Publications by authors named "Miguel J Beltrán-García"

9 Publications

  • Page 1 of 1

, an Endophyte That Establishes a Nutrient-Transfer Symbiosis With Banana Plants and Protects Against the Black Sigatoka Pathogen.

Front Microbiol 2019 7;10:804. Epub 2019 May 7.

Department of Chemistry, Universidad Autónoma de Guadalajara, Zapopan, Mexico.

Banana ( spp.) is an important crop worldwide, but black Sigatoka disease caused by the fungus threatens fruit production. In this work, we examined the potential of the endophytes of banana plants and , as antagonists of and support plant growth in nutrient limited soils by N-transfer. The two bacterial isolates were identified by MALDI-TOF mass spectrometry and corroborated by 16S rRNA sequence analysis. Both bacteria were positive for beneficial traits such as N-fixation, indole acetic acid production, phosphate solubilization, negative for 1-aminocyclopropane 1-carboxylic acid deaminase and were antagonistic to . To measure the effects on plant growth, the two plant bacteria and an strain (as non-endophyte), were inoculated weekly for 60 days as active cells (AC) and heat-killed cells (HKC) into plant microcosms without nutrients and compared to a water only treatment, and a mineral nutrients solution (MMN) treatment. Bacterial treatments increased growth parameters and prevented accelerated senescence, which was observed for water and mineral nutrients solution (MMN) treatments used as controls. Plants died after the first 20 days of being irrigated with water; irrigation with MMN enabled plants to develop some new leaves, but plants lost weight (-30%) during the same period. Plants treated with bacteria showed good growth, but AC treated plants had significantly greater biomass than the HKC. After 60 days, plants inoculated with AC showed intracellular bacteria within root cells, suggesting that a stable symbiosis was established. To evaluate the transference of organic N from bacteria into the plants, the 3 bacteria were grown with NHCl or NaNO as the nitrogen source. The N transferred from bacteria to plant tissues was measured by pheophytin isotopomer abundance. The relative abundance of the isotopomers 872.57, 873.57, 874.57, 875.57, 876.57 unequivocally demonstrated that plants acquired N atoms directly from bacterial cells, using them as a source of N, to support plant growth in restricted nutrient soils. might be a new alternative to promote growth and health of banana crops.
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http://dx.doi.org/10.3389/fmicb.2019.00804DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6513882PMC
May 2019

Role of Lipid Composition, Physicochemical Interactions, and Membrane Mechanics in the Molecular Actions of Microbial Cyclic Lipopeptides.

J Membr Biol 2019 06 16;252(2-3):131-157. Epub 2019 May 16.

Department of Chemistry ICET, Universidad Autónoma de Guadalajara, Patria 1201, Lomas del Valle, 45129, Zapopan, Jalisco, Mexico.

Several experimental and theoretical studies have extensively investigated the effects of a large diversity of antimicrobial peptides (AMPs) on model lipid bilayers and living cells. Many of these peptides disturb cells by forming pores in the plasma membrane that eventually lead to the cell death. The complexity of these peptide-lipid interactions is mainly related to electrostatic, hydrophobic and topological issues of these counterparts. Diverse studies have shed some light on how AMPs act on lipid bilayers composed by different phospholipids, and how mechanical properties of membranes could affect the antimicrobial effects of such compounds. On the other hand, cyclic lipopeptides (cLPs), an important class of microbial secondary metabolites, have received comparatively less attention. Due to their amphipathic structures, cLPs exhibit interesting biological activities including interactions with biofilms, anti-bacterial, anti-fungal, antiviral, and anti-tumoral properties, which deserve more investigation. Understanding how physicochemical properties of lipid bilayers contribute and determining the antagonistic activity of these secondary metabolites over a broad spectrum of microbial pathogens could establish a framework to design and select effective strategies of biological control. This implies unravelling-at the biophysical level-the complex interactions established between cLPs and lipid bilayers. This review presents, in a systematic manner, the diversity of lipidated antibiotics produced by different microorganisms, with a critical analysis of the perturbing actions that have been reported in the literature for this specific set of membrane-active lipopeptides during their interactions with model membranes and in vivo. With an overview on the mechanical properties of lipid bilayers that can be experimentally determined, we also discuss which parameters are relevant in the understanding of those perturbation effects. Finally, we expose in brief, how this knowledge can help to design novel strategies to use these biosurfactants in the agronomic and pharmaceutical industries.
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http://dx.doi.org/10.1007/s00232-019-00067-4DOI Listing
June 2019

In-vivo electrochemical monitoring of HO production induced by root-inoculated endophytic bacteria in Agave tequilana leaves.

Biosens Bioelectron 2018 Jan 18;99:108-114. Epub 2017 Jul 18.

Department of Fundamental Chemistry, Institute of Chemistry, University of São Paulo, Av. Prof. Lineu Prestes, 748, 05508-000 São Paulo, SP, Brazil. Electronic address:

A dual-function platinum disc microelectrode sensor was used for in-situ monitoring of HO produced in A. tequilana leaves after inoculation of their endophytic bacteria (Enterobacter cloacae). Voltammetric experiments were carried out from 0.0 to -1.0V, a potential range where HO is electrochemically reduced. A needle was used to create a small cavity in the upper epidermis of A. tequilana leaves, where the fabricated electrochemical sensor was inserted by using a manual three-dimensional micropositioner. Control experiments were performed with untreated plants and the obtained electrochemical results clearly proved the formation of HO in the leaves of plants 3h after the E. cloacae inoculation, according to a mechanism involving endogenous signaling pathways. In order to compare the sensitivity of the microelectrode sensor, the presence of HO was detected in the root hairs by 3,3-diaminobenzidine (DAB) stain 72h after bacterial inoculation. In-situ pH measurements were also carried out with a gold disc microelectrode modified with a film of iridium oxide and lower pH values were found in A. tequilana leaves treated with bacteria, which may indicate the plant produces acidic substances by biosynthesis of secondary metabolites. This microsensor could be an advantageous tool for further studies on the understanding of the mechanism of HO production during the plant-endophyte interaction.
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http://dx.doi.org/10.1016/j.bios.2017.07.039DOI Listing
January 2018

Nitrogen acquisition in Agave tequilana from degradation of endophytic bacteria.

Sci Rep 2014 Nov 6;4:6938. Epub 2014 Nov 6.

Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, 05508-000, São Paulo, SP, Brazil.

Plants form symbiotic associations with endophytic bacteria within tissues of leaves, stems, and roots. It is unclear whether or how plants obtain nitrogen from these endophytic bacteria. Here we present evidence showing nitrogen flow from endophytic bacteria to plants in a process that appears to involve oxidative degradation of bacteria. In our experiments we employed Agave tequilana and its seed-transmitted endophyte Bacillus tequilensis to elucidate organic nitrogen transfer from (15)N-labeled bacteria to plants. Bacillus tequilensis cells grown in a minimal medium with (15)NH4Cl as the nitrogen source were watered onto plants growing in sand. We traced incorporation of (15)N into tryptophan, deoxynucleosides and pheophytin derived from chlorophyll a. Probes for hydrogen peroxide show its presence during degradation of bacteria in plant tissues, supporting involvement of reactive oxygen in the degradation process. In another experiment to assess nitrogen absorbed as a result of endophytic colonization of plants we demonstrated that endophytic bacteria potentially transfer more nitrogen to plants and stimulate greater biomass in plants than heat-killed bacteria that do not colonize plants but instead degrade in the soil. Findings presented here support the hypothesis that some plants under nutrient limitation may degrade and obtain nitrogen from endophytic microbes.
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http://dx.doi.org/10.1038/srep06938DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4221784PMC
November 2014

Chemical management in fungicide sensitivity of Mycosphaerella fijiensis collected from banana fields in México.

Braz J Microbiol 2014 19;45(1):359-64. Epub 2014 May 19.

Departamento de Química Universidad Autónoma de Guadalajara GuadalajaraJalisco México.

The chemical management of the black leaf streak disease in banana caused by Mycosphaerella fijiensis (Morelet) requires numerous applications of fungicides per year. However this has led to fungicide resistance in the field. The present study evaluated the activities of six fungicides against the mycelial growth by determination of EC50 values of strains collected from fields with different fungicide management programs: Rustic management (RM) without applications and Intensive management (IM) more than 25 fungicide application/year. Results showed a decreased sensitivity to all fungicides in isolates collected from IM. Means of EC50 values in mg L(-1) for RM and IM were: 13.25 ± 18.24 and 51.58 ± 46.14 for azoxystrobin, 81.40 ± 56.50 and 1.8575 ± 2.11 for carbendazim, 1.225 ± 0.945 and 10.01 ± 8.55 for propiconazole, 220 ± 67.66 vs. 368 ± 62.76 for vinclozolin, 9.862 ± 3.24 and 54.5 ± 21.08 for fludioxonil, 49.2125 ± 34.11 and 112.25 ± 51.20 for mancozeb. A molecular analysis for β-tubulin revealed a mutation at codon 198 in these strains having an EC50 greater than 10 mg L(-1) for carbendazim. Our data indicate a consistency between fungicide resistance and intensive chemical management in banana fields, however indicative values for resistance were also found in strains collected from rustic fields, suggesting that proximity among fields may be causing a fungus interchange, where rustic fields are breeding grounds for development of resistant strains. Urgent actions are required in order to avoid fungicide resistance in Mexican populations of M. fijiensis due to fungicide management practices.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4059323PMC
http://dx.doi.org/10.1590/s1517-83822014000100051DOI Listing
February 2015

Singlet molecular oxygen generation by light-activated DHN-melanin of the fungal pathogen Mycosphaerella fijiensis in black Sigatoka disease of bananas.

PLoS One 2014 19;9(3):e91616. Epub 2014 Mar 19.

Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP, Brazil.

In pathogenic fungi, melanin contributes to virulence, allowing tissue invasion and inactivation of the plant defence system, but has never been implicated as a factor for host cell death, or as a light-activated phytotoxin. Our research shows that melanin synthesized by the fungal banana pathogen Mycosphaerella fijiensis acts as a virulence factor through the photogeneration of singlet molecular oxygen O2 (1Δg). Using analytical tools, including elemental analysis, ultraviolet/infrared absorption spectrophometry and MALDI-TOF mass spectrometry analysis, we characterized both pigment content in mycelia and secreted to the culture media as 1,8-dihydroxynaphthalene (DHN)-melanin type compound. This is sole melanin-type in M. fijiensis. Isolated melanins irradiated with a Nd:YAG laser at 532 nm produced monomol light emission at 1270 nm, confirming generation of O2 (1Δg), a highly reactive oxygen specie (ROS) that causes cellular death by reacting with all cellular macromolecules. Intermediary polyketides accumulated in culture media by using tricyclazole and pyroquilon (two inhibitors of DHN-melanin synthesis) were identified by ESI-HPLC-MS/MS. Additionally, irradiation at 532 nm of that mixture of compounds and whole melanized mycelium also generated O2 (1Δg). A pigmented-strain generated more O2 (1Δg) than a strain with low melanin content. Banana leaves of cultivar Cavendish, naturally infected with different stages of black Sigatoka disease, were collected from field. Direct staining of the naturally infected leaf tissues showed the presence of melanin that was positively correlated to the disease stage. We also found hydrogen peroxide (H2O2) but we cannot distinguish the source. Our results suggest that O2 (1Δg) photogenerated by DHN-melanin may be involved in the destructive effects of Mycosphaerella fijiensis on banana leaf tissues. Further studies are needed to fully evaluate contributions of melanin-mediated ROS to microbial pathogenesis.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0091616PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3960117PMC
December 2014

Cultivable endophytic bacteria from leaf bases of Agave tequilana and their role as plant growth promoters.

Braz J Microbiol 2014 4;45(4):1333-9. Epub 2015 Mar 4.

Departamento de Química Instituto de Ciencias Exactas y Terrestres Universidad Autónoma de Guadalajara Jalisco México Departamento de Química, Instituto de Ciencias Exactas y Terrestres, Universidad Autónoma de Guadalajara, Jalisco, México.

Agave tequilana Weber var. 'Azul' is grown for the production of tequila, inulin and syrup. Diverse bacteria inhabit plant tissues and play a crucial role for plant health and growth. In this study culturable endophytic bacteria were extracted from leaf bases of 100 healthy Agave tequilana plants. In plant tissue bacteria occurred at mean population densities of 3 million CFU/g of fresh plant tissue. Three hundred endophytic strains were isolated and 16s rDNA sequences grouped the bacteria into eight different taxa that shared high homology with other known sequences. Bacterial endophytes were identified as Acinectobacter sp., A. baumanii, A. bereziniae, Cronobacter sakazakii, Enterobacter hormaechei, Bacillus sp. Klebsiella oxytoca, Pseudomonas sp., Enterococcus casseliflavus, Leuconostoc mesenteroides subsp. mesenteroides and Gluconobacter oxydans. Isolates were confirmed to be plant growth promoting bacteria (PGPB) by their capacities for nitrogen fixation, auxin production, phosphate solubilization, or antagonism against Fusarium oxysporum AC132. E. casseliflavus JM47 and K. oxytoca JM26 secreted the highest concentrations of IAA. The endophyte Acinectobacter sp. JM58 exhibited the maximum values for nitrogen fixation and phosphate solubilization index (PSI). Inhibition of fungi was found in Pseudomonas sp. JM9p and K. oxytoca JM26. Bacterial endophytes show promise for use as bio-inoculants for agave cultivation. Use of endophytes to enhance cultivation of agave may be particularly important for plants produced by micropropagation techniques, where native endophytes may have been lost.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4323307PMC
http://dx.doi.org/10.1590/s1517-83822014000400025DOI Listing
October 2015

Oxidative stress response of Mycosphaerella fijiensis, the causal agent of black leaf streak disease in banana plants, to hydrogen peroxide and paraquat.

Can J Microbiol 2009 Jul;55(7):887-94

Departamento de Química ICET, Universidad Autónoma de Guadalajara, Patria 1201, Lomas del Valle 44100, Guadalajara, Jalisco, México.

Mycosphaerella fijiensis causes black leaf streak disease in banana and plantain. This fungus is usually attacked by reactive oxygen species secreted by the plant or during exposure to fungicide, however, little is known about the antioxidant response of the fungus. In this study, mycelia were observed to totally decompose 30 mmol/L of hydrogen peroxide (H2O2) within 120 min, liberating oxygen bubbles, and also to survive in concentrations as high as 100 mmol/L H2O2. The oxidative stress responses to H2O2, paraquat, and hydroquinone were characterized in terms of the activities of catalase and superoxide dismutase (SOD). Two active catalase bands were seen in native PAGE induced by H2O2. Band I had monofunctional activity and band II had bifunctional catalase-peroxidase activity. Two isozymes of SOD, distinguishable by their cyanide sensitivity, were found; CuZnSOD was the main one. The combination of H2O2 and 3-aminotriazole reduced the accumulation of biomass up to 40% compared with exposure to H2O2 alone, suggesting that catalase is important for the rapid decomposition of H2O2 and has a direct bearing on cell viability. The results also suggest that the superoxide anion formed through the redox of paraquat and hydroquinone has a greater effect than H2O2 on the cellular viability of M. fijiensis.
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http://dx.doi.org/10.1139/w09-023DOI Listing
July 2009

Biodegradation of viticulture wastes by Pleurotus: a source of microbial and human food and its potential use in animal feeding.

J Agric Food Chem 2002 Apr;50(9):2537-42

Centro de Investigación en Alimentación y Desarrollo, A.C. P.O. Box 1735, Hermosillo, Sonora, Mexico 83000.

The bioconversion of vineyard pruning and grape pomace by Pleurotus spp. using a solid state fermentation (SSF) was evaluated. Fruiting body production and chemical changes in the substrates after harvesting were measured. Biological efficiency and bioconversion ranged from 37.2 to 78.7% and from 16.7 to 38.8%, respectively. The best substrates for mycelial growth and mushroom yield were the mixtures with higher vineyard pruning content. Inclusion of pruning content had higher phenolic components and total sugars, better C/N ratio, and lower crude fat and total nitrogen than pomace. On the contrary, mycelium grew more slowly and scarcely in all treatments with 100% grape pomace. Moisture, protein, fat, and lignin contents were generally higher in mixtures with higher pomace proportion, whereas neutral detergent fiber, hemicellulose, and cellulose contents were higher with pruning content. Pleurotus strains may act depending on the availability of fiber fractions of substrate, and dynamic changes in digestion might occur as these fractions change during fungal growth. The recycling of viticulture residues through SSF by Pleurotus has great potential to produce human food and yields an available high-fiber feed for limited use in ruminants.
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http://dx.doi.org/10.1021/jf011308sDOI Listing
April 2002