive responses to rMal f 1. Five patient sera were identified that had no reaction to ImmunoCAP complete allergens of M. furfur and also exhibited negative response to rMal f 1. All sera, except for one, had no reaction to the unrelated allergen Bet v 1. rMal f 1 upregulated the maturation surface marker CD83 on TDDCs. In addition, rMal f 1 also induced high levels of CD80 and CD86. Increased expression of HLA-DR, a first signal for T cell activation, was observed. Secretion of IL-6, TNF-α and IL-10 by TDDCs increased significantly (Pâ¯<â¯0.0001 for IL-6, Pâ¯<â¯0.01 for TNF-α and Pâ¯<â¯0.05 for IL-10) after stimulation by rMal f 1, while the IL-12p70 level was unaltered. CONCLUSION: We have shown that rMal f 1 has ideal IgE binding ability and good correlation with binding activity to M. furfur. Moreover, we have revealed a hitherto unknown DC activation profile after rMal f 1 stimulation whereby TNF-α, IL-6, and IL-10 were significantly increased and IL-12 was unaltered, suggesting that rMal f 1 can predispose a DC bias toward the TH22/TH17 pathway beyond the routine IgE-dependent TH2 pathway, thus providing intriguing clues for clinical treatment involving both pathways.
Assuntos
Alérgenos/imunologia , Proteínas Fúngicas/imunologia , Imunoglobulina E/imunologia , Células de Langerhans/imunologia , Adolescente , Adulto , Alérgenos/genética , Alérgenos/farmacologia , Antígenos CD/imunologia , Técnicas de Cultura de Células , Criança , Pré-Escolar , Dermatite Atópica/sangue , Dermatite Atópica/imunologia , Escherichia coli/genética , Feminino , Proteínas Fúngicas/genética , Proteínas Fúngicas/farmacologia , Humanos , Imunoglobulina E/sangue , Imunoglobulinas/imunologia , Células de Langerhans/efeitos dos fármacos , Malassezia/imunologia , Masculino , Glicoproteínas de Membrana/imunologia , Pessoa de Meia-Idade , Ligação Proteica , Proteínas Recombinantes , Células THP-1 , Adulto Jovem , Antígeno CD83RESUMO
Assessing correctness of an assembled chromosome architecture is a central challenge. We create a geometric analysis method (called GenomeLandscaper) to conduct landscape analysis of genome-fingerprints maps (GFM), trace large-scale repetitive regions, and assess their impacts on the global architectures of assembled chromosomes. We develop an alignment-free method for phylogenetics analysis. The human Y chromosomes (GRCh.chrY, HuRef.chrY and YH.chrY) are analysed as a proof-of-concept study. We construct a galaxy of genome-fingerprints maps (GGFM) for them, and a landscape compatibility among relatives is observed. But a long sharp straight line on the GGFM breaks such a landscape compatibility, distinguishing GRCh38p1.chrY (and throughout GRCh38p7.chrY) from GRCh37p13.chrY, HuRef.chrY and YH.chrY. We delete a 1.30-Mbp target segment to rescue the landscape compatibility, matching the antecedent GRCh37p13.chrY. We re-locate it into the modelled centromeric and pericentromeric region of GRCh38p10.chrY, matching a gap placeholder of GRCh37p13.chrY. We decompose it into sub-constituents (such as BACs, interspersed repeats, and tandem repeats) and trace their homologues by phylogenetics analysis. We elucidate that most examined tandem repeats are of reasonable quality, but the BAC-sized repeats, 173U1020C (176.46 Kbp) and 5U41068C (205.34 Kbp), are likely over-repeated. These results offer unique insights into the centromeric and pericentromeric regions of the human Y chromosomes.
Assuntos
Mapeamento Cromossômico/métodos , Biologia Computacional/métodos , Impressões Digitais de DNA/métodos , Genômica/métodos , Software , Algoritmos , Bases de Dados GenéticasRESUMO
Ethylene biosynthesis and function in Agaricus bisporus (the button mushroom) remain uncertain. The enzyme activities of 1-aminocyclopropane-1-carboxylate (ACC) synthase (ACS) and ACC oxidase (ACO) were detectable in A. bisporus AS2796 and inhibited by α-aminooxyacetic acid and Co2+. We cloned and sequenced 2 ACS genes (Ab-ACS1 and Ab-ACS2) and 1 ACO gene (Ab-ACO) from the mushroom strain. Ab-ACS1 and Ab-ACS2 demonstrated low amino acid sequence similarity. Ab-ACO demonstrated an amino acid sequence completely identical to that of ACO1_AGABI from A. bisporus. Antisense ACO significantly reduced ACO gene expression level, ACO enzyme activity, and ethylene production in the mushroom transformants. The transformants grew faster than the wild-type strain in sterilized compost and normally formed primordia when cultivated in sterilized compost with the sterilized casing vermiculite, but the wild-type strain did not. Our results show that ethylene is synthesized in button mushrooms via the ACC pathway. Ethylene inhibited button mushroom mycelial growth and development.
Assuntos
Agaricus/crescimento & desenvolvimento , Agaricus/metabolismo , Aminoácido Oxirredutases/metabolismo , Etilenos/metabolismo , Proteínas Fúngicas/metabolismo , Liases/metabolismo , Micélio/crescimento & desenvolvimento , Aminoácido Oxirredutases/genética , Clonagem Molecular , Proteínas Fúngicas/genética , Expressão Gênica , Liases/genética , Análise de Sequência de DNA , Homologia de Sequência de AminoácidosRESUMO
BACKGROUND: No attention has been paid on comparing a set of genome sequences crossing genetic components and biological categories with far divergence over large size range. We define it as the systematic comparative genomics and aim to develop the methodology. RESULTS: First, we create a method, GenomeFingerprinter, to unambiguously produce a set of three-dimensional coordinates from a sequence, followed by one three-dimensional plot and six two-dimensional trajectory projections, to illustrate the genome fingerprint of a given genome sequence. Second, we develop a set of concepts and tools, and thereby establish a method called the universal genome fingerprint analysis (UGFA). Particularly, we define the total genetic component configuration (TGCC) (including chromosome, plasmid, and phage) for describing a strain as a systematic unit, the universal genome fingerprint map (UGFM) of TGCC for differentiating strains as a universal system, and the systematic comparative genomics (SCG) for comparing a set of genomes crossing genetic components and biological categories. Third, we construct a method of quantitative analysis to compare two genomes by using the outcome dataset of genome fingerprint analysis. Specifically, we define the geometric center and its geometric mean for a given genome fingerprint map, followed by the Euclidean distance, the differentiate rate, and the weighted differentiate rate to quantitatively describe the difference between two genomes of comparison. Moreover, we demonstrate the applications through case studies on various genome sequences, giving tremendous insights into the critical issues in microbial genomics and taxonomy. CONCLUSIONS: We have created a method, GenomeFingerprinter, for rapidly computing, geometrically visualizing, intuitively comparing a set of genomes at genome fingerprint level, and hence established a method called the universal genome fingerprint analysis, as well as developed a method of quantitative analysis of the outcome dataset. These have set up the methodology of systematic comparative genomics based on the genome fingerprint analysis.
Assuntos
Impressões Digitais de DNA , Genoma Bacteriano , Genoma Viral , Genoma , Genômica , Bactérias/genética , Mapeamento Cromossômico , Modelos Teóricos , Análise de Sequência de DNA , Vírus/genéticaRESUMO
The artificial neural network (ANN) and genetic algorithm (GA) were combined to optimize the fermentation process for enhancing production of marine bacteriocin 1701 in a 5-L-stirred-tank. Fermentation time, pH value, dissolved oxygen level, temperature and turbidity were used to construct a "5-10-1" ANN topology to identify the nonlinear relationship between fermentation parameters and the antibiotic effects (shown as in inhibition diameters) of bacteriocin 1701. The predicted values by the trained ANN model were coincided with the observed ones (the coefficient of R(2) was greater than 0.95). As the fermentation time was brought in as one of the ANN input nodes, fermentation parameters could be optimized by stages through GA, and an optimal fermentation process control trajectory was created. The production of marine bacteriocin 1701 was significantly improved by 26% under the guidance of fermentation control trajectory that was optimized by using of combined ANN-GA method.
Assuntos
Bacteriocinas/biossíntese , Biotecnologia/métodos , Fermentação , Redes Neurais de Computação , Algoritmos , Análise de Variância , Bacteriocinas/economia , Reatores Biológicos/microbiologia , Oxigênio/metabolismo , Reprodutibilidade dos Testes , Fatores de TempoRESUMO
The phage display technology (PDT) was unique in genetic engineering and recombinant expression. The phage display systems (PDS) were platforms (kits) composed of genetic modified phages, helper phages, and host bacteria. This review concisely summarized the development of four types of PDS, based on M13, λ, T4, and T7 phages, in terms of phage molecular genetics and genetic (gene or genome) engineering. We addressed on the key components and their genetic (genomic) engineering for modifications, the technical features of different anchors, and the development progress and selection reference of those different kits.
Assuntos
Engenharia Genética , Biblioteca de Peptídeos , Bacteriófago M13/genética , Bacteriófago T4/genética , Bacteriófago T7/genética , Bacteriófago lambda/genéticaRESUMO
With PCR-denaturing gradient gel electrophoresis analysis, two bacterial 16S rRNA gene V3 region sequences, 7A and 7B, were detected in association with the crushed spores of the arbuscular mycorrhizal fungus Gigaspora margarita W.N. Becker & I.R. Hall 1976 MAFF520054. DNA sequencing and phylogenetic analysis revealed that 7B was mostly related to the documented cytoplasm endosymbiotic bacterium Candidatus Glomeribacter gigasporarum of G. margarita, but 7A could not be confidently assigned to a known taxon. Further characterization of 7A was conducted by obtaining its almost complete 16S rRNA gene sequence via PCR amplification and sequencing. BLAST search indicates that the 16S rRNA gene sequence did not match any identified species sequences in the GenBank database. Further detection revealed that 7A was also associated with the clean G. margarita MAFF520054 spores that were obtained by the surface-sterilized method or dual culture with Ri T-DNA transformed carrot roots. Many ellipse-shaped or egg-shaped bacterium-like organisms were clustered in layer 3 of the fungal spore wall by transmission electron microscopy observation. Our results indicate that 7A represents a novel bacterial population associated with G. margarita MAFF520054 spores, and its doubtless location (wall or cytoplasm) remains unclear based on the present data.
Assuntos
Bactérias/isolamento & purificação , Glomeromycota/fisiologia , Micorrizas/fisiologia , Raízes de Plantas/microbiologia , Bactérias/classificação , Bactérias/genética , Fenômenos Fisiológicos Bacterianos , DNA Bacteriano/genética , DNA Ribossômico/genética , Glomeromycota/química , Dados de Sequência Molecular , Micorrizas/química , Filogenia , RNA Ribossômico 16S/genética , Esporos Fúngicos/química , Esporos Fúngicos/fisiologia , SimbioseRESUMO
OBJECTIVE: We analyzed bacterial colonization associated with spores of arbuscular mycorrhizal fungi (AMF) Gigaspora margarita, to indicate their ecological niche, and to provide information for further researches on their populations or functions. METHODS: Six bacteria strains (Peanibacillus sp. M060106-1, Peanibacillus sp. M061122-2, Peanibacillus sp. M061122-6, Bacillus sp. M061122-4, Bacillus sp. M061122-10 and Brevibacillus sp. M061122-12) isolated from G. margarita spores were tagged with green fluorescence protein (GFP) using the carrier plasmid pNF8 (gfp-mut1). We analyzed the ecological niche and population dynamics of tagged strains on G. margarita under different conditions by using fluorescent microscope and/or plate counts. RESULTS: Four strains (M060106-1, M061122-6, M061122-10 and M061122-12) were tagged with GFP, showing high plasmid stability. These tagged strains possessed the basic characteristics identical to their original strains and, hence, were fit for short-term study of environmental colonization. All four GFP-tagged strains colonized the spore wall of G. margarita, and M061122-6 and M061122-12 further colonized the fungal hyphae. Under different pH conditions,the population dynamic of each GFP-tagged strain on the spores showed the same trend, i.e. first increased and then decreased, and the effects on the population size varied with different pH value. GFP-tagged strains colonized the spores of low viability more easily than those of high viability, and the population dynamic on the spores of high viability was different for each tagged strain. CONCLUSION: The isolated bacteria associated with G. margarita spores can re-colonize the fungal spores, whereas their colonizing ability depends on their characteristics and environmental factors. These data contributes to the further understanding of populations and functions of AMF-associated bacteria.
Assuntos
Bactérias/metabolismo , Técnicas de Laboratório Clínico , Glomeromycota/fisiologia , Proteínas de Fluorescência Verde/metabolismo , Esporos Fúngicos/fisiologia , Bactérias/genética , Plasmídeos de Bacteriocinas/genética , Proteínas de Fluorescência Verde/genéticaRESUMO
Submarine hydrothermal vents are among the least-understood habitats on Earth but have been the intense focus of research in the past 30 years. An active hydrothermal sulfide chimney collected from the Dudley site in the Main Endeavour vent Field (MEF) of Juan de Fuca Ridge was investigated using mineralogical and molecular approaches. Mineral analysis indicated that the chimney was composed mainly of Fe-, Zn-and Cu-rich sulfides. According to phylogenetic analysis, within the Crenarchaeota, clones of the order Desulfurococcales predominated, comprising nearly 50% of archaeal clones. Euryarchaeota were composed mainly of clones belonging to Thermococcales and deep-sea hydrothermal vent Euryarchaeota (DHVE), each of which accounted for about 20% of all clones. Thermophilic or hyperthermophilic physiologies were common to the predominant archaeal groups. More than half of bacterial clones belonged to epsilon-Proteobacteria, which confirmed their prevalence in hydrothermal vent environments. Clones of Proteobacteria (gamma-, delta-, beta-), Cytophaga-Flavobacterium-Bacteroides (CFB) and Deinococcus-Thermus occurred as well. It was remarkable that methanogens and methanotrophs were not detected in our 16S rRNA gene library. Our results indicated that sulfur-related metabolism, which included sulfur-reducing activity carried out by thermophilic archaea and sulfur-oxidizing by mesophilic bacteria, was common and crucial to the vent ecosystem in Dudley hydrothermal site.