Antibiosis and dark-pigments secretion by the phytopathogenic and environmental fungal species after interaction in vitro with a Bacillus subtilis isolate

In this work, different reactions in vitro between an environmental bacterial isolate and fungal species were related. The Gram-positive bacteria had terminal and subterminal endospores, presented metabolic characteristics of mesophilic and acidophilic growth, halotolerance, positive to nitrate reduction and enzyme production, as caseinase and catalase. The analysis of partial sequences containing 400 to 700 bases of the 16S ribosomal RNA gene showed identity with the genus Bacillus. However, its identity as B. subtilis was confirmed after analyses of the rpoB, gyrA, and 16S rRNA near-full-length sequences. Strong inhibitory activity of environmental microorganisms, such as Penicillium sp, Aspergillus flavus, A. niger, and phytopathogens, such as Colletotrichum sp, Alternaria alternata, Fusarium solani and F. oxysporum f.sp vasinfectum, was shown on co-cultures with B. subtilis strain, particularly on Sabouraud dextrose agar (SDA) and DNase media. Red and red-ochre color pigments, probably phaeomelanins, were secreted by A. alternata and A. niger respectively after seven days of co-culture.

Volatile organic compounds produced by Saccharomyces cerevisiae inhibit the in vitro development of Guignardia citricarpa, the causal agent of citrus black spot

Due to the low chemical control effectiveness of citrus black spot, caused by the fungus Guignardia citricarpa at postharvest, and to the search for alternative control methods, this study aimed to evaluate the in vitro effect of volatile organic compounds (VOCs), produced by yeast Saccharomyces cerevisiae, on G. citricarpa. It was observed that the yeast strains evaluated acted as antagonists by VOC production, whose maximum inhibitory capacity was as high as 87.2%. The presence of fermentable carbon sources in the medium was essential for the bioactive VOC production by the yeast. The analysis of VOCs produced in PDA medium by SPME-GC-MS indicated the presence of high quantities of alcohols as well as esters. An artificial VOC mixture prepared on the basis of the composition of the VOCs mimicked the inhibitory effects of the natural VOCs released by S. cerevisiae. Thus, the VOCs produced by the yeast or the artificial mixtures can be a promising control method for citrus black spot or others postharvest diseases.

Estudos metabolômicos entre interações da rizosfera de Senna spectabilis e rizobacterias associadas

Pós-graduação em Química - IQ

Modulação da doença periodontal por curcumin modificado quimicamente. Estudo de dose-resposta em roedor

Pós-graduação em Odontologia - FOAR

Metabolomic insights into the intricate gut microbial-host interaction in the development of obesity and type 2 diabetes.

Gut microbiota has recently been proposed as a crucial environmental factor in the development of metabolic diseases such as obesity and type 2 diabetes, mainly due to its contribution in the modulation of several processes including host energy metabolism, gut epithelial permeability, gut peptide hormone secretion, and host inflammatory state. Since the symbiotic interaction between the gut microbiota and the host is essentially reflected in specific metabolic signatures, much expectation is placed on the application of metabolomic approaches to unveil the key mechanisms linking the gut microbiota composition and activity with disease development. The present review aims to summarize the gut microbial-host co-metabolites identified so far by targeted and untargeted metabolomic studies in humans, in association with impaired glucose homeostasis and/or obesity. An alteration of the co-metabolism of bile acids, branched fatty acids, choline, vitamins (i.e., niacin), purines, and phenolic compounds has been associated so far with the obese or diabese phenotype, in respect to healthy controls. Furthermore, anti-diabetic treatments such as metformin and sulfonylurea have been observed to modulate the gut microbiota or at least their metabolic profiles, thereby potentially affecting insulin resistance through indirect mechanisms still unknown. Despite the scarcity of the metabolomic studies currently available on the microbial-host crosstalk, the data-driven results largely confirmed findings independently obtained from in vitro and animal model studies, putting forward the mechanisms underlying the implication of a dysfunctional gut microbiota in the development of metabolic disorders.

Colonization-induced host-gut microbial metabolic interaction

The gut microbiota enhances the host's metabolic capacity for processing nutrients and drugs and modulate the activities of multiple pathways in a variety of organ systems. We have probed the systemic metabolic adaptation to gut colonization for 20 days following exposure of axenic mice (n = 35) to a typical environmental microbial background using high-resolution (1)H nuclear magnetic resonance (NMR) spectroscopy to analyze urine, plasma, liver, kidney, and colon (5 time points) metabolic profiles. Acquisition of the gut microbiota was associated with rapid increase in body weight (4%) over the first 5 days of colonization with parallel changes in multiple pathways in all compartments analyzed. The colonization process stimulated glycogenesis in the liver prior to triggering increases in hepatic triglyceride synthesis. These changes were associated with modifications of hepatic Cyp8b1 expression and the subsequent alteration of bile acid metabolites, including taurocholate and tauromuricholate, which are essential regulators of lipid absorption. Expression and activity of major drug-metabolizing enzymes (Cyp3a11 and Cyp2c29) were also significantly stimulated. Remarkably, statistical modeling of the interactions between hepatic metabolic profiles and microbial composition analyzed by 16S rRNA gene pyrosequencing revealed strong associations of the Coriobacteriaceae family with both the hepatic triglyceride, glucose, and glycogen levels and the metabolism of xenobiotics. These data demonstrate the importance of microbial activity in metabolic phenotype development, indicating that microbiota manipulation is a useful tool for beneficially modulating xenobiotic metabolism and pharmacokinetics in personalized health care. IMPORTANCE: Gut bacteria have been associated with various essential biological functions in humans such as energy harvest and regulation of blood pressure. Furthermore, gut microbial colonization occurs after birth in parallel with other critical processes such as immune and cognitive development. Thus, it is essential to understand the bidirectional interaction between the host metabolism and its symbionts. Here, we describe the first evidence of an in vivo association between a family of bacteria and hepatic lipid metabolism. These results provide new insights into the fundamental mechanisms that regulate host-gut microbiota interactions and are thus of wide interest to microbiological, nutrition, metabolic, systems biology, and pharmaceutical research communities. This work will also contribute to developing novel strategies in the alteration of host-gut microbiota relationships which can in turn beneficially modulate the host metabolism.

The interaction of sodium chlorite with phospholipids and glutathione:a comparison of effects in vitro, in mammalian and in microbial cells

In this study the interaction of the preservative sodium chlorite with unsaturated lipids and glutathione was investigated, in comparison with peroxides, sodium hypochlorite, and benzalkonium chloride. The aim was to determine whether the action of sodium chlorite could involve membrane lipid damage or antioxidant depletion, and how this related to toxicity in both mammalian and microbial cells. The treatment of phospholipids with chlorite yielded low levels of hydroperoxides, but sodium chlorite oxidized the thiol-containing antioxidant glutathione to its disulfide form very readily in vitro, with a 1:4 oxidant:GSH stoichiometry. In cultured cells, sodium chlorite also caused a substantial depletion of intracellular glutathione, whereas lipid oxidation was not very prominent. Sodium chlorite had a lower toxicity to ocular mammalian cells than benzalkonium chloride, which could be responsible for the different effects of long-term application in the eye. The fungal cells, which were most resistant to sodium chlorite, maintained higher percentage levels of intracellular glutathione during treatment than the mammalian cells. The results show that sodium chlorite can cause oxidative stress in cells, and suggest that cell damage is more likely to be due to interaction with thiol compounds than with cell membrane lipids. The study also provides important information about the differential resistance of ocular cells and microbes to various preservatives and oxidants.

High levels of Bifidobacteria are associated with increased levels of anthocyanin microbial metabolites: a randomized clinical trial.

The health benefits associated with the consumption of polyphenol-rich foods have been studied in depth, however, the full mechanism of action remains unknown. One of the proposed mechanisms is through microbiota interaction. In the present study, we aimed to explore the relationship between changes in fecal microbiota and changes in urinary phenolic metabolites after wine interventions. Nine participants followed a randomized, crossover, controlled interventional trial. After the washout period, they received red wine, dealcoholized red wine or gin for 20 days each. Polyphenol metabolites (n > 60) in urine were identified and quantified by UPLC-MS/MS and the microbial content of fecal samples was quantified by real-time quantitative PCR. Interventions with both red wine and dealcoholized red wine increased the fecal concentration of Bifidobacterium, Enterococcus and Eggerthella lenta, compared to gin intervention and baseline. When participants were categorized in tertiles of changes in fecal bacteria, those in the highest tertile of Bifidobacteria had higher urinary concentration changes in syringic acid, p-coumaric acid, 4-hydroxybenzoic acid and homovanillic acid (all anthocyanin metabolites) than those in tertile 1 (P < 0.05, all). In addition, changes of Bifidobacteria correlated positively with changes of these metabolites (r = 0.5-0.7, P < 0.05, all). Finally, the 68.5% changes in Bifidobacteria can be predicted by syringic acid and 4-hydroxybenzoic acid changes. This study confirms the important role of polyphenols as bacterial substrates and their modulatory capacity as an important field in the research of new products with prebiotic and probiotic characteristics for the food industry.

Interaction d'Escherichia coli entérohémorragique (EHEC) avec Acanthamoeba castellanii et rôle du régulon Pho chez les EHEC

Les EHEC de sérotype O157:H7 sont des agents zoonotiques d’origine alimentaire ou hydrique. Ce sont des pathogènes émergeants qui causent chez l’humain des épidémies de gastro-entérite aiguë et parfois un syndrome hémolytique-urémique. Les EHEC réussissent leur transmission à l’humain à partir de leur portage commensal chez l’animal en passant par l’étape de survie dans l’environnement. L’endosymbiose microbienne est une des stratégies utilisées par les bactéries pathogènes pour survivre dans les environnements aquatiques. Les amibes sont des protozoaires vivants dans divers écosystèmes et connus pour abriter plusieurs agents pathogènes. Ainsi, les amibes contribueraient à transmettre les EHEC à l'humain. La première partie de mon projet de thèse est centrée sur l'interaction de l’amibe Acanthamoeba castellanii avec les EHEC. Les résultats montrent que la présence de cette amibe prolonge la persistance des EHEC, et ces dernières survivent à leur phagocytose par les amibes. Ces résultats démontrent le potentiel réel des amibes à héberger les EHEC et à contribuer à leur transmission. Cependant, l’absence de Shiga toxines améliore leur taux de survie intra-amibe. Par ailleurs, les Shiga toxines sont partiellement responsables de l’intoxication des amibes par les EHEC. Cette implication des Shiga toxines dans le taux de survie intracellulaire et dans la mortalité des amibes démontre l’intérêt d’utiliser les amibes comme modèle d'interaction hôte/pathogène pour étudier la pathogénicité des EHEC. Durant leur cycle de transmission, les EHEC rencontrent des carences en phosphate inorganique (Pi) dans l’environnement. En utilisant conjointement le système à deux composantes (TCS) PhoB-R et le système Pst (transport spécifique de Pi), les EHEC détectent et répondent à cette variation en Pi en activant le régulon Pho. La relation entre la virulence des EHEC, le PhoB-R-Pst et/ou le Pi environnemental demeure inconnue. La seconde partie de mon projet explore le rôle du régulon Pho (répondant à un stress nutritif de limitation en Pi) dans la virulence des EHEC. L’analyse transcriptomique montre que les EHEC répondent à la carence de Pi par une réaction complexe impliquant non seulement un remodelage du métabolisme général, qui est critique pour sa survie, mais aussi en coordonnant sa réponse de virulence. Dans ces conditions le régulateur PhoB contrôle directement l’expression des gènes du LEE et de l’opéron stx2AB. Ceci est confirmé par l’augmentation de la sécrétion de l’effecteur EspB et de la production et sécrétion de Stx2 en carence en Pi. Par ailleurs, l’activation du régulon Pho augmente la formation de biofilm et réduit la motilité chez les EHEC. Ceci corrèle avec l’induction des gènes régulant la production de curli et la répression de la voie de production d’indole et de biosynthèse du flagelle et du PGA (Polymère β-1,6-N-acétyle-D-glucosamine).

Haematology of Fenneropenaeus indicus in response to environmental alterations and microbial infections

Aquaculture has developed to become one of the fastest growing food producing sectors in the world.Today India is one among the major shrimp producing countries in the world.There are extensive and intensive shrimp culture practices. In extensive shrimp culture, shrimps are stocked at low densities (< 25 PLs m'2)in large ponds or tidal enclosures in which little or no management is exercised or possible. Farmers depend almost entirely on natural conditions in extensive cultures. Intensive shrimp culture is carried out in high densities (>200 PLs m'2). Much of the world shrimp production still comes from extensive culture.There is a growing demand for fish and marine products for human and animal consumption. This demand has led to rapid growth of aquaculture, which some times has been accompanied by ecological impacts and economic loss due to diseases. The expansion of shrimp culture always accompanies local environmental degradation and occurrence of diseases.Disease out breaks is recognised as a significant constraint to aquaculture production. Environmental factors, water quality, pollution due to effluent discharge and pathogenic invasion due to vertical and horizontal transmission are the main causes of shrimp disease out breaks. Nutritional imbalance, toxicant and other pollutants also account for the onset of diseases. pathogens include viruses, bacteria, fungi and parasites.Viruses are the most economically significant pathogens of the cultured shrimps world wide. Disease control in shrimp aquaculture should focus first on preventive measures for eliminating disease promoting factors.ln order to design prophylactic and proactive measures against shrimp diseases, it is mandatory to understand the immune make up of the cultivable species, its optimum culture conditions and the physico chemical parameters of the rearing environment. It has been proven beyond doubt that disease is an end result of complex interaction of environment, pathogen and the host animal. The aquatic environment is abounded with infectious microbes.The transmission of disease in this environment is extremely easy, especially under dense, culture conditions. Therefore, a better understanding of the immune responses of the cultured animal in relation to its environmental alterations and microbial invasions is essential indevising strategic measures against aquaculture loss due to diseases. This study accentuate the importance of proper and regular health monitoring in shrimps employing the most appropriate haematological biomarkers for application of suitable prophylactic measures in order to avoid serious health hazards in shrimp culture systems.

Microbial response to restoration of tantalite mine soils in western Rwanda

Artisanal columbite-tantalite (coltan) mining has had negative effects on the rural economy in the great Lakes region of Africa through labor deficits, degradation and loss of farmland, food insecurity, high cost of living, and reduced traditional export crop production alongside secondary impacts that remotely affect the quality of air, water, soil, plants, animals, and human wellbeing. The situation is multifaceted and calls for a holistic approach for short and long-term mitigation of such negative effects. This study focuses on the effects of mine land restoration on soil microbiological quality in the Gatumba Mining District of western Rwanda. Some coltan mine wastelands were afforested with pine and eucalyptus trees while farmers directly cultivated others due to land scarcity. Farmyard manure (FYM) is the sole fertilizer applied on the wastelands although it is insufficient to achieve the desired crop yields. Despite this, several multi-purpose plants such as Tithonia diversifolia, Markhamia lutea, and Canavalia brasiliensis thrive in the area and could supplement FYM. The potential for these “new” amendments to improve soil microbial properties, particularly in the tantalite mine soils was investigated. The specific objectives of the study were to: (a) evaluate the effects of land use on soil microbial indices of the tantalite mine soils; (b) investigate the restorative effects of organic amendments on a Technosol; and (c) estimate the short-term N and P supply potential of the soil amendments in the soils. Fresh soils (0-20 cm) from an unmined native forest, two mine sites afforested with pine and eucalyptus forests (pine and eucalyptus Technosols), an arable land, and two cultivated Technosols (Kavumu and Kirengo Technosols) were analyzed for the physicochemical properties. Afterwards, a 28-day incubation (22oC) experiment was conducted followed by measurements of mineral N, soil microbial biomass C, N, P, and fungal ergosterol contents using standard methods. This was followed by a 12-week incubation study of the arable soil and the Kavumu Technosol amended with FYM, Canavalia and Tithonia biomass, and Markhamia leaf litter after which soil microbial properties were measured at 2, 8, and 12 weeks of incubation. Finally, two 4-week incubation experiments each were conducted in soils of the six sites to estimate (i) potential mineralizable N using a soil-sand mixture (1:1) amended with Canavalia and goat manure and (ii) P mineralization mixtures (1:1) of soil and anion exchange resins in bicarbonate form amended with Tithonia biomass and goat manure. In study one, afforestation increased soil organic carbon and total N contents in the pine and eucalyptus Technosols by 34-40% and 28-30%, respectively of that in the native forest soil. Consequently, the microbial biomass and activity followed a similar trend where the cultivated Technosols were inferior to the afforested ones. The microbial indices of the mine soils were constrained by soil acidity, dithionite-extractable Al, and low P availability. In study two, the amendments substantially increased C and N mineralization, microbial properties compared with non-amended soils. Canavalia biomass increased CO2 efflux by 340%, net N mineralization by 30-140%, and microbial biomass C and N by 240-600% and 240-380% (P < 0.01), respectively after four weeks of incubation compared with the non-amended soils. Tithonia biomass increased ergosterol content by roughly 240%. The Kavumu Technosol showed a high potential for quick restoration of its soil quality due to its major responses to the measured biological parameters. In study three, Canavalia biomass gave the highest mineralizable N (130 µg g-1 soil, P < 0.01) in the Kavumu Technosol and the lowest in the native forest soil (-20 µg g-1 soil). Conversely, the mineralizable N of goat manure was negative in all soils ranging from -2.5 µg N g-1 to -7.7 µg N g-1 soil except the native forest soil. However, the immobilization of goat manure N in the “cultivated soils” was 30-70% lower than in the “forest soils” signifying an imminent recovery of the amended soils from N immobilization. The mineralization of goat manure P was three-fold that of Tithonia, constituting 61-71% of total P applied. Phosphorus mineralization slightly decreased after four weeks of incubation due to sulfate competition as reflected in a negative correlation, which was steeper in the Tithonia treatment. In conclusion, each amendment used in this research played a unique role in C, N, and P mineralization and contributed substantially to microbial properties in the tantalite mine soils. Interestingly, the “N immobilizers” exhibited potentials for P release and soil organic carbon storage. Consequently, the combined use of the amendments in specific ratios, or co-composting prior to application is recommended to optimize nutrient release, microbial biomass dynamics and soil organic matter accrual. Transport of organic inputs seems more feasible for smallholder farmers who typically manage small field sizes. To reduce acidity in the soils, liming with wood ash was recommended to also improve P availability and enhance soil biological quality, even if it may only be possible on small areas. Further, afforestation with mixed-species of fast-growing eucalyptus and legume or indigenous tree species are suggested to restore tantalite mine wastelands. It is emphasized most of this research was conducted under controlled laboratory conditions, which exclude interaction with environmental variables. Also fine fractions of the amendments were used compared with the usual practice of applying a mixture of predominantly coarser fractions. Therefore, the biological dynamics reported in the studies here may not entirely reflect those of farmers’ field conditions.

The role of fungi and bacteria on the organic matter decomposition process in streams: interaction and relevance in biofilms

L'objectiu d'aquest estudi és el d'investigar sobre l'ús de matèria orgànica per part dels fongs i bacteris que colonitzen diferents substrats bentònics en rius Mediterranis i analitzar l'efecte dels factors ambientals i antròpics sobre l'estabilitat estructural i funcional de les comunitats del biofilm. La metodologia emprada en aquest estudi consisteix en: i) anàlisi de la biomassa bacteriana i fúngica, ii) anàlisi de la composició de les comunitats bentòniques (identificació d'hifomicets aquàtics i anàlisi del 16S rDNA bacterià), i iii) anàlisi de l'activitat enzimàtica extracel·lular relacionada amb el reciclatge de matèria orgànica en rius.

Microbial impacts on plant-herbivore interactions: the indirect effects of a birch pathogen on a birch aphid

The role of indirect interactions in structuring communities is becoming increasingly recognised. Plant fungi can bring about changes in plant chemistry which may affect insect herbivores that share the same plant, and hence the two may interact indirectly. This study investigated the indirect effects of a fungal pathogen (Marssonina betulae) of silver birch (Betula pendula) on an aphid (Euceraphis betulae), and the processes underpinning the interaction. There was a strong positive association between natural populations of the aphid and leaves bearing high fungal infection. In choice tests, significantly more aphids settled on leaves inoculated with the fungus than on asymptomatic leaves. Individual aphids reared on inoculated leaves were heavier, possessed longer hind tibiae and displayed enhanced embryo development compared with aphids reared on asymptomatic leaves; population growth rate was also positively correlated with fungal infection when groups of aphids were reared on inoculated branches. Changes in leaf chemistry were associated with fungal infection with inoculated leaves containing higher concentrations of free-amino acids. This may reflect a plant-initiated response to fungal attack in which free amino acids from the degradation of mesophyll cells are translocated out of infected leaves via the phloem. These changes in plant chemistry are similar to those occurring during leaf senescence, and are proposed as the mechanistic basis for the positive interaction between the fungus and aphid.

Development and application of in vivo expression technology (IVET) for analysing microbial gene expression in complex environments

Establishing the mechanisms by which microbes interact with their environment, including eukaryotic hosts, is a major challenge that is essential for the economic utilisation of microbes and their products. Techniques for determining global gene expression profiles of microbes, such as microarray analyses, are often hampered by methodological restraints, particularly the recovery of bacterial transcripts (RNA) from complex mixtures and rapid degradation of RNA. A pioneering technology that avoids this problem is In Vivo Expression Technology (IVET). IVET is a 'promoter-trapping' methodology that can be used to capture nearly all bacterial promoters (genes) upregulated during a microbe-environment interaction. IVET is especially useful because there is virtually no limit to the type of environment used (examples to date include soil, oomycete, a host plant or animal) to select for active microbial promoters. Furthermore, IVET provides a powerful method to identify genes that are often overlooked during genomic annotation, and has proven to be a flexible technology that can provide even more information than identification of gene expression profiles. A derivative of IVET, termed resolvase-IVET (RIVET), can be used to provide spatio-temporal information about environment-specific gene expression. More recently, niche-specific genes captured during an IVET screen have been exploited to identify the regulatory mechanisms controlling their expression. Overall, IVET and its various spin-offs have proven to be a valuable and robust set of tools for analysing microbial gene expression in complex environments and providing new targets for biotechnological development.

Variation in antibiotic-induced microbial recolonization impacts on the host metabolic phenotypes of rats

The interaction between the gut microbiota and their mammalian host is known to have far-reaching consequences with respect to metabolism and health. We investigated the effects of eight days of oral antibiotic exposure (penicillin and streptomycin sulfate) on gut microbial composition and host metabolic phenotype in male Han-Wistar rats (n = 6) compared to matched controls. Early recolonization was assessed in a third group exposed to antibiotics for four days followed by four days recovery (n = 6). Fluorescence in situ hybridization analysis of the intestinal contents collected at eight days showed a significant reduction in all bacterial groups measured (control, 1010.7 cells/g feces; antibiotic-treated, 108.4). Bacterial suppression reduced the excretion of mammalian-microbial urinary cometabolites including hippurate, phenylpropionic acid, phenylacetylglycine and indoxyl-sulfate whereas taurine, glycine, citrate, 2-oxoglutarate, and fumarate excretion was elevated. While total bacterial counts remained notably lower in the recolonized animals (109.1 cells/g faeces) compared to the controls, two cage-dependent subgroups emerged with Lactobacillus/Enterococcus probe counts dominant in one subgroup. This dichotomous profile manifested in the metabolic phenotypes with subgroup differences in tricarboxylic acid cycle metabolites and indoxyl-sulfate excretion. Fecal short chain fatty acids were diminished in all treated animals. Antibiotic treatment induced a profound effect on the microbiome structure, which was reflected in the metabotype. Moreover, the recolonization process was sensitive to the microenvironment, which may impact on understanding downstream consequences of antibiotic consumption in human populations.