Microbiologia Agrícola

URI permanente para esta coleçãohttps://locus.ufv.br/handle/123456789/190

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2020 - 202390

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Data inicial: 2020Data final: 2023

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Agora exibindo 1 - 10 de 90
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    Desenvolvimento de teste rápido imunocromatográfico de fluxo lateral para neosporose bovina
    (Universidade Federal de Viçosa, 2023-07-17) Pinto, Maria Eduarda Almeida; Mendes, Tiago Antônio de Oliveira; http://lattes.cnpq.br/9379279240915140
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    Otimização da produção de lipopeptídeos por Bacillus velezensis visando ao controle de fungos fitopatogênicos
    (Universidade Federal de Viçosa, 2023-09-13) Félix, Jéssica da Silva; Tótola, Marcos Rogério
    O gênero Bacillus destaca-se como um dos mais empregados no biocontrole de doenças de plantas. Esses microrganismos são encontrados em diversos ambientes, sendo reconhecidos por sua capacidade de produzir lipopeptídeos. Essas moléculas são caracterizadas como biossurfactantes de baixo peso molecular e exibem atividade antibacteriana, antifúngica, antiviral, cicatrizante e antitumoral. Os lipopeptídeos se dividem em três famílias principais: surfactina, iturina e fengicina, sendo que as duas últimas apresentam elevada atividade antifúngica. Este trabalho teve como objetivo definir condições de cultivo adequadas para a produção de lipopeptídeos com atividade antifúngica por três isolados de Bacillus velezensis (LBBMA TR59II, LBBMA TR47II e LBBMA AP01), a fim de promover o controle de fungos fitopatogênicos. Para isso, realizou-se a otimização da produção de lipopeptídeos dos isolados em três diferentes meios de cultura (Meio Farelo de Soja adicionado de açúcar cristal, Meio 523 e Meio Mineral). Posteriormente, diferentes combinações de pH e temperatura foram investigadas. O fungo fitopatogênico modelo adotado no trabalho foi o Colletotrichum truncatum isolado UCBV258. Os estudos de otimização foram conduzidos utilizando-se o método de Delineamento Composto Central Rotacional (DCCR). O meio de cultura Meio 523 propiciou as maiores produções de lipopeptídeos para B. velezensis LBBMA TR59ll e LBBMA TR47ll; para B. velezensis LBBMA AP01, as maiores produções de lipopeptídeos foram obtidas em Meio Mineral. Ao se considerar o custo dos ingredientes para a produção dos meios de cultivo, o Meio Farelo de Soja se mostrou o mais indicado para a produção de lipopeptídeos com ação fungicida para todos os isolados de B. velezensis estudados. As combinações das variáveis concentração de farelo de soja, de açúcar cristal, pH e temperatura propiciaram ganhos de eficiência de produção de lipopeptídeos (medida pela razão entre a produção de extrato bruto de lipopeptídeos liofilizado – EBEL - e a concentração inibitória mínima dos extratos brutos obtidos em cada condição) de até 59 vezes para o isolado B. velezensis LBBMA TR59ll, 17 vezes para B. velezensis LBBMA TR47ll 16 vezes para B. velezensis LBBMA AP01. Os ganhos de eficiência reportados, juntamente com o baixo custo dos ingredientes empregados no preparo dos meios de cultivo, têm o potencial de reduzir significativamente os custos deprodução de bioinsumos à base de Bacillus spp. visando ao controle de patógenos agrícolas. Palavras-chave: Bacillus velezensis. Lipopeptídeos. Surfactina. Fengicina. Iturina. Colletotrichum truncatum. Controle biológico.
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    Exploring RNA virus diversity in Metarhizium spp
    (Universidade Federal de Viçosa, 2023-07-26) Oliveira, Cauê Neves; Zerbini, Poliane Alfenas; http://lattes.cnpq.br/8002246970195948
    Viruses are ubiquitous acellular organisms since there is evidence of them hosting any living cell in all kingdoms. Mycoviruses are characterized by infecting fungi. The International Committee on Taxonomy of Virus (ICTV) recently recognized 29 families with representatives of mycoviruses. Most of these families have an RNA genome, and only one has a circular ssDNA genome. The non-retroviral RNA viruses are sorted into three Baltimore groups. Group III is characterized by having a dsRNA genome, while groups IV and V correspond to a ssRNA positive and negative sense, respectively. The unique hallmark gene in these three groups is RNA-dependent RNA polymerase (RdRp). This protein is responsible for viral genome replication and is conserved in all groups. Moreover, mycovirus infection can change the host phenotype. Most of the time, these changes are related to modification of their host pathogenic trait. For example, hypovirulent-associated mycoviruses are characterized by reducing the aggressiveness of phytopathogenic fungi by slowing down their mycelial growth and sporulation rate, as well as downregulating pathogenic genes. On the other hand, hypervirulent-associated mycoviruses are characterized by promoting mycelial growth and sporulation rate and upregulating host pathogenic genes. Therefore, mycoviruses are a more eco-friendly alternative to pest-management strategies than the use of chemical compounds related to ecosystem harm. Hypovirulent-associated mycovirus can be used to control the incidence of phytopathogenic fungi, and hypervirulent-associated mycovirus can increase the efficacy of mycopesticides. Studies already demonstrate that hypervirulent-associated mycoviruses hosting entomopathogenic fungi (EPF) can increase their potential to control insect pests. This work used two approaches to identify novel mycoviruses infecting the EPF in the genus Metarhizium. The first strategy was carried out by the total nucleic acid extraction of a Metarhizium robertsii isolate SCJAN-21.11 followed by identifying and sequencing the dsRNA viral genome. As we know, we report the first case of a polymycovirus hosting a Metarhizium in Brazil. This polymycovirus has at least three dsRNA genome segments. Each segment encodes only one ORF representing the RdRp, a hypothetical protein, and a methyltransferase. The RdRp aminoacid phylogenetic analyses revealed that this mycovirus represents a new species, and we attempted to call it Polymycovirus mineiro. In the second strategy, we used a database with 77 viral family-level profile Hidden Markov Models (pHMM) to look for RdRp-like sequences in the public RNAseq data available on NCBI tagged as “Metarhizium.” The pHMM is a statistical model to identify sequence patterns. As a result, we could identify 42 virus-like sequences in approximately 20% of the analyzed data. In conclusion, this study helps elucidate the virosphere in Metarhizium genera and investigate the possibility of using mycovirus as a biocontrol agent. Keywords: Mycovirus; Biocontrol; Entomopathogenic fungi; Metarhizium
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    LIPOPEPTÍDEOS PRODUZIDOS POR Bacillus subtilis LBBMA AP01 E SEU POTENCIAL DE APLICAÇÃO COMO AGENTES ANTIFÚNGICOS
    (Universidade Federal de Viçosa, 2022-08-26) Paula, Maria Theresa Rafaela; Tótola, Marcos Rogério; 9908682517903478
    O Brasil se destaca como um dos principais produtores agrícolas mundiais, contudo as perdas causadas por doenças e pragas agrícolas e os elevados custos para seu controle são limitantes a alta produtividade. Fungos fitopatógenos causam sérios danos às culturas e demandam o uso de compostos, em sua maioria químicos, para seu controle. Visando investigar estratégias de controle menos danosas ao ambiente e à saúde humana, o presente trabalho tem como objetivos a obtenção dos lipopeptídeos produzidos por Bacillus subtilis LBBMA AP01 e a caracterização dos mesmos quanto às propriedades tensoativas e potencial de inibir o desenvolvimento de fungos fitopatógenos isoladamente e em associação com nanomateriais de carbono, prata e ouro. Os pontos quânticos de carbono e nanopartículas metálicas foram produzidos pela síntese pirolítica e química, respectivamente, e posteriormente funcionalizados com os lipopeptídeos. O potencial de inibição do crescimento micelial dos lipopeptídeos foi avaliado a partir do ensaio em placa para Sclerotinia sclerotiorum, Botrytis cinerea, Fusarium sp. e Rhizoctonia solani em meio Ágar Batata Dextrose (BDA) acrescido dos lipopeptídeos em concentrações entre 20 e 1000 mg L-1. A atividade antifúngica dos lipopeptídeos e nanomateriais contra Botrytis cinerea foi avaliada in vivo em folhas destacadas de tomate e repolho. Adicionalmente, foi avaliada a inibição da germinação de conídios de B. cinerea em microplaca. A atividade herbicida dos lipopeptídeos foi investigada em sementes de modelos de mono e dicotiledônea. Os lipopeptídeos foram capazes de inibir todos os fitopatógenos e a concentração mínima inibitória (CMI) para todos os isolados foi de 80 mg L -1, exceto para Fusarium sp., cuja CMI foi 40 mg L-1. O extrato bruto a 500 e 1000 mg L-1 inibiu 100% a germinação de conídios de B. cinerea. Os lipopeptídeos não apresentaram efeito inibitório considerável sobre a germinação de sementes. Os tratamentos preventivos se mostraram eficientes no controle dos fitopatógenos. Embora B. cinerea tenha exibido sensibilidade a algumas formulações de nanopartículas e pontos quânticos de carbono, não foi possível determinar o potencial de inibição dos nanomateriais in vivo. Conclui-se que os lipopeptídeos são capazes de inibir todos os fungos fitopatógenos testados de forma eficiente e que seu uso na funcionalização de nanopartículas pode potencializar este efeito. Deste modo, este trabalho evidencia o potencial dos lipopeptídeos produzidos por Bacillus subtilis AP01 e sua versatilidade de aplicações e associações com outros materiais, para o controle de fitopatógenos. Palavras-chave: Biossurfactantes. Sclerotinia sclerotiorum. Botrytis cinérea. Rhizoctonia solani. Fusarium sp. Nanomateriais.
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    Genômica comparativa possibilita a identificação de fatores de virulência no genoma de Colletotrichum lindemuthianum
    (Universidade Federal de Viçosa, 2021-02-01) Correia, Hilberty Lucas Nunes; Queiroz, Marisa Vieira de; 5718975445923498
    O fungo Colletotrichum lindemuthianum é um fitopatógeno hemibiotrófico, responsável pela antracnose do feijoeiro-comum (Phaseolus vulgaris L.). Entre as características mais marcantes do C. lindemuthianum destacam-se o alto poder destrutivo e sua ampla varabilidade patogênica. O objetivo do presente trabalho foi realizar o estudo dos genomas pertencentes aos isolados 83.501 (raça 83) e A2-2-3 (raça 89), de C. lindemuthianum por meio da análise comparativa, com o intuito de avaliar as suas características genômicas e identificar fatores de virulência envolvidos na infecção de plantas de feijoeiro. Considerando somente os dados de genomas obtidos com tecnologias de sequenciamento de primeira e segunda geração, C. lindemuthianum apresenta o maior percentual de elementos genéticos móveis já predito para os genomas de Colletotrichum spp. Esta espécie também apresentou os maiores índices de ocorrência de repeat-induced point mutation (RIP) dentre todas as espécies de Colletotrichum avaliadas, possuindo os genes que codificam as metiltransferases RID e DIM-2, que participam deste mecanismo silenciamento. Colletotrichum lindemuthianum possui toda a maquinaria necessária a ocorrência do ciclo sexual, no entanto, o locus Mat1-1 não foi encontrado em seu genoma, bem como no genoma das demais espécies do gênero avaliadas. Foram identificados 55 genes no genoma do isolado 83.501 que estão ausentes no genoma do isolado A2-2-3 e 41 genes do isolado A2-2-3 que estão ausentes no genoma do isolado 83.501. Os genomas de C. lindemuthianum apresentam alterações significativas no número de cópias de genes pertencentes a 20 famílias de P450 mono-oxigenases, 10 famílias de CAZymes envolvidas na degradação de material vegetal e em 99 tipos de transportadores quando comparados às demais espécies de Colletotrichum. Dentre as espécies do gênero Colletotrichum analisadas, o fungo C. ivlindemuthianum é a espécie com as características genômicas mais distintas. Palavras chave: patogenicidade, adaptação, secretoma.
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    Selection of oleaginous yeasts able to accumulate lipids from culture media containing glycerol as the sole carbon source
    (Universidade Federal de Viçosa, 2023-08-30) Abimbola, Olufemi Gabriel; Silveira, Wendel Batista da; http://lattes.cnpq.br/0122511141140643
    The biodiesel is currently produced mainly from edible oils as feedstock, generating glycerol as a by-product. The use of glycerol as a carbon source for oil production by oleaginous yeasts contributes to the development of alternative routes of oil production in the context of the circular economy. Therefore, this study aimed to select oleaginous yeasts capable of producing oil in culture media containing glycerol as a carbon source. A total of 10 yeasts capable of assimilating glycerol as carbon source were evaluated for their ability to produce lipids in a glycerol culture medium to select a promising oleaginous yeast. Herein, the isolate CA8 was selected as the best lipid producer from glycerol considering the achieved values of lipid content (%), lipid titer (g/L), and biomass (g/L). This isolate was taxonomically identified as Meyerozyma guilliermondii and designated as Meyerozyma guilliermondii UFV-2. Since the C: N ratio notably influences lipid production by oleaginous yeasts, the effect of different C:N ratios on biomass production, lipid content and lipid titer were evaluated. The highest values of biomass formation, lipid content, and lipid titer, recorded in a C: N of 150:1, were 6.77 g/L, 25 % (w/w), and 1.71 g/L, respectively recorded. The lowest biomass formation 4.67 g/L was recorded in a C: N ratio of 75:1. Taken together, these results indicate that M. guilliermondii UFV-2 displays the potential to produce lipids from glycerol, opening perspectives for further studies that aim to optimize the production parameters. Keywords: Biodiesel; Meyerozyma guilliermondii; Yeast oil; Circular economy; Carbon- nitrogen ratios.
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    Potential use of multiplex real-time PCR to develop a kit for identification of beer-spoilage microorganisms
    (Universidade Federal de Viçosa, 2022-09-29) Machado, Túlio Iglésias; Tótola, Marcos Rogério; http://lattes.cnpq.br/9358020634360780
    Beer is the most consumed alcoholic beverage globally. Despite being considered a microbiologically stable beverage due to its intrinsic properties such as low pH, high CO2, presence of antimicrobial compounds from hop and other factors, microbiological contamination in beer does happen, leading to off-flavor production with changes in flavor and aroma, viscosity, acidification, among other unwanted effects. This study explores the use of multiplex real-time PCR (qPCR) coupled to High Resolution Melting (HRM) analysis for the simultaneous detection and discrimination of beer-spoilage microorganisms genera. Orthologous sequences were identified using the OrthoMCL pipeline for primer design. The designed primers exhibited high specificity, generating distinct melting peaks for the target genera. Sensitivity was confirmed, with successful amplification at low DNA concentrations. The perfect alignment of primers with target regions significantly influenced sensitivity. The multiplex qPCR-HRM approach demonstrated efficacy in detecting beer-spoilage microorganisms in multiplex reactions. Nonetheless, sensitivity variations among primers underscore the importance of thoughtful design for multiplex reactions with primers within the same sensitivity range. Our pipeline is highly adaptable and can be applied not only to the detection of various beer-spoilage microorganisms but also to other segments within the food industry, pharmaceutical, oil & gas industry, among others, effectively enhancing cost-efficient quality control measures. Keywords: Beer-spoilage. Quality control. Multiplex qPCR. HRM. Orthologous genes.
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    Tales from a genome heavily affected by RIP: unraveling Tc1/mariner and MITE transposable elements in Colletotrichum lindemuthianum
    (Universidade Federal de Viçosa, 2023-07-21) Ferst, Lara Mattana; Queiroz, Marisa Vieira de; http://lattes.cnpq.br/4408827815801006
    The Colletotrichum genus comprises fungal pathogens that inflict severe diseases on a wide range of hosts, including economically important plants. Colletotrichum lindemuthianum is the causative agent of anthracnose in common beans (Phaseolus vulgaris), leading to significant production and quality losses in this essential legume crop. Transposable elements are mobile genetic units found across all life domains that play a significant role in genomic plasticity, which is particularly relevant for phytopathogenic fungi such as C. lindemuthianum. These elements are classified into two distinct classes based on their transposition mode: Class I transposons rely on an RNA intermediate to transpose, while Class II transposons transpose directly as DNA. Additionally, transposons can be divided as autonomous and non- autonomous elements, with the latter depending on other transposable elements for their mobility due to the absence of essential functional components. The Tc1/mariner is a widely distributed Class II transposon superfamily. Miniature- inverted repeat elements (MITEs) are non-autonomous elements derived from Class II elements like the Tc1/mariner family. Considering the crucial role of transposons in generating genetic variability, this study aimed to identify and characterize Tc1/mariner and MITE transposable elements within the genome of C. lindemuthianum, using an in-silico approach. A total of 615 sequences related to Tc1/mariner elements were identified, which represented 0.78% of the genome. Among them, 536 copies were considered degenerated due to nearly unrecognizable terminal inverted repeats. Only one family of complete elements was found, and a derived MITE family was identified. Both the parental and derived families displayed a strong tendency to be found inserted within gene promoter regions. Moreover, two other MITE families were characterized, but the elements that originated them could not be identified. The elements from both families were primarily located within transposon-enriched regions. All complete elements showed putative transposase ORFs interrupted with multiple stop codons, suggesting that there are no active Tc1/mariner elements in C. lindemuthianum. Our analysis revealed that these elements were heavily affected by repeat-induced point mutations (RIP). Furthermore, we identified copies of both methyltransferases involved in RIP, which further supports that this mechanism or a RIP-like mechanism is active in C. lindemuthianum. The insights gained from this study contribute to a better understanding of the Tc1/mariner and MITE landscape as well as defense mechanisms against transposon proliferation of C. lindemuthianum, providing a foundation for further investigations into the genetic variability and transposable element exaptation of this economically important plant pathogen. Keywords: Class II transposons. Non-autonomous transposons. Repetitive DNA. Genome defense.
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    Isolation and characterization of yeasts from brazilian coffee beans for brewing application
    (Universidade Federal de Viçosa, 2023-11-11) Siqueira, Tatiane de Paula; Tótola, Marcos Rogério; http://lattes.cnpq.br/1740176026695532
    The use of new non-Saccharomyces yeasts is a strategy for obtaining beers with new sensory profiles. This makes it possible to attend to the expectations of consumers who are increasingly looking for innovative beverages. In this study, we proposed the isolation of yeasts from coffee fruits, samples that have a wide range of microbial diversity. The study was conducted on samples obtained from the Alto da Mogiana region, a place in Minas Gerais known for its high-quality coffees. Twenty-seven isolates were obtained and of these, 52 % were able to use maltose as a carbon source and 87.5 % exhibited low hydrogen sulphide production. In addition, most of the isolates exhibited tolerance to factors such as alcohol content, low pH and temperature variation. Based on these results, two isolates (F702 and F605) were identified and selected for laboratory-scale fermentation characterization. Isolate F605 belongs to the Wickerhamomyces anomalus species, while F702 is a strain of Torulaspora delbrueckii. Laboratory fermentation trials have shown that these yeasts are unable to attenuate beer wort and, consequently, do not produce ethanol. In this context, the use of isolate F605 in co-fermentation with a conventional yeast was proposed. The F605 isolate was selected because of the better sensory profile observed throughout the experiments. The beer produced had an alcohol content of 5.6 %, indicating that the conventional yeast was able to grow and ferment in the presence of the isolate. An assay was also carried out to check consumer acceptance, which showed positive evaluations for criteria such as appearance and aroma. It can therefore be concluded that the coffee fruits evaluated were promising environments for isolating yeast with potential for application in the brewing industry. This reinforces the importance of sustainably exploiting Brazilian environments to obtain new national brewing strains, which can generate financial returns for the economy, as well as scientific and technological advances.
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    Isolation and characterization of lytic bacteriophages for the composition of a phage cocktail capable of reducing bacterial biofilms in oil-related environments
    (Universidade Federal de Viçosa, 2023-07-06) Silva, Jéssica Duarte da; Paula, Sérgio Oliveira de; http://lattes.cnpq.br/0374413607226322
    Phages are viruses that infect bacteria and since their discovery in the early 20th century, they have been used to control bacterial growth. This work was divided into three chapters. The first genomically and biologically characterizes the Enterobacter phage vB_EclM-UFV01, presenting general characteristics of its genus, the Karamviruses. Chapter 2, examines the capacity of a phage cocktail (composed by enterobacteria-isolated phages) to reduce the biofilm of sulfate-reducing bacteria. The formulation of the cocktail, named Petro01, was performed by selecting phages from the collection of the Laboratory of Molecular Immunovirology at UFV and ended up being composed of three phages of the Tequatrovirus genus (briefly characterized genomically in this chapter) and one of the Karamvirus genus (described in Chapter 1). The stability of Petro01 under conditions similar to those found in petroleum-related environments, its shelf life at high storage temperatures, the supernatant composition (LB enriched with conservative compounds), as well as the propagation capacity of the phages that make up the cocktail in bench and a 12 L bioreactor, was also evaluated. Petro01 showed significant biotechnological potential and positive capacity for the composition of a phage-based product. Chapter 3 moves away from the environmental application of bacteriophages and characterizes, biologically and genomically, the Proteus mirabilis phages BigMira UFV01 and MidiMira UFV02, providing an overview of the common features of their genus, the Acadeviruses. It also presents the genomic evaluation of 10 strains of P. mirabilis, isolated in clinical environments, focusing on the LPS locus and the antimicrobial resistance profile. All phages described in this study have good potential as antibacterial agents. Keywords: Bacteriophages. Industrial microbiology. Sulfate-reducing bacteria. Bioproducing.