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1.
Allergy ; 2024 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-38932655

RESUMO

BACKGROUND: The pathological mechanism of the gastrointestinal forms of food allergies is less understood in comparison to other clinical phenotypes, such as asthma and anaphylaxis Importantly, high-IgE levels are a poor prognostic factor in gastrointestinal allergies. METHODS: This study investigated how high-IgE levels influence the development of intestinal inflammation and the metabolome in allergic enteritis (AE), using IgE knock-in (IgEki) mice expressing high levels of IgE. In addition, correlation of the altered metabolome with gut microbiome was analysed. RESULTS: Ovalbumin-sensitized and egg-white diet-fed (OVA/EW) BALB/c WT mice developed moderate AE, whereas OVA/EW IgEki mice induced more aggravated intestinal inflammation with enhanced eosinophil accumulation. Untargeted metabolomics detected the increased levels of N-tau-methylhistamine and 2,3-butanediol, and reduced levels of butyric acid in faeces and/or sera of OVA/EW IgEki mice, which was accompanied with reduced Clostridium and increased Lactobacillus at the genus level. Non-sensitized and egg-white diet-fed (NC/EW) WT mice did not exhibit any signs of AE, whereas NC/EW IgEki mice developed marginal degrees of AE. Compared to NC/EW WT mice, enhanced levels of lysophospholipids, sphinganine and sphingosine were detected in serum and faecal samples of NC/EW IgEki mice. In addition, several associations of altered metabolome with gut microbiome-for example Akkermansia with lysophosphatidylserine-were detected. CONCLUSIONS: Our results suggest that high-IgE levels alter intestinal and systemic levels of endogenous and microbiota-associated metabolites in experimental AE. This study contributes to deepening the knowledge of molecular mechanisms for the development of AE and provides clues to advance diagnostic and therapeutic strategies of allergic diseases.

2.
Int J Mol Sci ; 24(11)2023 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-37298198

RESUMO

Cow's milk allergy (CMA) is one of the most prevalent food allergies in children. Several studies have demonstrated that gut microbiota influences the acquisition of oral tolerance to food antigens at initial stages of life. Changes in the gut microbiota composition and/or functionality (i.e., dysbiosis) have been linked to inadequate immune system regulation and the emergence of pathologies. Moreover, omic sciences have become an essential tool for the analysis of the gut microbiota. On the other hand, the use of fecal biomarkers for the diagnosis of CMA has recently been reviewed, with fecal calprotectin, α-1 antitrypsin, and lactoferrin being the most relevant. This study aimed at evaluating functional changes in the gut microbiota in the feces of cow's milk allergic infants (AI) compared to control infants (CI) by metagenomic shotgun sequencing and at correlating these findings with the levels of fecal biomarkers (α-1 antitrypsin, lactoferrin, and calprotectin) by an integrative approach. We have observed differences between AI and CI groups in terms of fecal protein levels and metagenomic analysis. Our findings suggest that AI have altered glycerophospholipid metabolism as well as higher levels of lactoferrin and calprotectin that could be explained by their allergic status.


Assuntos
Microbioma Gastrointestinal , Hipersensibilidade a Leite , Feminino , Animais , Bovinos , Leite/química , Lactoferrina/metabolismo , Hipersensibilidade a Leite/diagnóstico , Fezes/química , Biomarcadores/análise
3.
J Antimicrob Chemother ; 77(3): 719-726, 2022 02 23.
Artigo em Inglês | MEDLINE | ID: mdl-34888660

RESUMO

BACKGROUND: Patients with a significant decrease in hepatic venous pressure gradient (HVPG) have a considerable reduction of liver complications and higher survival after HCV eradication. OBJECTIVES: To evaluate the association between the baseline blood microbiome and the changes in HVPG after successful direct-acting antiviral (DAA) therapy in patients with HCV-related cirrhosis. METHODS: We performed a prospective study in 32 cirrhotic patients (21 HIV positive) with clinically significant portal hypertension (HVPG ≥10 mmHg). Patients were assessed at baseline and 48 weeks after HCV treatment completion. The clinical endpoint was a decrease in HVPG of ≥20% or HVPG <12 mmHg at the end of follow-up. Bacterial 16S ribosomal DNA was sequenced using MiSeq Illumina technology, inflammatory plasma biomarkers were investigated using ProcartaPlex immunoassays and the metabolome was investigated using GC-MS. RESULTS: During the follow-up, 47% of patients reached the clinical endpoint. At baseline, those patients had a higher relative abundance of Corynebacteriales and Diplorickettsiales order, Diplorickettsiaceae family, Corynebacterium and Aquicella genus and Undibacterium parvum species organisms and a lower relative abundance of Oceanospirillales and Rhodospirillales order, Halomonadaceae family and Massilia genus organisms compared with those who did not achieve the clinical endpoint according to the LEfSe algorithm. Corynebacteriales and Massilia were consistently found within the 10 bacterial taxa with the highest differential abundance between groups. Additionally, the relative abundance of the Corynebacteriales order was inversely correlated with IFN-γ, IL-17A and TNF-α levels and the Massilia genus with glycerol and lauric acid. CONCLUSIONS: Baseline-specific bacterial taxa are related to an HVPG decrease in patients with HCV-related cirrhosis after successful DAA therapy.


Assuntos
Hepatite C Crônica , Hipertensão Portal , Microbiota , Antivirais/uso terapêutico , Hepatite C Crônica/complicações , Hepatite C Crônica/tratamento farmacológico , Humanos , Hipertensão Portal/tratamento farmacológico , Hipertensão Portal/etiologia , Cirrose Hepática/complicações , Estudos Prospectivos
4.
Metabolomics ; 17(6): 53, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-34061256

RESUMO

INTRODUCTION: The silver/graphitic carbon nitride (Ag/g-C3N4) composite system exerts biocidal activity against the pathogenic bacterium Escherichia coli 1337-H that is stronger than that of well-known silver and titanium oxide (TiO2)-based composites. However, whether the Ag/g-C3N4 composite system has biocidal properties that the parent components do or do not have as separate chemical entities and whether they differ from those in Ag/TiO2 composite photocatalysts have not been clarified. OBJECTIVE: We investigated the chemical (cooperative charge handling and electronic properties) and biological (metabolic) effects exerted by the addition of Ag to g-C3N4 and to TiO2. METHODS: In this work, we undertook metabolome-wide analysis by liquid chromatography-electrospray ionization-quadrupole-time of flight-mass spectrometry to compare the metabolite profiles of untreated E. coli 1337-H cells or those subjected to disinfection with Ag, g-C3N4, 2Ag/g-C3N4, TiO2 and 2Ag/TiO2. RESULTS: While Ag or g-C3N4 moderately affected microbial metabolism according to the mean of the altered metabolites, multiple cell systems contributing to rapid cell death were immediately affected by the light-triggered radical species produced when Ag and g-C3N4 were as xAg/g-C3N4. The effects include drastically reduced production of small metabolites essential for detoxifying reactive oxygen species and those that regulate DNA replication fidelity, cell morphology and energy status. These biological consequences were different from those caused by Ag/TiO2-based biocides, demonstrating the uniqueness of the Ag/g-C3N4 system. CONCLUSIONS: Our results support the idea that the unique Ag/g-C3N4 biocidal properties are based on synergistic action and reveal new directions for designing future photocatalysts for use in disinfection and microbial control.


Assuntos
Escherichia coli , Prata , Catálise , Luz , Metabolômica , Prata/farmacologia
5.
Appl Environ Microbiol ; 86(24)2020 11 24.
Artigo em Inglês | MEDLINE | ID: mdl-32978141

RESUMO

Enzymatic processing of fish by-products for recovery of peptides (hydrolysates) is a promising technology to reach food grade ingredients of high nutritional quality. Despite this, their bitter taste and "fish" odor block implementation in food products and limit their economic potential. Trimethylamine (TMA) is a known contributor to malodor in fish. Current strategies to mask or remove the odor either are not effective or give rise to undesirable side effects. As an alternative approach to remediate TMA, we propose a novel enzymatic strategy to convert TMA into the odorless trimethylamine N-oxide (TMAO) using TMA monooxygenases (Tmms). We identified a diverse set of bacterial Tmms using a sequence similarity network. Purified, recombinant enzymes were assessed for their biocatalytic capacity by monitoring NADPH consumption and TMAO generation. Selected Tmms were subjected to biochemical characterization and investigated for their ability to oxidize TMA in an industry-relevant substrate. From the 45 bacterial Tmm candidates investigated, eight enzymes from four different taxa were selected for their high activity toward TMA. The three most active enzymes were shown to vary in temperature optimum, with the highest being 45°C. Enzymatic activity dropped at high temperatures, likely due to structural unfolding. The enzymes were all active from pH 6.0 to 8.5, with functional stability being lowest around the optimal pH. All three Tmms, given sufficient NADPH cofactor, were found to generate TMAO in the TMA-rich salmon protein hydrolysate. The Tmms serve as unique starting points for engineering and should be useful for guiding process development for marine biorefineries.IMPORTANCE Enzyme-based conversion of marine biomass to high-quality peptide ingredients leaves a distinct smell of "fish" caused by the presence of trimethylamine, which limits their economic potential. We suggest an enzymatic solution for converting trimethylamine to the odorless trimethylamine N-oxide as a novel strategy to improve the smell quality of marine protein hydrolysates. Following a systematic investigation of 45 putative bacterial trimethylamine monooxygenases from several phyla, we expand the repertoire of known active trimethylamine monooxygenases. As a proof-of-concept, we demonstrate that three of these enzymes oxidized trimethylamine in an industry-relevant salmon protein hydrolysate. Our results add new oxidoreductases to the industrial biocatalytic toolbox and provide a new point of departure for enzyme process developments in marine biorefineries.


Assuntos
Proteínas de Bactérias/metabolismo , Escherichia coli/metabolismo , Metilaminas/metabolismo , Oxigenases/metabolismo , Sequência de Aminoácidos , Proteínas de Bactérias/química , Oxigenases/química , Filogenia , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência
6.
Liver Int ; 40(9): 2215-2227, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32593189

RESUMO

BACKGROUND & AIMS: Hepatitis C virus (HCV), human immunodeficiency virus (HIV) and cirrhosis induce metabolic disorders. Here, we aimed to evaluate the association of plasma metabolites with Child-Turcotte-Pugh (CTP) score and hepatic decompensation in HIV/HCV-coinfected and HCV-monoinfected patients with advanced cirrhosis. METHODS: A cross-sectional study was carried out in 62 HIV/HCV-coinfected and 28 HCV-monoinfected patients. Metabolomics analysis was performed by gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS). The statistical association analysis was performed by partial least squares discriminant analysis (PLS-DA) and generalized linear model (GLM) with binomial distribution (to analyse HIV coinfection, high alcohol intake, treatment with statins, previous HCV therapy failure and decompensation) and ordinal logistic regression (OLR) models to analyse different stages of cirrhosis (CTP score). RESULTS: The statistical analysis identified plasma metabolites associated with HIV coinfection, high alcohol intake, CTP score and hepatic decompensation. Overall, fatty acids, bile acids, aromatic and sulphur amino acids, butyrate derivatives, oxidized phospholipids, energy-related metabolites and bacterial fermentation-related metabolites were increased in more advanced cirrhosis stages; while lysophosphatidylcholines and lysophosphatidylethanolamines, branched-chain amino acids (BCAA) and metabolites of tricarboxylic acid cycle, among others, were decreased in more advanced cirrhosis. Most of the significant metabolites displayed a similar trend after stratifying for HIV/HCV- and HCV-infected patients. Glycolic acid, LPC (16:0) and taurocholic acid had high accuracy for discriminating patients according to decompensated cirrhosis (CTP ≥ 7). CONCLUSION: Altered plasma metabolomic profile was associated with advanced stages of cirrhosis in HIV/HCV-coinfected and HCV-monoinfected patients.


Assuntos
Coinfecção , Infecções por HIV , Hepatite C Crônica , Hepatite C , Criança , Estudos de Coortes , Estudos Transversais , Infecções por HIV/complicações , Infecções por HIV/tratamento farmacológico , Hepacivirus , Hepatite C/complicações , Hepatite C Crônica/complicações , Humanos , Cirrose Hepática , Metabolômica
7.
J Proteome Res ; 18(1): 169-181, 2019 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-30362351

RESUMO

The characterization of specialized cell subpopulations in a heterogeneous tissue is essential for understanding organ function in health and disease. A popular method of cell isolation is fluorescence-activated cell sorting (FACS) based on probes that bind surface or intracellular markers. In this study, we analyze the impact of FACS on the cell metabolome of mouse peritoneal macrophages. Compared with directly pelleted macrophages, FACS-treated cells had an altered content of metabolites related to the plasma membrane, activating a mechanosensory signaling cascade causing inflammation-like stress. The procedure also triggered alterations related to energy consumption and cell damage. The observed changes mostly derive from the physical impact on cells during their passage through the instrument. These findings provide evidence of FACS-induced biochemical changes, which should be taken into account in the design of robust metabolic assays of cells separated by flow cytometry.


Assuntos
Separação Celular , Citometria de Fluxo/normas , Metaboloma , Animais , Células Cultivadas , Macrófagos Peritoneais/citologia , Macrófagos Peritoneais/metabolismo , Camundongos , Projetos de Pesquisa
8.
Appl Environ Microbiol ; 85(2)2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30413473

RESUMO

Amination of bulky ketones, particularly in (R) configuration, is an attractive chemical conversion; however, known ω-transaminases (ω-TAs) show insufficient levels of performance. By applying two screening methods, we discovered 10 amine transaminases from the class III ω-TA family that were 38% to 76% identical to homologues. We present examples of such enzymes preferring bulky ketones over keto acids and aldehydes with stringent (S) selectivity. We also report representatives from the class III ω-TAs capable of converting (R) and (S) amines and bulky ketones and one that can convert amines with longer alkyl substituents. The preference for bulky ketones was associated with the presence of a hairpin region proximal to the conserved Arg414 and residues conforming and close to it. The outward orientation of Arg414 additionally favored the conversion of (R) amines. This configuration was also found to favor the utilization of putrescine as an amine donor, so that class III ω-TAs with Arg414 in outward orientation may participate in vivo in the catabolism of putrescine. The positioning of the conserved Ser231 also contributes to the preference for amines with longer alkyl substituents. Optimal temperatures for activity ranged from 45 to 65°C, and a few enzymes retained ≥50% of their activity in water-soluble solvents (up to 50% [vol/vol]). Hence, our results will pave the way to design, in the future, new class III ω-TAs converting bulky ketones and (R) amines for the production of high-value products and to screen for those converting putrescine.IMPORTANCE Amine transaminases of the class III ω-TAs are key enzymes for modification of chemical building blocks, but finding those capable of converting bulky ketones and (R) amines is still challenging. Here, by an extensive analysis of the substrate spectra of 10 class III ω-TAs, we identified a number of residues playing a role in determining the access and positioning of bulky ketones, bulky amines, and (R)- and (S) amines, as well as of environmentally relevant polyamines, particularly putrescine. The results presented can significantly expand future opportunities for designing (R)-specific class III ω-TAs to convert valuable bulky ketones and amines, as well as for deepening the knowledge into the polyamine catabolic pathways.


Assuntos
Proteínas de Bactérias/genética , Bioprospecção , Genes Bacterianos , Cetonas/metabolismo , Poliaminas/metabolismo , Pseudomonas oleovorans/genética , Transaminases/genética , Sequência de Aminoácidos , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Pseudomonas oleovorans/enzimologia , Pseudomonas oleovorans/metabolismo , Alinhamento de Sequência , Transaminases/metabolismo
9.
J Bacteriol ; 200(3)2018 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-28874411

RESUMO

In 1680, Antonie van Leeuwenhoek noted compositional differences in his oral and fecal microbiota, pioneering the study of the diversity of the human microbiome. From Leeuwenhoek's time to successful modern attempts at changing the gut microbial landscape to cure disease, there has been an exponential increase in the recognition of our resident microbes as part of ourselves. Thus, the human host and microbiome have evolved in parallel to configure a balanced system in which microbes survive in homeostasis with our innate and acquired immune systems, unless disease occurs. A growing number of studies have demonstrated a correlation between the presence/absence of microbial taxa and some of their functional molecules (i.e., genes, proteins, and metabolites) with health and disease states. Nevertheless, misleading experimental design on human subjects and the cost and lack of standardized animal models pose challenges to answering the question of whether changes in microbiome composition are cause or consequence of a certain biological state. In this review, we evaluate the state of the art of methodologies that enable the study of the gut microbiome, encouraging a change in broadly used analytic strategies by choosing effector molecules (proteins and metabolites) in combination with coding nucleic acids. We further explore microbial and effector microbial product imbalances that relate to disease and health.


Assuntos
Microbioma Gastrointestinal , Biologia de Sistemas/métodos , Animais , Fezes/microbiologia , Saúde , Homeostase , Humanos
10.
Trop Med Int Health ; 23(6): 616-621, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29660815

RESUMO

OBJECTIVE: Preventive chemotherapy is the WHO-recommended control method for soil-transmitted helminthiases. In the Bolivian Chaco, 6-monthly single-dose mebendazole delivery to school-age children achieved a dramatic decrease in soil-transmitted helminthiases prevalence between 1987 and 2013. Consequently, in September 2016, preventive chemotherapy delivery was interrupted in nine rural communities. In compliance with WHO recommendations, we intensified surveillance to monitor soil-transmitted helminthiases prevalence and detect potential changes that would require interventions. METHODS: We conducted two cross-sectional parasitology surveys 12 months apart (September 2016-2017) among school-age children living in the communities where preventive chemotherapy delivery had been halted. Study design, methods of sampling and sample analysis technique (direct microscopy, Kato-Katz technique) followed WHO recommendations, aiming to obtain data representative of the Bolivian Chaco ecological zone. RESULTS: We collected 426 samples in 2016 and 520 in 2017. Soil-transmitted helminthiasis prevalence was unremarkable: 0.7% (95% CI 0-1.5%) in 2016 and 0.8% (0-1.5%) in 2017. Conversely, the prevalence of tapeworms (13% in 2016, 12% in 2017) and intestinal protozoan infections (81% in 2016 and 75% in 2017) continued to be high. CONCLUSIONS: Our findings support the role of preventive chemotherapy in reducing soil-transmitted helminthiases transmission, as otherwise poor hygienic and health conditions persist in the Bolivian Chaco. A national survey, involving areas from all the ecological zones of Bolivia, is now warranted.


Assuntos
Anti-Helmínticos/uso terapêutico , Helmintíase/prevenção & controle , Mebendazol/uso terapêutico , Bolívia/epidemiologia , Quimioprevenção/métodos , Criança , Estudos Transversais , Feminino , Helmintíase/epidemiologia , Humanos , Masculino , Prevalência , Avaliação de Programas e Projetos de Saúde , População Rural/estatística & dados numéricos , Solo/parasitologia
11.
Electrophoresis ; 38(18): 2275-2286, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28605027

RESUMO

The analysis of the microbial metabolome is crucial to fully understand the symbiotic relation between humans and microbes. That is why an explosion of metabolomics took place in the area. So far, at least several hundreds of microbial metabolites have been shown to be statistically altered when humans undergo a plethora of commonly faced perturbations. NMR and MS, usually coupled with GC, LC and CE have revealed their identities. CE is a robust analytical platform for the analysis of polar and ionic metabolites that are essential in order to assess the cells' activity. Due to its novelty, only 5% of the metabolomics studies investigate gut microbiota using CE, even though the metabolites found by CE as being significantly altered in human microbiota represent around 23% of the total number of metabolites identified by metabolomics tools. Herein, we discuss the advances of metabolomics in the frame of other OMICS techniques for human gut microbiota analysis. Afterwards, we focus on sample treatment, analytical methods and data processing in CE coupled to any detector that have been reported to date in order to enhance metabolite coverage in the art, and to identify metabolite markers that cannot be covered by other platforms but are of key importance for determining microbial activity and human health.


Assuntos
Eletroforese Capilar , Microbioma Gastrointestinal , Metabolômica , Animais , Humanos , Metaboloma/genética , Metaboloma/fisiologia , Fenótipo
12.
Bioinformatics ; 31(11): 1771-9, 2015 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-25618865

RESUMO

MOTIVATION: With the advent of meta-'omics' data, the use of metabolic networks for the functional analysis of microbial communities became possible. However, while network-based methods are widely developed for single organisms, their application to bacterial communities is currently limited. RESULTS: Herein, we provide a novel, context-specific reconstruction procedure based on metaproteomic and taxonomic data. Without previous knowledge of a high-quality, genome-scale metabolic networks for each different member in a bacterial community, we propose a meta-network approach, where the expression levels and taxonomic assignments of proteins are used as the most relevant clues for inferring an active set of reactions. Our approach was applied to draft the context-specific metabolic networks of two different naphthalene-enriched communities derived from an anthropogenically influenced, polyaromatic hydrocarbon contaminated soil, with (CN2) or without (CN1) bio-stimulation. We were able to capture the overall functional differences between the two conditions at the metabolic level and predict an important activity for the fluorobenzoate degradation pathway in CN1 and for geraniol metabolism in CN2. Experimental validation was conducted, and good agreement with our computational predictions was observed. We also hypothesize different pathway organizations at the organismal level, which is relevant to disentangle the role of each member in the communities. The approach presented here can be easily transferred to the analysis of genomic, transcriptomic and metabolomic data.


Assuntos
Bactérias/metabolismo , Naftalenos/metabolismo , Poluentes do Solo/metabolismo , Bactérias/classificação , Bactérias/genética , Redes e Vias Metabólicas , Proteômica
13.
World J Microbiol Biotechnol ; 32(12): 201, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27785708

RESUMO

The analysis of catabolic capacities of microorganisms is currently often achieved by cultivation approaches and by the analysis of genomic or metagenomic datasets. Recently, a microarray system designed from curated key aromatic catabolic gene families and key alkane degradation genes was designed. The collection of genes in the microarray can be exploited to indicate whether a given microbe or microbial community is likely to be functionally connected with certain degradative phenotypes, without previous knowledge of genome data. Herein, this microarray was applied to capture new insights into the catabolic capacities of copper-resistant actinomycete Amycolatopsis tucumanensis DSM 45259. The array data support the presumptive ability of the DSM 45259 strain to utilize single alkanes (n-decane and n-tetradecane) and aromatics such as benzoate, phthalate and phenol as sole carbon sources, which was experimentally validated by cultivation and mass spectrometry. Interestingly, while in strain DSM 45259 alkB gene encoding an alkane hydroxylase is most likely highly similar to that found in other actinomycetes, the genes encoding benzoate 1,2-dioxygenase, phthalate 4,5-dioxygenase and phenol hydroxylase were homologous to proteobacterial genes. This suggests that strain DSM 45259 contains catabolic genes distantly related to those found in other actinomycetes. Together, this study not only provided new insight into the catabolic abilities of strain DSM 45259, but also suggests that this strain contains genes uncommon within actinomycetes.


Assuntos
Actinobacteria/genética , Proteínas de Bactérias/genética , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Análise de Sequência de DNA/métodos , Actinobacteria/metabolismo , Alcanos/metabolismo , Proteínas de Bactérias/metabolismo , Biodegradação Ambiental , Cobre/metabolismo , Evolução Molecular , Metabolismo
14.
Proteomics ; 15(20): 3508-20, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26201687

RESUMO

Crude oil is one of the most important natural assets for humankind, yet it is a major environmental pollutant, notably in marine environments. One of the largest crude oil polluted areas in the word is the semi-enclosed Mediterranean Sea, in which the metabolic potential of indigenous microbial populations towards the large-scale chronic pollution is yet to be defined, particularly in anaerobic and micro-aerophilic sites. Here, we provide an insight into the microbial metabolism in sediments from three chronically polluted marine sites along the coastline of Italy: the Priolo oil terminal/refinery site (near Siracuse, Sicily), harbour of Messina (Sicily) and shipwreck of MT Haven (near Genoa). Using shotgun metaproteomics and community metabolomics approaches, the presence of 651 microbial proteins and 4776 metabolite mass features have been detected in these three environments, revealing a high metabolic heterogeneity between the investigated sites. The proteomes displayed the prevalence of anaerobic metabolisms that were not directly related with petroleum biodegradation, indicating that in the absence of oxygen, biodegradation is significantly suppressed. This suppression was also suggested by examining the metabolome patterns. The proteome analysis further highlighted the metabolic coupling between methylotrophs and sulphate reducers in oxygen-depleted petroleum-polluted sediments.


Assuntos
Metabolômica , Poluição por Petróleo , Proteômica , Biodegradação Ambiental , Sedimentos Geológicos/microbiologia , Itália , Mar Mediterrâneo , Petróleo/toxicidade , Microbiologia da Água
16.
Gut ; 62(11): 1591-601, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-23236009

RESUMO

OBJECTIVE: Antibiotic (AB) usage strongly affects microbial intestinal metabolism and thereby impacts human health. Understanding this process and the underlying mechanisms remains a major research goal. Accordingly, we conducted the first comparative omic investigation of gut microbial communities in faecal samples taken at multiple time points from an individual subjected to ß-lactam therapy. METHODS: The total (16S rDNA) and active (16S rRNA) microbiota, metagenome, metatranscriptome (mRNAs), metametabolome (high-performance liquid chromatography coupled to electrospray ionisation and quadrupole time-of-flight mass spectrometry) and metaproteome (ultra high performing liquid chromatography coupled to an Orbitrap MS(2) instrument [UPLC-LTQ Orbitrap-MS/MS]) of a patient undergoing AB therapy for 14 days were evaluated. RESULTS: Apparently oscillatory population dynamics were observed, with an early reduction in Gram-negative organisms (day 6) and an overall collapse in diversity and possible further colonisation by 'presumptive' naturally resistant bacteria (day 11), followed by the re-growth of Gram-positive species (day 14). During this process, the maximum imbalance in the active microbial fraction occurred later (day 14) than the greatest change in the total microbial fraction, which reached a minimum biodiversity and richness on day 11; additionally, major metabolic changes occurred at day 6. Gut bacteria respond to ABs early by activating systems to avoid the antimicrobial effects of the drugs, while 'presumptively' attenuating their overall energetic metabolic status and the capacity to transport and metabolise bile acid, cholesterol, hormones and vitamins; host-microbial interactions significantly improved after treatment cessation. CONCLUSIONS: This proof-of-concept study provides an extensive description of gut microbiota responses to follow-up ß-lactam therapy. The results demonstrate that ABs targeting specific pathogenic infections and diseases may alter gut microbial ecology and interactions with host metabolism at a much higher level than previously assumed.


Assuntos
Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Trato Gastrointestinal/microbiologia , Microbiota/efeitos dos fármacos , beta-Lactamas/farmacologia , Idoso , Bactérias/classificação , Bactérias/isolamento & purificação , Técnicas de Tipagem Bacteriana/métodos , Biodiversidade , DNA Bacteriano/análise , Fezes/microbiologia , Trato Gastrointestinal/metabolismo , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Metaboloma/efeitos dos fármacos , RNA Bacteriano/análise , RNA Ribossômico 16S/análise
17.
Front Cell Infect Microbiol ; 14: 1340610, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38550617

RESUMO

Background: Combination antiretroviral therapy (ART) has transformed human immunodeficiency virus (HIV) infection in people with HIV (PWH). However, a chronic state of immune activation and inflammation is maintained despite achieving HIV suppression and satisfactory immunological recovery. We aimed to determine whether the plasma metabolomic profile of PWH on long-term suppressive ART and immunologically recovered approximates the normality by comparison with healthy controls with similar age and gender. Methods: We carried out a cross-sectional study in 17 PWH on long-term ART (HIV-RNA <50 copies/mL, CD4+ ≥500 cells/mm3, and CD4+/CD8+ ≥1) and 19 healthy controls with similar age and gender. Metabolomics analysis was performed by gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS). The statistical association analysis was performed by principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), and Generalized Linear Models (GLM) with a gamma distribution (log-link). Significance levels (p-value) were corrected for multiple testing (q-value). Results: PCA and PLS-DA analyses found no relevant differences between groups. Adjusted GLM showed 14 significant features (q-value<0.20), of which only three could be identified: lysophosphatidylcholine (LysoPC) (22:6) (q-value=0.148), lysophosphatidylethanolamine (LysoPE) (22:6) (q-value=0.050) and hydroperoxy-octadecatrienoic acid (HpOTrE)/dihydroperoxy-octadecatrienoic acid (DiHOTrE)/epoxy-octadecadienoic acid (EpODE) (q-value=0.136). These significant identified metabolites were directly correlated to plasma inflammatory biomarkers in PWH and negatively correlated in healthy controls. Conclusion: PWH on long-term ART have a metabolomic profile that is almost normal compared to healthy controls. Nevertheless, residual metabolic alterations linked to inflammatory biomarkers persist, which could favor the development of age-related comorbidities among this population.


Assuntos
Infecções por HIV , Metabolômica , Humanos , Estudos Transversais , Metabolômica/métodos , Biomarcadores , Inflamação/metabolismo
18.
Biomolecules ; 13(1)2023 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-36671530

RESUMO

An oxidative degradation product of the polyunsaturated fatty acids, 4-hydroxynonenal (4-HNE), is of particular interest in cancer research due to its concentration-dependent pleiotropic activities affecting cellular antioxidants, metabolism, and growth control. Although an increase in oxidative stress and lipid peroxidation was already associated with prostate cancer progression a few decades ago, the knowledge of the involvement of 4-HNE in prostate cancer tumorigenesis is limited. This study investigated the appearance of 4-HNE-protein adducts in prostate cancer tissue by immunohistochemistry using a genuine 4-HNE monoclonal antibody. Plasma samples of the same patients and samples of the healthy controls were also analyzed for the presence of 4-HNE-protein adducts, followed by metabolic profiling using LC-ESI-QTOF-MS and GC-EI-Q-MS. Finally, the analysis of the metabolic pathways affected by 4-HNE was performed. The obtained results revealed the absence of 4-HNE-protein adducts in prostate carcinoma tissue but increased 4-HNE-protein levels in the plasma of these patients. Metabolomics revealed a positive association of different long-chain and medium-chain fatty acids with the presence of prostate cancer. Furthermore, while linoleic acid positively correlated with the levels of 4-HNE-protein adducts in the blood of healthy men, no correlation was obtained for cancer patients indicating altered lipid metabolism in this case. The metabolic pathway of unsaturated fatty acids biosynthesis emerged as significantly affected by 4-HNE. Overall, this is the first study linking 4-HNE adduction to plasma proteins with specific alterations in the plasma metabolome of prostate cancer patients. This study revealed that increased 4-HNE plasma protein adducts could modulate the unsaturated fatty acids biosynthesis pathway. It is yet to be determined if this is a direct result of 4-HNE or whether they are produced by the same underlying mechanisms. Further mechanistic studies are needed to grasp the biological significance of the observed changes in prostate cancer tumorigenesis.


Assuntos
Aldeídos , Neoplasias da Próstata , Masculino , Humanos , Peroxidação de Lipídeos , Aldeídos/metabolismo , Proteínas/metabolismo , Biomarcadores/metabolismo , Ácidos Graxos Insaturados , Carcinogênese
19.
Artigo em Inglês | MEDLINE | ID: mdl-37454721

RESUMO

Alzheimer's disease (AD) is often not recognized or is diagnosed very late, which significantly reduces the effectiveness of available pharmacological treatments. Metabolomic analyzes have great potential for improving existing knowledge about the pathogenesis and etiology of AD and represent a novel approach towards discovering biomarkers that could be used for diagnosis, prognosis, and therapy monitoring. In this study, we applied the untargeted metabolomic approach to investigate the changes in biochemical pathways related to AD pathology. We used gas chromatography and liquid chromatography coupled to mass spectrometry (GC-MS and LC-MS, respectively) to identify metabolites whose levels have changed in subjects with AD diagnosis (N = 40) compared to healthy controls (N = 40) and individuals with mild cognitive impairment (MCI, N = 40). The GC-MS identified significant differences between groups in levels of metabolites belonging to the classes of benzene and substituted derivatives, carboxylic acids and derivatives, fatty acyls, hydroxy acids and derivatives, keto acids and derivatives, and organooxygen compounds. Most of the compounds identified by the LC-MS were various fatty acyls, glycerolipids and glycerophospholipids. All of these compounds were decreased in AD patients and in subjects with MCI compared to healthy controls. The results of the study indicate disturbed metabolism of lipids and amino acids and an imbalance of metabolites involved in energy metabolism in individuals diagnosed with AD, compared to healthy controls and MCI subjects.


Assuntos
Doença de Alzheimer , Humanos , Doença de Alzheimer/metabolismo , Metabolômica , Metaboloma , Espectrometria de Massas , Biomarcadores
20.
Anal Chim Acta ; 1210: 339043, 2022 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-35595356

RESUMO

GC-MS for untargeted metabolomics is a well-established technique. Small molecules and molecules made volatile by derivatization can be measured and those compounds are key players in main biological pathways. This tutorial provides ready-to-use protocols for GC-MS-based metabolomics, using either the well-known low-resolution approach (GC-Q-MS) with nominal mass or the more recent high-resolution approach (GC-QTOF-MS) with accurate mass, discussing their corresponding strengths and limitations. Analytical procedures are covered for different types of biofluids (plasma/serum, bronchoalveolar lavage, urine, amniotic fluid) tissue samples (brain/hippocampus, optic nerve, lung, kidney, liver, pancreas) and samples obtained from cell cultures (adipocytes, macrophages, Leishmania promastigotes, mitochondria, culture media). Together with the sample preparation and data acquisition, data processing strategies are described specially focused on Agilent equipments, including deconvolution software and database annotation using spectral libraries. Manual curation strategies and quality control are also deemed. Finally, considerations to obtain a semiquantitative value for the metabolites are also described. As a case study, an illustrative example from one of our experiments at CEMBIO Research Centre, is described and findings discussed.


Assuntos
Líquidos Corporais , Metabolômica , Bases de Dados Factuais , Cromatografia Gasosa-Espectrometria de Massas/métodos , Metabolômica/métodos , Software
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