Search

Colletotrichum truncatum, the most relevant fungal species associated with soybean anthracnose, is responsible for major losses in the crop. Chemical control via fungicide application is still the most effective strategy for the control of soybean foliar diseases. However, the increase in anthracnose incidence in some regions of Brazil indicates that current chemical control has not been effective against anthracnose. In this study, we assessed the fungicide sensitivity of C. truncatum lineages using isolates representing two important regions of soybean production in Brazil to the fungicides azoxystrobin, thiophanate-methyl, difenoconazole, and fludioxonil. We characterized the molecular resistance to quinone-outside inhibitors (QoI), methyl benzimidazole carbamates (MBC) and demethylation inhibitors (DMI) fungicide groups based on amino acid sequences of the cytochrome b (cytb), β-tubulin gene (β-tub), and P450 sterol 14a-demethylases (CYP51) genes. Multiple resistance of C. truncatum isolates to QoI and MBC was observed associated with mutation points in the β-tub (E198A and F200Y) and cytb (G143A). Alternatively, low EC50 values were found for fludioxonil and difenoconazole indicating high efficacy. Analysis of C. truncatum genomes revealed two potential DMI targets, CYP51A and CYP51B, and higher genetic variability in the CYP51A gene. A slight correlation between genetic differentiation of C. truncatum populations and fungicide sensibility was found (Student’s t-test C. truncatum in Brazil.Highlights-Multiple resistance of C. truncatum to azoxystrobin and thiophanate-methyl-C. truncatum isolates are sensitive to difenoconazole and fludioxonil-Presence of E198A and F200Y β-tubulin mutations and G143A cytochrome b mutation-Presence of CYP51A and CYP51B paralogues and higher genetic variability in the CYP51A

Currently, the common bean (Phaseolus vulgaris L.) productivity has been reduced by the fungus Fusarium oxysporum f. sp. phaseoli (Fop), the causative agent of Fusarium wilt. Considering the integrated management of diseases, the objectives of the present work were to verify the compatibility between chemical and biological fungicides for Fusarium oxysporum Schlecht. f. sp. phaseoli Kendrick & Snyder (Fop) control in common bean seeds. In laboratory, the effects of the treatments were evaluated by sanity, germination, seedling total length and seedling dry matter tests. In greenhouse conditions, the emergence speed rate, the percentage of emergence and the rate of pathogen transmission through the pathogen infestation in a substrate to plants were evaluated. Common bean seeds BRS Estilo were artificially inoculated with Fop isolate (IAC 11.299-1). In the seeds’ treatment, the chemical fungicides fludyoxonyl, flutriafol, methyl tiofanate, and biological products of Trichoderma sp. (isolates SF04, GF 422 and strain 1306), separately and mixed, were used. Treatments that promoted the best pathogen control in seeds were the combination of methyl tiophanate with biological products. Both flutriafol and GF 422 isolated and in mixed treatments affected the seeds’ physiological quality. The protective effect of the products was noted in the transmission test, whose Fop incidence was from 5 to 40% in the hypocotyl and from 5 to 30% in common bean roots. RESUMO: Atualmente, a produtividade do feijão comum (Phaseolus vulgaris L.) foi reduzida pelo fungo Fusarium oxysporum f. sp. phaseoli (Fop), o agente causador da murcha de Fusarium. Considerando o manejo integrado de doenças, os objetivos do presente trabalho foram verificar a compatibilidade entre fungicidas químicos e biológicos para Fusarium oxysporum Schlecht. f. sp. phaseoli Kendrick & Snyder (Fop) em sementes de feijão. Em laboratório, foram avaliados os efeitos dos tratamentos pelos testes de sanidade, germinação, comprimento total de plântulas e matéria seca de plântulas. Em uma estufa, foram avaliadas a velocidade de emergência, a porcentagem de emergência e a taxa de transmissão de patógenos através da infestação de patógenos no substrato às plantas. As sementes de feijão comum BRS Estilo foram inoculadas artificialmente com isolado Fop (IAC 11.299-1). No tratamento das sementes, foram utilizados os fungicidas químicos fludioxoxil, flutriafol, tiofanato metílico e produtos biológicos de Trichoderma sp. (isolados SF04, GF 422 e cepa 1306), separadamente e misturados. Os tratamentos que promoveram o melhor controle de patógenos nas sementes foram a combinação de tiofanato de metila com produtos biológicos. Tanto o flutriafol quanto o GF 422 isolados e em tratamentos com misturas afetaram a qualidade fisiológica das sementes. O efeito protetor dos produtos foi observado no teste de transmissão, cuja incidência de Fop foi de 5 a 40% no hipocótilo e de 5 a 30% nas raízes do feijão comum.

The genus Stagonosporopsis is classified within the Didymellaceae family and has around 40 associated species. Among them, several species are important plant pathogens responsible for significant losses in economically important crops worldwide. Stagonosporopsis vannaccii is a newly described species pathogenic to soybean. Here, we present the draft whole-genome sequence, gene prediction, and annotation of S. vannaccii isolate LFN0148 (also known as IMI 507030). To our knowledge, this is the first genome sequenced of this species and represents a new useful source for future research on fungal comparative genomics studies.

Soybean (Glycine max L.) is one of the most important crops worldwide as a source of protein-rich foods and animal feeds. Anthracnose, one of the major limiting factors to soybean production (Dias et al. 2016), is caused by species such as Colletotrichum truncatum, C. sojae, and C. plurivorum (Damm et al. 2009, 2019). In December 2016 and 2017, soybean plants of cultivars Monsoy 8768 and Pioneer y-70 with typical symptoms of anthracnose (necrotic and irregular brown lesions on stems, leaves, and pods) were collected in Mato Grosso, Brazil. Commercial fields sampled showed 10 to 15% incidence of anthracnose in 1 ha in each sampled area. In total, 10 different geographic locations were sampled. Colletotrichum strains were isolated and cultured on potato dextrose agar at 25°C with a 12-h light photoperiod from surface-disinfected (70% alcohol followed by 0.5% sodium hypochlorite) plant tissues. Among others, three single-spore isolates (LFN0048 from Sinop, LFN0074 and LFN0090 from Lucas do Rio Verde) showed different morphology; isolates LFN0048 and LFN0074 were selected for further characterization. Total DNA was extracted and partial glyceraldehyde 3-phosphate dehydrogenase (GAPDH), histone H3 (HIS3), and β-tubulin (TUB2) genes were amplified and sequenced. The sequences were deposited in GenBank (accession numbers MN604249 and MK163893 for HIS3, MN604248 and MK142674 for GAPDH, and MN604250 and MK142675 for TUB) and were compared with most similar reference sequences of Colletotrichum (Damm et al. 2019). Both isolates clustered with Colletotrichum musicola epitype (CBS 132885), showing 100 and 98.5% similarity in GAPDH, 99.5 and 98.9% in HIS3, and 99.2% in TUB2. On PDA, colonies showed dark-gray aerial mycelium with entire margins, reverse violaceous-black. Conidia and ascospore size and shape match those previously described by Damm et al. (2019): 12.12 to 15.86 × 4.93 to 6.95 µm and 15.5 to 19.34 × 5 to 7.84 µm, respectively (n = 100). Appressoria (n = 50) were single or in loose groups, violaceous-black with predominant obovoid, truncated, and cylindrical shapes, with smooth, undulate, or lobate margin, and 9.25 to 29.79 × 7.22 to 21.06 µm. Perithecia, paraphyses; and unitunicate eight-spored asci were also observed. Asci were cylindrical to clavate, smooth-walled, and 48.12 to 68.78 × 9.59 to 14.47 µm (n = 50). Soybean anthracnose is seed-borne (Dias et al. 2018; Rogério et al. 2017); therefore, pathogenicity tests were carried out on pregerminated seeds. Five seeds of Brasmax 8579 cultivar were inoculated with 10 µl of a conidial suspension (106 conidia/ml) that was placed in the emerging radicle, and five mock-inoculated seeds were used as a control. Seedlings were planted in vermiculite and incubated at 25°C with a 12-h photoperiod. After 7 days, inoculated plants showed necrotic lesions on the cotyledons, leaflets, and hypocotyl, whereas control plants remained asymptomatic. The experiment was repeated three times. C. musicola was reisolated from the symptomatic tissues, and the identity was confirmed by morphology and multilocus phylogeny. Until now, C. musicola has been reported to be associated with Musa sp. (Damm et al. 2019) and Colocasia esculenta (Vásquez-López et al. 2019) in Mexico, and with Phaseolus lunatus in Brazil (Cavalcante et al. 2018). To our knowledge, this is the first report of C. musicola joining a group of new and emergent species of Colletotrichum causing anthracnose in soybean-producing regions around the world.

A cultura da soja tem grande importância na segurança alimentar mundial, sendo o Brasil o segundo maior produtor do grão. Dentre os fatores que afetam a produção da cultura, as doenças se destacam e uma das principais é a antracnose. Diferentes espécies de Colletotrichum têm sido relacionadas à antracnose da soja, como C. truncatum, geralmente reportado como o principal agente causal da doença, e o C. plurivorum, anteriormente nomeado C. cliviae, uma espécie recentemente descrita e associada ao patossistema. Desta forma, objetivou-se caracterizar de forma filogenética, morfológica e cultural a espécie C. plurivorum comparando-a com C. truncatum; comparar a agressividade e a capacidade de transmissão semente-planta e planta-semente entre C. plurivorum e C. truncatum na cultura da soja; testar a reação de diferentes cultivares de soja a C. plurivorum; e verificar a sensibilidade in vitro de C. plurivorum a fungicidas registrados no Brasil para controle da doença. Baseado na análise filogenética Bayesiana, os isolados anteriormente chamados de C. cliviae foram reclassificados como C. plurivorum, agrupando-se juntamente com novos isolados de coletas recentes. Um isolado foi classificado como C. musicola, primeiro relato desta espécie isolada de planta sintomática de soja. Os isolados de C. plurivorum apresentaram taxas de crescimento micelial maior do que os de C. truncatum, enquanto que a esporulação dos isolados da segunda espécie foi superior aos da primeira. No teste de comparação de agressividade, o isolado de C. truncatum se mostrou mais agressivo do que os isolados de C. plurivorum à cultivar AMS Tibagi, após inoculações com suspensões de esporos no estágio de crescimento V2-V3. No teste de transmissão semente-planta, as taxas de transmissão de C. plurivorum e de C. truncatum da semente para a planta variaram entre 37% e 79%. Não foi observada transmissão dos patógenos para os grãos em teste de sanidade quando as plantas foram inoculadas nos estágios R2 e R5, além de não afetarem a produção de vagens e grãos. No teste de reação de cultivares de soja a C. plurivorum, a AMS Tibagi foi a que apresentou maior resistência, enquanto que as demais cultivares avaliadas foram afetadas significativamente nos parâmetros de qualidade fisiológica avaliados. Nas avaliações de sensibilidade a fungicidas, isolados de C. plurivorum e C. musicola, apresentaram insensibilidade, baixa ou média sensibilidade aos fungicidas trifloxistrobina e piraclostrobina. Para esses mesmos fungicidas, os isolados de C. truncatum e C. sojae foram classificados como altamente sensíveis (CE50 menor que 1 µg mL-1). Os isolados de C. plurivorum e C. musicola apresentaram a mutação E198A no gene da beta-tubulina, que confere resistência ao tiofanato-metílico, enquanto que os isolados de C. truncatum foram considerados altamente sensíveis ao produto e não apresentaram a mutação no gene analisado. Isolados de C. plurivorum e de C. truncatum, avaliados no teste de inibição do crescimento micelial, foram considerados altamente sensíveis aos fungicidas difenoconazol e fludioxonil. Identificar e conhecer as características das espécies Colletotrichum que afetam uma das principais commodities do Brasil é fundamental para entender a epidemiologia da doença e obter sucesso no desenvolvimento de estratégias de controle mais eficientes. Soybean cultivation has great importance in world food security with Brazil being the second largest grain producer. Among the factors that affect crop production, diseases stand out and one of the main ones is anthracnose. Different species of Colletotrichum have been related to soybean anthracnose, such as C. truncatum, generally reported as the main causal agent of the disease, and C. plurivorum, previously named C. cliviae and a recently described pathosystem associated species. The aim of this work was to characterize in a phylogenetic, morphological and cultural way the species C. plurivorum comparing it with C. truncatum; to compare the aggressiveness and seed-plant and plant-seed transmission capacity between C. plurivorum and C. truncatum in soybean cultivation; to test the reaction of different soybean cultivars to C. plurivorum; and to verify C. plurivorum in vitro sensitivity to fungicides registered in Brazil for disease control. Based on Bayesian phylogenetic analysis, the strains previously called C. cliviae were reclassified as C. plurivorum, grouped together with new strains from recent collections. One strain was classified as C. musicola, the first report of this species on soybean symptomatic plant. The C. plurivorum strains had higher mycelial growth rates than those of C. truncatum, whereas the sporulation of the second species was superior. C. truncatum was more aggressive than C. plurivorum to the AMS Tibagi cultivar after inoculations with spore suspensions at the V2-V3 growth stage. In the seed-plant transmission test, C. plurivorum and C. truncatum transmission rates from the seed to the plant varied between 37% and 79%. Transmission of the pathogens to the grains was not observed when the plants were inoculated in stages R2 and R5, besides not affecting the production of pods and grains. Among the tested cultivars, \'AMS Tibagi\'showed the highest resistance to C. plurivorum, while the other cultivars were significantly affected in the physiological quality parameters. C. plurivorum and C. musicola strains showed insensitivity, low or medium sensitivity to trifloxystrobin and pyraclostrobin fungicides. For these same fungicides, C. truncatum and C. sojae strains were classified as highly sensitive (EC50 less than 1 µg mL-1). C. plurivorum and C. musicola strains showed the E198A mutation in the beta-tubulin gene, which confers resistance to thiophanate-methyl, whereas C. truncatum strains were considered highly sensitive to this fungicide and did not present the mutation in the beta-tubulin gene. C. plurivorum and C. truncatum strains were considered highly sensitive to difenoconazol and fludioxonil fungicides. Identifying and knowing the characteristics of the Colletotrichum species that affect one of the main commodities in Brazil is fundamental to understand the epidemiology of the disease and to succeed in the development of more efficient control strategies.