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Orange processing waste (OPW) generated by the processing of oranges, as well as other citrus fruits, is a major source of pectin in the market nowadays. The residues generated during the pectin extraction process may contain many phytochemicals, including flavonoids. We use state-of-the-art techniques such as liquid chromatography high-resolution mass spectrometry (LC-HRMS/MS) and feature-based molecular network (FBMN) to annotate the flavonoids in OPWs. In particular, four flavonoids, hesperidin, naringin, diosmin, and hesperetin were quantified in the samples by LC-TDQ-MS. In total, 32 flavonoids from different classes were annotated, of which 16 were polymethoxylated flavonoids, 13 were flavonoid glycosides and 3 were flavanone aglycones. The results showed that flavonoid glycosides remain in high concentrations in OPWs from pectin factories even after pectin extraction by harsh conditions. The results show an exciting opportunity to harness the untapped potential of pectin factory waste as a renewable source for the extraction of glycoside flavonoids.
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Histoplasma capsulatum is the causative agent of histoplasmosis. Treating this fungal infection conventionally has significant limitations, prompting the search for alternative therapies. In this context, fungal extracellular vesicles (EVs) hold relevant potential as both therapeutic agents and targets for the treatment of fungal infections. To explore this further, we conducted a study using pharmacological inhibitors of chitinase (methylxanthines) to investigate their potential to reduce EV release and its subsequent impact on fungal virulence in an in vivo invertebrate model. Our findings revealed that a subinhibitory concentration of the methylxanthine, caffeine, effectively reduces EV release, leading to a modulation of H. capsulatum virulence. To the best of our knowledge, this is the first reported instance of a pharmacological inhibitor that reduces fungal EV release without any observed fungicidal effects.
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Metabolomics has been extensively used in clinical studies in the search for new biomarkers of human diseases. However, this approach has also been highlighted in agriculture and biological sciences, once metabolomics studies have been assisting researchers to deduce new chemical mechanisms involved in biological interactions that occur between microorganisms and plants. In this sense, the knowledge of the biological role of each metabolite (virulence factors, signaling compounds, antimicrobial metabolites, among others) and the affected biochemical pathways during the interaction contribute to a better understand of different ecological relationships established in nature. The current chapter addresses five different applications of the metabolomics approach in fungal-plant interactions research: (1) Discovery of biomarkers in pathogen-host interactions, (2) plant diseases diagnosis, (3) chemotaxonomy, (4) plant defense, and (5) plant resistance; using mass spectrometry and/or nuclear magnetic resonance spectroscopy, which are the techniques most used in metabolomics.
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Metabolómica , Plantas , Humanos , Metabolómica/métodos , Plantas/microbiología , Espectrometría de Masas/métodos , Biomarcadores/metabolismo , Hongos/metabolismoRESUMEN
The urgent need for new antimicrobials arises from antimicrobial resistance. Actinobacteria, especially Streptomyces genus, are responsible for production of numerous clinical antibiotics and anticancer agents. Genome mining reveals the biosynthetic gene clusters (BGCs) related to secondary metabolites and the genetic potential of a strain to produce natural products. However, this potential may not be expressed under laboratory conditions. In the present study, the Antarctic bacterium was taxonomically affiliated as Streptomyces albidoflavus ANT_B131 (CBMAI 1855). The crude extracts showed antimicrobial activity against both fungi, Gram-positive and Gram-negative bacteria and antiproliferative activity against five human tumor cell lines. Whole-genome sequencing reveals a genome size of 6.96 Mb, and the genome mining identified 24 BGCs, representing 13.3% of the genome. The use of three culture media and three extraction methods reveals the expression and recovery of 20.8% of the BGCs. The natural products identified included compounds, such as surugamide A, surugamide D, desferrioxamine B + Al, desferrioxamine E, and ectoine. This study reveals the potential of S. albidoflavus ANT_B131 as a natural product producer. Yet, the diversity of culture media and extraction methods could enhance the BGCs expression and recovery of natural products, and could be a strategy to intensify the BGC expression of natural products.
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Antiinfecciosos , Productos Biológicos , Streptomyces , Humanos , Antibacterianos/metabolismo , Bacterias Gramnegativas/genética , Bacterias Grampositivas/genética , Antiinfecciosos/metabolismo , Productos Biológicos/farmacología , Productos Biológicos/metabolismo , Medios de Cultivo/metabolismo , Familia de MultigenesRESUMEN
Granulomas are important immunological structures in the host defense against the fungus Paracoccidioides brasiliensis, the main etiologic agent of Paracoccidioidomycosis (PCM), a granulomatous systemic mycosis endemic in Latin America. We have performed transcriptional and proteomic studies of yeasts present in the pulmonary granulomas of PCM aiming to identify relevant genes and proteins that act under stressing conditions. C57BL/6 mice were infected with 1x106 yeasts and after 8- and 12-weeks of infection, granulomatous lesions were obtained for extraction of fungal and murine RNAs and fungal proteins. Dual transcriptional profiling was done comparing lung cells and P. brasiliensis yeasts from granulomas with uninfected lung cells and the original yeast suspension used in the infection, respectively. Mouse transcripts indicated a lung malfunction, with low expression of genes related to muscle contraction and organization. In addition, an increased expression of transcripts related to the activity of neutrophils, eosinophils, macrophages, lymphocytes as well as an elevated expression of IL-1ß, TNF-α, IFN-γ, IL-17 transcripts were observed. The increased expression of transcripts for CTLA-4, PD-1 and arginase-1, provided evidence of immune regulatory mechanisms within the granulomatous lesions. Also, our results indicate iron as a key element for the granuloma to function, where a high number of transcripts related to fungal siderophores for iron uptake was observed, a mechanism of fungal virulence not previously described in granulomas. Furthermore, transcriptomics and proteomics analyzes indicated a low fungal activity within the granuloma, as demonstrated by the decreased expression of genes and proteins related to energy metabolism and cell cycle.
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Paracoccidioides , Paracoccidioidomicosis , Animales , Ratones , Paracoccidioides/genética , Proteómica , Ratones Endogámicos C57BL , Hierro/metabolismo , Inmunidad , GranulomaRESUMEN
Microbial biostimulants have emerged as a sustainable alternative to increase the productivity and quality of important crops. Despite this, the effects of the treatment on plant metabolism are poorly understood. Thus, this study investigated the metabolic response of common bean (Phaseolus vulgaris) related to the treatment with a biostimulant obtained from the extract of Corynebacterium glutamicum that showed positive effects on the development, growth, and yield of crops previously. By untargeted metabolomic analysis using UHPLC-MS/MS, plants and seeds were subjected to treatment with the biostimulant. Under ideal growth conditions, the plants treated exhibited higher concentration levels of glutamic acid, nicotiflorin and glycosylated lipids derived from linolenic acid. The foliar application of the biostimulant under water stress conditions increased the chlorophyll content by 17% and induced the accumulation of flavonols, mainly quercetin derivatives. Also, germination seed assays exhibited longer radicle lengths for seeds treated compared to the untreated control even in the absence of light (13-18% increase, p-value <0.05). Metabolomic analysis of the seeds indicated changes in concentration levels of amino acids (tryptophan, phenylalanine, tyrosine, glutamine, and arginine) and their derivatives. The results point out the enhancement of abiotic stress tolerance and the metabolic processes triggered in this crop associated with the treatment with the biostimulant, giving the first insights into stress tolerance mechanisms in P. vulgaris.
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Corynebacterium glutamicum , Phaseolus , Phaseolus/química , Phaseolus/metabolismo , Phaseolus/microbiología , Espectrometría de Masas en Tándem , Estrés Fisiológico , Clorofila/metabolismoRESUMEN
Candida albicans is a commensal fungus in healthy humans that causes infection in immunocompromised individuals through the secretion of several virulence factors. The successful establishment of infection is owing to elaborate strategies to cope with defensive molecules secreted by the host, including responses toward oxidative stress. Extracellular vesicle (EV) release is considered an alternative to the biomolecule secretory mechanism that favors fungal interactions with the host cells. During candidiasis establishment, the host environment becomes oxidative, and it impacts EV release and cargo. To simulate the host oxidative environment, we added menadione (an oxidative stress inducer) to the culture medium, and we explored C. albicans EV metabolites by metabolomics analysis. This study characterized lipidic molecules transported to an extracellular milieu by C. albicans after menadione exposure. Through Liquid Chromatography coupled with Mass Spectrometry (LC-MS) analyses, we identified biomolecules transported by EVs and supernatant. The identified molecules are related to several biological processes, such as glycerophospholipid and sphingolipid pathways, which may act at different levels by tuning compound production in accordance with cell requirements that favor a myriad of adaptive responses. Taken together, our results provide new insights into the role of EVs in fungal biology and host-pathogen interactions.
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Germination and enzymatic hydrolysis are biological processes with well-recognized positive effects on phenolic composition and antioxidant potential. This study aimed to apply those processes to white (Sinapsis alba) and black (Brassica nigra) mustard grains and to analyze the influences on the total phenolic content (TPC); phenolic and peptide profile determined by ultra-performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS); antioxidant potential (DPPH, ABTS, and FRAP assays); and cytotoxicity against Caco-2, a human colorectal adenocarcinoma cell line. Enzyme combinations for hydrolysis were different for each mustard grain, but for both species, enzymatic hydrolysis and germination showed a positive effect on antioxidant properties. From UPLC-HRMS analysis and molecular network studies, 14 peptides and 17 phenolic compounds were identified as metabolites released from mustard after processes application, which were strongly correlated with increased antioxidant activity. In addition, enzymatic hydrolysis applied in germinated mustard grains for both mustards increased the cytotoxic activity against Caco-2 human colorectal adenocarcinoma cell line.
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Antioxidantes , Planta de la Mostaza , Humanos , Antioxidantes/análisis , Planta de la Mostaza/química , Células CACO-2 , Hidrólisis , Fenoles/análisis , Semillas/química , BiotransformaciónRESUMEN
Rhizobacteria are valuable sources of compounds that can be used for the integrated management of diseases in rice. Here, we aimed to explore the metabolism and organize and annotate the metabolites of Bacillus sp. and Serratia marcescens isolates using molecular networking and evaluate their fungicidal effects against Magnaporthe oryzae and Bipolaris oryzae. We obtained bacterial extracts after 6 and 16-h incubation via liquid-liquid extraction using ethyl acetate as solvent. We performed UHPLC-MS analysis and data processing using molecular networking and conducted biological assays in rice plants. Using the Global Natural Product Social spectral libraries, we annotated the following compounds: austinoneol, Phe-Pro, N-acetyl-l-leucine, Leu-Gly, Ile-Leu, Phe-Pro, 2,5-piperazinedione, 3-(1H-indol-3-methyl)-6-methyl-cyclo(d-Trp-l-Pro), and cholic acid. Results of the biological assays showed that the bacterial extracts reduced the mycelial growth of both pathogens in all treatments compared to the control. In the greenhouse setup, 8 days after the challenge for leaf gray spot and leaf blast, all treatments affected up to 4.4% of the leaf area, with an area under disease progress curve of 13.24, showing significant difference compared to the control, which affected 23% of the leaf area, with an AUDPC of 44.65. Our study provides potential new sources of natural products to be applied in the integrated management of rice. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03547-6.
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BACKGROUND: Lignin is an attractive alternative for producing biobased chemicals. It is the second major component of the plant cell wall and is an abundant natural source of aromatic compounds. Lignin degradation using microbial oxidative enzymes that depolymerize lignin and catabolize aromatic compounds into central metabolic intermediates is a promising strategy for lignin valorization. However, the intrinsic heterogeneity and recalcitrance of lignin severely hinder its biocatalytic conversion. In this context, examining microbial degradation systems can provide a fundamental understanding of the pathways and enzymes that are useful for lignin conversion into biotechnologically relevant compounds. RESULTS: Lignin-degrading catabolism of a novel Rhodosporidium fluviale strain LM-2 was characterized using multi-omic strategies. This strain was previously isolated from a ligninolytic microbial consortium and presents a set of enzymes related to lignin depolymerization and aromatic compound catabolism. Furthermore, two catabolic routes for producing 4-vinyl guaiacol and vanillin were identified in R. fluviale LM-2. CONCLUSIONS: The multi-omic analysis of R. fluviale LM-2, the first for this species, elucidated a repertoire of genes, transcripts, and secreted proteins involved in lignin degradation. This study expands the understanding of ligninolytic metabolism in a non-conventional yeast, which has the potential for future genetic manipulation. Moreover, this work unveiled critical pathways and enzymes that can be exported to other systems, including model organisms, for lignin valorization.
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Most of the biosynthetic gene clusters (BGCs) found in microbes are silent under standard laboratory cultivation conditions due to the lack of expression triggering stimuli, representing a considerable drawback in drug discovery. To access the full biosynthetic potential, studies towards the activation of cryptic BGCs are essential. Histone acetylation status is an important regulator of chromatin structure, which impacts cell physiology and the expression of BGCs. In this study, clr3, a gene encoding a histone deacetylase in Penicillium brasilianum LaBioMMi 136, is deleted and associated phenotypic and metabolic changes are evaluated. The results indicate reduced growth under oxidative stress conditions in the ∆clr3 strain, higher intracellular reactive oxygen species (ROS) levels, and a different transcriptional profile of 13 ROS-related genes of both strains under basal and ROS-induced conditions. Moreover, the production of 14 secondary metabolites, including austin-related meroterpenoids, brasiliamides, verruculogen, penicillic acid, and cyclodepsipeptides was evaluated in the ∆clr3 strain, most of them being reduced. Accordingly, the addition of epigenetic modulators responsible for HDAC inhibition into P. brasilianum's growth media also culminated in the reduction in secondary metabolite production. The results suggest that Clr3 plays an essential role in secondary metabolite biosynthesis in P. brasilianum, thus offering new strategies for the regulation of natural product synthesis by assessing chromatin modification.
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The biocatalytic production of fuels and chemicals from plant biomass represents an attractive alternative to fossil fuel-based refineries. In this context, the mining and characterization of novel biocatalysts can promote disruptive innovation opportunities in the field of lignocellulose conversion and valorization. In the present work, we conducted the biochemical and structural characterization of two novel hydroxycinnamic acid catabolic enzymes, isolated from a lignin-degrading microbial consortium, a feruloyl-CoA synthetase, and a feruloyl-CoA hydratase-lyase, named LM-FCS2 and LM-FCHL2, respectively. Besides establishing the homology model structures for novel FCS and FCHL members with unique characteristics, the enzymes presented interesting biochemical features: LM-FCS2 showed stability in alkaline pHs and was able to convert a wide array of p-hydroxycinnamic acids to their respective CoA-thioesters, including sinapic acid; LM-FCHL2 efficiently converted feruloyl-CoA and p-coumaroyl-CoA into vanillin and 4-hydroxybenzaldehyde, respectively, and could produce vanillin directly from ferulic acid. The coupled reaction of LM-FCS2 and LM-FCHL2 produced vanillin, not only from commercial ferulic acid but also from a crude lignocellulosic hydrolysate. Collectively, this work illuminates the structure and function of two critical enzymes involved in converting ferulic acid into high-value molecules, thus providing valuable concepts applied to the development of plant biomass biorefineries. KEY POINTS: ⢠Comprehensive characterization of feruloyl-CoA synthetase from metagenomic origin. ⢠Novel low-resolution structures of hydroxycinnamate catabolic enzymes. ⢠Production of vanillin via enzymatic reaction using lignocellulosic hydrolysates.
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Lignina , Metagenoma , Escherichia coli/genética , Hiperlipidemia Familiar Combinada , Lignina/metabolismo , SueloRESUMEN
Citrus fruits are the most produced fruits in the world, but they are threatened by several pathogens, including the fungus Phyllosticta citricarpa, the causal agent of citrus black spot (CBS). The fungus affects most citrus species and the infection results in economic losses in citrus-producing areas. This disease causes the aesthetic depreciation of fresh fruit, impairing its commercialization. As an alternative to the use of synthetic fungicides to control the pathogen, the biological control, using bacteria of the genus Bacillus, is highlighted. Such microorganisms enable biocontrol by the production of volatile organic compounds (VOC) or non-volatile. Therefore, this work aimed to evaluate the production of VOC by isolates of Bacillus spp. grown in different culture media; to evaluate the effects of these compounds on the evolution of CBS lesions in orange fruits; to study the effects of VOC on resistance induction in orange fruits; to evaluate the effects of VOC on P. citricarpa morphology in CBS lesions, and to identify the produced VOC. Tryptone soya agar (TSA) and tryptone soya broth (TSB) media used to culture the bacterium resulted in up to 73% pathogen inhibition by VOC. Volatile compounds from Bacillus spp. ACB-65 and Bacillus spp. ACB-73 when cultured in TSB culture medium provided 86% inhibition of freckles that evolved to hard spots. The volatile fractions produced by the bacteria were identified as alcohols, ketones, amines, ethers, aldehydes and carboxylic acids that can serve as arsenal against the phytopathogen. The present work demonstrated the potential of VOC produced by Bacillus spp. in the control of P. citricarpa.
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Ascomicetos/patogenicidad , Bacillus , Agentes de Control Biológico , Citrus , Enfermedades de las Plantas/prevención & control , Bacillus/fisiología , Citrus/microbiología , Interacciones Microbianas , Enfermedades de las Plantas/microbiología , Esporas FúngicasRESUMEN
Microbial pigments have a distinguished potential for applications in food and pharmaceutical industries, stimulating the research in this field. The present study evaluated the ideal conditions for extracting bikaverin (red pigment) from the biomass of Fusarium oxysporum CCT7620. Among the solvents tested, ethyl acetate extraction resulted in the highest bikaverin concentration and the kinetic study revealed a saturation in bikaverin concentration from 256 min on. Based on a preliminary economic study, three sequential extractions with ethyl acetate was considered the ideal protocol to recover bikaverin. After extraction, chromatographic methods were tested to purify bikaverin. The use of silica gel or Sephadex (open column) could not successfully purify bikaverin, but the semi-preparative HPLC resulted in a bikaverin-enriched fraction with a purity degree equivalent to the commercial analytical standard. This work provides relevant information regarding the extraction and purification of bikaverin, which may be useful for other downstraming processes.
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Aspergillus fumigatus produces diverse secondary metabolites whose biological functions and regulation remain to be understood. Despite the importance of the conidia for this fungus, the role of the conidia-born metabolite fumiquinazoline C (FqC) is unclear. Here, we describe a dual function of the cell-wall integrity pathway in regulating FqC biosynthesis dictated by the MAPK kinase MpkA, which phosphorylates one of the nonribosomal peptide synthetases enzymes of the cluster (FmqC), and the transcription factor RlmA, which directly regulates the expression of fmq genes. Another level of crosstalk between the FqC regulation and the cell physiology is described since the deletion of the stress-responsive transcription factor sebA provokes derepression of the fmq cluster and overproduction of FqC. Thus, we describe a mechanism by which A. fumigatus controls FqC biosynthesis orchestrated by MpkA-RlmA and SebA and hence enabling survival and adaptation to the environmental niche, given that FqC is a deterrent of ameba predation.
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Aspergillus fumigatus/genética , Quinazolinas/metabolismo , Aspergillus fumigatus/metabolismo , Pared Celular/genética , Proteínas Fúngicas/genética , Expresión Génica , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Fagocitosis/fisiología , Transducción de Señal , Esporas Fúngicas/genética , Esporas Fúngicas/metabolismo , Transcripción GenéticaRESUMEN
Penicillium digitatum is the most aggressive pathogen of citrus fruits. Tryptoquialanines are major indole alkaloids produced by P. digitatum It is unknown if tryptoquialanines are involved in the damage of citrus fruits caused by P. digitatum. To investigate the pathogenic roles of tryptoquialanines, we initially asked if tryptoquialanines could affect the germination of Citrus sinensis seeds. Exposure of the citrus seeds to tryptoquialanine A resulted in a complete inhibition of germination and an altered metabolic response. Since this phytotoxic effect requires the extracellular export of tryptoquialanine A, we investigated the mechanisms of extracellular delivery of this alkaloid in P. digitatum We detected extracellular vesicles (EVs) released by P. digitatum both in culture and during infection of citrus fruits. Compositional analysis of EVs produced during infection revealed the presence of a complex cargo, which included tryptoquialanines and the mycotoxin fungisporin. The EVs also presented phytotoxicity activity in vitro and caused damage to the tissues of citrus seeds. Through molecular networking, it was observed that the metabolites present in the P. digitatum EVs are produced in all of its possible hosts. Our results reveal a novel phytopathogenic role of P. digitatum EVs and tryptoquialanine A, implying that this alkaloid is exported in EVs during plant infection.IMPORTANCE During the postharvest period, citrus fruits can be affected by phytopathogens such as Penicillium digitatum, which causes green mold disease and is responsible for up to 90% of total citrus losses. Chemical fungicides are widely used to prevent green mold disease, leading to concerns about environmental and health risks. To develop safer alternatives to control phytopathogens, it is necessary to understand the molecular basis of infection during the host-pathogen interaction. In the P. digitatum model, the virulence strategies are poorly known. Here, we describe the production of phytotoxic extracellular vesicles (EVs) by P. digitatum during the infection of citrus fruits. We also characterized the secondary metabolites in the cargo of EVs and found in this set of molecules an inhibitor of seed germination. Since EVs and secondary metabolites have been related to virulence mechanisms in other host-pathogen interactions, our data are important for the comprehension of how P. digitatum causes damage to its primary hosts.
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Alcaloides/metabolismo , Alcaloides/farmacología , Citrus/microbiología , Vesículas Extracelulares/química , Penicillium/patogenicidad , Semillas/crecimiento & desarrollo , Alcaloides/biosíntesis , Frutas/microbiología , Fungicidas Industriales/farmacología , Interacciones Huésped-Patógeno , Enfermedades de las Plantas/microbiología , Metabolismo Secundario , Semillas/efectos de los fármacos , Semillas/metabolismo , Semillas/microbiologíaRESUMEN
The exponential growth of plastic consumption in the last decade became a large economic and ecological issue; therefore, strategies have been used to mitigate the environmental impacts, including the manufacture of biodegradable bio-based plastics and biodegradation strategies. Herein, a new bio-based plastic was developed consisting of a polymeric recyclable matrix (polyethylene or polypropylene) with a vegetal polymeric material from cocoa husk. Mechanical and rheological properties were evaluated and the new material showed interesting tensile strength compared to completely non-biodegradable plastics. The new polymeric material was submitted to biodegradation processes using different fungi species. The biodegradation caused by Colletotrichum gloeosporioides, Xylaria sp. and Fusarium graminearum in the new polymeric material was analyzed through scanning electron microscopy with energy-dispersive X-ray spectroscopy (SEM-EDS) and tensile tests. Furthermore, ultra performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) and mass spectrometry imaging (MSI) were applied to identify metabolites produced in consequence to the biodegradation process. Interestingly, some compounds produced present high economic value.
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In the agricultural sector, citrus is one of the most important fruit genus in the world. In this scenario, Brazil is the largest producer of oranges; 34% of the global production, and exporter of concentrated orange juice; 76% of the juice consumed in the planet, summing up US$ 6.5 billion to Brazilian GDP. However, the orange production has been considerable decreasing due to unfavorable weather conditions in recent years and the increasing number of pathogen infections. One of the main citrus post-harvest phytopathogen is Penicillium italicum, responsible for the blue mold disease, which is currently controlled by pesticides, such as Imazalil, Pyrimethanil, Fludioxonil, and Tiabendazole, which are toxic chemicals harmful to the environment and also to human health. In addition, P. italicum has developed considerable resistance to these chemicals as a result of widespread applications. To address this growing problem, the search for new control methods of citrus post-harvest phytopathogens is being extensively explored, resulting in promising new approaches such as biocontrol methods as "killer" yeasts, application of essential oils, and antimicrobial volatile substances. The alternative methodologies to control P. italicum are reviewed here, as well as the fungal virulence factors and infection strategies. Therefore, this review will focus on a general overview of recent research carried out regarding the phytopathological interaction of P. italicum and its citrus host.
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Huanglongbing (HLB) is a disease of worldwide incidence that affects orange trees, among other commercial varieties, implicating in great losses to the citrus industry. The disease is transmitted through Diaphorina citri vector, which inoculates Candidatus Liberibacter spp. in the plant sap. HLB disease lead to blotchy mottle and fruit deformation, among other characteristic symptoms, which induce fruit drop and affect negatively the juice quality. Nowadays, the disease is controlled by eradication of sick, symptomatic plants, coupled with psyllid control. Polymerase chain reaction (PCR) is the technique most used to diagnose the disease; however, this methodology involves high cost and extensive sample preparation. Mass spectrometry imaging (MSI) technique is a fast and easily handled sample analysis that, in the case of Huanglongbing allows the detection of increased concentration of metabolites associated to the disease, including quinic acid, phenylalanine, nobiletin and sucrose. The metabolites abieta-8,11,13-trien-18-oic acid, suggested by global natural product social molecular networking (GNPS) analysis, and 4-acetyl-1-methylcyclohexene showed a higher distribution in symptomatic leaves and have been directly associated to HLB disease. Desorption electrospray ionization coupled to mass spectrometry imaging (DESI-MSI) allows the rapid and efficient detection of biomarkers in sweet oranges infected with Candidatus Liberibacter asiaticus and can be developed into a real-time, fast-diagnostic technique.
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Citrus/microbiología , Espectrometría de Masas/métodos , Hojas de la Planta/química , Animales , Citrus/crecimiento & desarrollo , Citrus/metabolismo , Ciclohexanos/análisis , ADN Bacteriano/química , Diagnóstico , Vectores de Enfermedades , Hemípteros/genética , Enfermedades de las Plantas/etiología , Reacción en Cadena de la Polimerasa/métodosRESUMEN
Flavonoids are involved in citrus defense against phytopathogens. In this study, we applied in vitro biocatalysis assays using the flavanones glycosides hesperidin and naringin to explore the enzymatic activities involved in such interaction. The main enzymatic activity observed was the hydrolysis catalyzed by fungi naringinases and hesperidinases. Withing 7 days, the two citrus phytopathogenic fungi, Penicillium digitatum and Penicillium italicum, exhibited the highest hydrolyzing rate on the flavanones, reaching conversion values higher than 90%. In addition, Geothrichum citri-aurantii exhibited no enzymatic activity and Penicillium expansum only hydrolyzed hesperidin. In order to evaluate flavonoid biotransformation by the fungi in vivo, citrus fruits infected with P. digitatum were analyzed through molecular networking and Imaging Mass Spectrometry (IMS). In vivo assays revealed that citrus fruit in response to the infection is able to hydroxylate flavonoids, and novel flavonoid structures were associated to the citrus' defense. The data reported here present a new point of view in the relation between citrus flavonoids and phytopathogenic fungi and can be useful to understand the infection processes and host-pathogen interaction.