Microbiologia

URI permanente desta comunidadehttps://locus.ufv.br/handle/123456789/11840

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The repertoire of effector candidates in Colletotrichum lindemuthianum reveals important information about Colletotrichum genus lifestyle
(Applied Microbiology and Biotechnology, 2019) Queiroz, Casley Borges de; Correia, Hilberty L. Nunes; Santana, Mateus Ferreira; Batista, Diego Silva; Vidigal, Pedro M. Pereira; Brommonschenkel, Sérgio Hermínio; Queiroz, Marisa Vieira de
The fungus Colletotrichum lindemuthianum is the causal agent of anthracnose in the common bean (Phaseolus vulgaris), and anthracnose is one of the most devastating diseases of this plant species. However, little is known about the proteins that are essential for the fungus-plant interactions. Knowledge of the fungus’ arsenal of effector proteins is of great importance for understanding this pathosystem. In this work, we analyzed for the first time the arsenal of Colletotrichum lindemuthianum effector candidates (ClECs) and compared them with effector proteins from other species of the genus Colletotrichum, providing a valuable resource for studying the infection mechanisms of these pathogens in their hosts. Isolates of two physiological races (83.501 and 89 A2 2-3) of C. lindemuthianum were used to predict 353 and 349 ClECs, respectively. Of these ClECs, 63% were found to be rich in cysteine, have repetitive sequences of amino acids, and/or possess nuclear localization sequences. Several conserved domains were found between the ClECs. We also applied the effector prediction to nine species in the genus Colletotrichum, and the results ranged from 247 predicted effectors in Colletotrichum graminicola to 446 in Colletotrichum orbiculare. Twelve conserved domains were predicted in the effector candidates of all analyzed species of Colletotrichum. An expression analysis of the eight genes encoding the effector candidates in C. lindemuthianum revealed their induction during the biotrophic phase of the fungus on the bean.
PacCl, a pH-responsive transcriptional regulator, is essential in the pathogenicity of Colletotrichum lindemuthianum, a causal agent of anthracnose in bean plants
(European Journal of Plant Pathology, 2014-08-08) Nogueira, Guilherme Bicalho; Soares, Marcos Antônio; Bazzolli, Denise Mara Soares; Araújo, Elza Fernandes de; Langin, Thierry; Queiroz, Marisa Vieira de
In fungi, the expression of genes encoding proteins related to parasitism is regulated by several factors, including pH. This study reports the structural and functional characterization of the pacCl gene, which encodes the transcription factor PacC of C. lindemuthianum. The pacCl gene showed reduced expression in acidic pH, and its transcription was activated by elevated extracellular pH. The importance of this gene was demonstrated by the development of a pacC1 disruption mutant line of C. lindemuthianum. The mutant line was able to penetrate the host tissue through differentiation of primary hyphae. However, it was not able to cause maceration on the infected plant tissue. The results suggest that PacCl is a regulator of gene activation, and its expression is required for fungal growth in alkaline conditions, as well as for the transcription of genes necessary for the passage from the biotrophic to the necrotrophic phase.
Differential expression of genes during the interaction between Colletotrichum lindemuthianum and Phaseolus vulgaris
(European Journal of Plant Pathology, 2016-08-26) Santana, Mateus F.; Cnossen, Andréia; Bazzolli, Denise M. S.; Araújo, Elza F. de; Queiroz, Marisa V. de; Bromonschenkel, Sérgio H.; Fontenelle, Mariana R.
The fungus Colletotrichum lindemuthianum is the causal agent of anthracnose, one of the most severe diseases of the common bean (Phaseolus vulgaris). The infection process begins with the adhesion of conidia to the plant’s surface. Appressoria are then formed, allowing penetration of the fungus. Next, the biotrophic phase begins, followed by the necrotrophic phase. Due to the peculiar nutrition mode of the fungus, including both of the previously mentioned stages, it is of great interest to determine which genes are involved in the transition between the two phases during the infection process. To determine this, suppression subtractive hybridization (SSH) was used in association with qRT-PCR in the present study. These methods allowed for the identification of genes that were differentially expressed at each developmental stage of the fungus in the plant. This is the first report on the use of the cited techniques to evaluate the infectious cycle of the fungus. A total of 175 sequences exhibited significant identity (e ≤ 10−5) with sequences present in the sequenced genomes of P. vulgaris and C. lindemuthianum; approximately 41 % of those were determined to belong to the fungus, and 59 % were determined to belong to the plant. Of the predicted sequences, 68 % were of unknown function or were not found in the databases. Among the analyzed expressed sequence tags (ESTs), sequences were found that encode proteins related to: primary and secondary metabolism; the transport of different compounds; the degradation/modification of proteins; cell regulation and signaling; cellular stress response; and the degradation of exogenous compounds. The obtained results allowed for the identification of sequences encoding proteins that are essential for the progression of anthracnose. Furthermore, it was possible to identify new genes, the functions of which have not yet been described, and even to identify unique genes of C. lindemuthianum that are involved in the pathogenicity and virulence of this fungus.
Development of new molecular markers for the Colletotrichum genus using RetroCl1 sequences
(World Journal of Microbiology and Biotechnology, 2011-10-01) Santos, Leandro Vieira dos; Queiroz, Marisa Vieira de; Santana, Mateus Ferreira; Soares, Marcos Antonio; Barros, Everaldo Goncalves de; Araujo, Elza Fernandes de; Langin, Thierry
A nonautonomous element of 624 bp, called RetroCl1 (Retroelement Colletotrichum lindemuthianum 1), was identified in the plant pathogenic fungus Colletotrichum lindemuthianum. RetroCl1 contains terminal direct repeats (223 bp) that are surrounded by CTAGT sequences. It has a short internal domain of 178 bp and shows characteristics of terminal-repeat retrotransposon in miniature (TRIM) family. We used RetroCl1 sequence to develop molecular markers for the Colletotrichum genus. IRAP (Inter-Retrotransposon Amplified Polymorphism) and REMAP (Retrotransposon-Microsatellite Amplified Polymorphism) markers were used to analyze the genetic diversity of C. lindemuthianum. Fifty-four isolates belonging to different races were used. A total of 45 loci were amplified. The Nei index showed significant differences among the populations divided according to race, indicating that they are structured according to pathotype. No clear correlation between IRAP and REMAP markers with pathogenic characterization was found. C. lindemuthianum has high genetic diversity, and the analysis of molecular variance showed that 51% of variability is found among the populations of different races. The markers were also tested in different Colletotrichum species. In every case, multiple bands were amplified, indicating that these markers can be successfully used in different species belonging to the Colletotrichum genus.
Beginning to understand the role of sugar carriers in Colletotrichum lindemuthianum: the function of the gene mfs1
(Journal of Microbiology, 2012-10-12) Pereira, Monalessa Fábia; Santos, Carolina Maria de Araújo dos; Araújo, Elza Fernandes de; Queiroz, Marisa Vieira de; Bazzolli, Denise Mara Soares
Fungi of the Colletotrichum genus are among the most prominent phytopathogens that cause diseases with a considerable economic impact, such as anthracnose. The hemibiotrophic fungus Colletotrichum lindemuthianum (teleomorph Glomerella cingulata f. sp. phaseoli) is the causal agent of the anthracnose of the common bean; and similarly to other phytopathogens, it uses multiple strategies to gain access to different carbon sources from its host. In this study, we examine mfs1, a newly identified C. lindemuthianum hexose transporter. The mfs1 gene is expressed only during the necrotrophic phase of the fungus’ interaction within the plant and allows it to utilize the available sugars during this phase. The deletion of mfs1 gene resulted in differential growth of the fungus in a medium that contained glucose, mannose or fructose as the only carbon source. This study is the first to describe a hexose transporter in the hemibiotrophic pathogen C. lindemuthianum and to demonstrate the central role of this protein in capturing carbon sources during the necrotrophic development of the plant/pathogen interaction.
Análise comparativa do genoma mitocondrial do fungo Colletotrichum lindemuthianum, agente causal da antracnose em feijão comum
(Applied Microbiology and Biotechnology, 2018-02-16) Queiroz, Casley Borges de; Santana, Mateus Ferreira; Vidigal, Pedro M. Pereira; Queiroz, Marisa Vieira de
Fungi of the genus Colletotrichum are economically important and are used as models in plant-pathogen interaction studies. In this study, the complete mitochondrial genomes of two Colletotrichum lindemuthianum isolates were sequenced and compared with the mitochondrial genomes of seven species of Colletotrichum. The mitochondrial genome of C. lindemuthianum is a typical circular molecule 37,446 bp (isolate 89 A2 2-3) and 37,440 bp (isolate 83.501) in length. The difference of six nucleotides between the two genomes is the result of a deletion in the ribosomal protein S3 (rps3) gene in the 83.501 isolate. In addition, substitution of adenine for guanine within the rps3 gene in the mitochondrial genome of the 83.501 isolate was observed. Compared to the previously sequenced C. lindemuthianum mitochondrial genome, an exon no annotated in the cytochrome c oxidase I (cox1) gene and a non-conserved open reading frame (ncORF) were observed. The size of the mitochondrial genomes of the seven species of Colletotrichum was highly variable, being attributed mainly to the ncORF, ranging from one to 10 and also from introns ranging from one to 11 and which encode a total of up to nine homing endonucleases. This paper reports for the first time by means of transcriptome that then ncORFs are transcribed in Colletotrichum spp. Phylogeny data revealed that core mitochondrial genes could be used as an alternative in phylogenetic relationship studies in Colletotrichum spp. This work contributes to the genetic and biological knowledge of Colletotrichum spp., which is of great economic and scientific importance.
Draft genome sequences of two isolates of colletotrichum lindemuthianum, the causal agent of anthracnose in common beans
(Genome Announcements, 2017-03-15) Queiroz, Casley Borges de; Correia, Hilberty L. Nunes; Menicucci, Renato Pedrozo; Vidigal, Pedro M. Pereira; Queiroz, Marisa Vieira de
Colletotrichum lindemuthianum is the causal agent of anthracnose in common beans, one of the main limiting factors of their culture. Here, we report for the first time, to our knowledge, a draft of the complete genome sequences of two isolates belonging to 83.501 and 89 A2 2-3 of C. lindemutuianum.
Structural and functional characterization of the Colletotrichum lindemuthianum nit1 gene, which encodes a nitrate eductase enzyme
(Genetics and Molecular Research, 2013-02-08) Nogueira, G.B.; Queiroz, M.V.; Ribeiro, R.A.; Araújo, E.F.
Colletotrichum lindemuthianum is the causal agent of plant bean anthracnose, one of the most important diseases affecting the common bean. We investigated the structure and expression of the nit1 gene (nitrate reductase) of C. lindemuthianum. The nit1 gene open reading frame contains 2787 bp, interrupted by a single 69-bp intron. The predicted protein has 905 amino acids; it shows high identity with the nitrate reductase of C. higginsianum (79%) and C. graminicola (73%). Expression of nit1 in C. lindemuthianum was evaluated in mycelia grown on different nitrogen sources under conditions of activation and repression. The gene was expressed after 15 min of induction with nitrate, reaching maximum expression at 360 min. The transcription was repressed in mycelia grown in media enriched with ammonia, urea or glutamine. Twenty nit1⁻ mutants were obtained in a medium treated with chlorate. Ten of these mutants were characterized by DNA hybridization, which identified point mutations, a deletion and an insertion. These rearrangements in the nit1 gene in the different mutants may have occurred through activity of transposable elements.

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