RESUMO
Fusarium graminearum of the 15-acetyl-deoxynivalenol (15-ADON) chemotype is the main cause of Fusarium head blight (FHB) of wheat in southern Brazil. However, 3-ADON and nivalenol (NIV) chemotypes have been found in other members of the species complex causing FHB in wheat. To improve our understanding of the pathogen biology and ecology, we assessed a range of fitness-related traits in a sample of 30 strains representatives of 15-ADON (F. graminearum), 3-ADON (F. cortaderiae and F. austroamericanum), and NIV (F. meridionale and F. cortaderiae). These included perithecia formation on three cereal-based substrates, mycelial growth at two suboptimal temperatures, sporulation and germination, pathogenicity toward a susceptible and a moderately resistant cultivar, and sensitivity to tebuconazole. The most important trait favoring F. graminearum was a two times higher sexual fertility (>40% perithecial production index [PPI]) than the other species (<30% PPI); PPI varied among substrates (maize > rice > wheat). In addition, sensitivity to tebuconazole appeared lower in F. graminearum, which had the only strain with effective fungicide concentration to reduce 50% of mycelial growth >1 ppm. In the pathogenicity assays, the deoxynivalenol producers were generally more aggressive (1.5 to 2× higher final severity) toward the two cultivars, with 3-ADON or 15-ADON leading to higher area under the severity curve than the NIV strains in the susceptible and moderately resistant cultivars, respectively. There was significant variation among strains of the same species with regards asexual fertility (mycelial growth, macroconidia production, and germination), which suggested a strain- rather than a species-specific difference. These results contribute new knowledge to improve our understanding of the pathogen-related traits that may explain the dominance of certain members of the species complex in specific wheat agroecosystems.
Assuntos
Fusarium/metabolismo , Fusarium/patogenicidade , Doenças das Plantas/microbiologia , Tricotecenos/biossíntese , Triticum/microbiologia , Farmacorresistência Fúngica/efeitos dos fármacos , Farmacorresistência Fúngica/genética , Fungicidas Industriais/farmacologia , Fusarium/genética , Genótipo , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Micélio/metabolismo , Micélio/patogenicidade , Especificidade da Espécie , Esporos Fúngicos/genética , Esporos Fúngicos/patogenicidade , Triazóis/farmacologia , Virulência/genéticaRESUMO
An efficient synthesis of a series of 4'-oxyalkyl-isocordoin analogues (2-8) is reported for the first time. Their structures were confirmed by ¹H-NMR, 13C-NMR, and HRMS. Their anti-oomycete activity was evaluated by mycelium and spores inhibition assay against two selected pathogenic oomycetes strains: Saprolegnia parasitica and Saprolegnia australis. The entire series of isocordoin derivatives (except compound 7) showed high inhibitory activity against these oomycete strains. Among them, compound 2 exhibited strong activity, with minimum inhibitory concentration (MIC) and minimum oomyceticidal concentration (MOC) values of 50 µg/mL and 75 µg/mL, respectively. The results showed that 4'-oxyalkylated analogues of isocordoin could be potential anti-oomycete agents.
Assuntos
Catecóis/química , Micélio/efeitos dos fármacos , Saprolegnia/efeitos dos fármacos , Esporos Fúngicos/efeitos dos fármacos , Antifúngicos/síntese química , Antifúngicos/classificação , Antifúngicos/farmacologia , Catecóis/síntese química , Catecóis/farmacologia , Compostos Inorgânicos/síntese química , Compostos Inorgânicos/química , Compostos Inorgânicos/farmacologia , Testes de Sensibilidade Microbiana , Micélio/patogenicidade , Saprolegnia/patogenicidade , Esporos Fúngicos/patogenicidadeRESUMO
The in vitro sensitivity of AvrPik allele isolates of Magnaporthe oryzae to isoprothiolane was examined and the virulence fitness costs of AvrPik allele isolates to isoprothiolane were assessed. Isoprothiolane was found to suppress the radial growth of AvrPik allele isolates at all concentrations (1, 5, 10, 15, and 20 µg/mL). Generally, a higher isoprothiolane concentration has a stronger inhibitory effect on mycelial growth in AvrPik allele isolates at 6 and 10 days after inoculation. The inhibitory effect of isoprothiolane also increased with treatment time. To determine whether a correlation existed between the in vitro sensitivity of AvrPik allele isolates and virulence, the half-maximal inhibitor concentration and 75% of the maximum inhibitor concentration were calculated for each mutation isolate and wild-type isolate. Based on these values and virulence, no significant correlation between the susceptibility of AvrPik allele isolates and virulence was detected. In summary, no fitness costs were associated with sensitivity of blast isolates carrying specific AvrPik alleles to different virulence.
Assuntos
Aptidão Genética/efeitos dos fármacos , Magnaporthe/efeitos dos fármacos , Mutação , Micélio/efeitos dos fármacos , Tiofenos/farmacologia , Alelos , Relação Dose-Resposta a Droga , Genótipo , Magnaporthe/genética , Magnaporthe/patogenicidade , Micélio/genética , Micélio/patogenicidade , Oryza/microbiologia , Doenças das Plantas/microbiologia , Fatores de Tempo , VirulênciaRESUMO
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.
Assuntos
Melaninas/metabolismo , Musa/microbiologia , Micélio/patogenicidade , Naftóis/metabolismo , Folhas de Planta/microbiologia , Saccharomycetales/patogenicidade , Oxigênio Singlete/metabolismo , Fatores de Virulência/metabolismo , Peróxido de Hidrogênio/metabolismo , Luz , Musa/efeitos da radiação , Micélio/metabolismo , Doenças das Plantas/microbiologia , Folhas de Planta/efeitos da radiação , Saccharomycetales/metabolismo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Espectrometria de Massas em TandemRESUMO
Corynespora Leaf Fall (CLF) is a major disease of rubber tree (Hevea brasiliensis) caused by the Ascomycota Corynespora cassiicola. Here we describe the cloning and characterization of a gene encoding cassiicolin (Cas), a glycosylated cystein-rich small secreted protein (SSP) identified as a potential CLF disease effector in rubber tree. Three isolates with contrasted levels of aggressiveness were analyzed comparatively. The cassiicolin gene was detected - and the toxin successfully purified - from the isolates with high and medium aggressiveness (CCP and CCAM3 respectively) but not from the isolate with the lowest aggressiveness (CCAM1), suggesting the existence of a different disease effector in the later. CCP and CCAM3 carried strictly identical cassiicolin genes and produced toxins of identical mass, as evidence by mass spectrometry analysis, thus suggesting conserved post-translational modifications in addition to sequence identity. The differences in aggressiveness between CCP and CCAM3 may be attributed to differences in cassiicolin transcript levels rather than qualitative variations in cassiicolin structure. Cassiicolin may play an important role in the early phase of infection since a peak of cassiicolin transcripts occurred in 1 or 2 days after inoculation (before the occurrence of the first symptoms), in both the tolerant and the susceptible cultivars.
Assuntos
Ascomicetos/genética , Proteínas Fúngicas/isolamento & purificação , Regulação Fúngica da Expressão Gênica/genética , Hevea/microbiologia , Micotoxinas/isolamento & purificação , Doenças das Plantas/microbiologia , Sequência de Aminoácidos , Ascomicetos/isolamento & purificação , Ascomicetos/patogenicidade , Sequência de Bases , Clonagem Molecular , Biologia Computacional , DNA Complementar/genética , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Dados de Sequência Molecular , Micélio/genética , Micélio/isolamento & purificação , Micélio/patogenicidade , Micotoxinas/química , Micotoxinas/genética , Folhas de Planta/microbiologia , RNA Fúngico/genética , RNA Fúngico/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Análise de Sequência de DNA , VirulênciaRESUMO
The effectiveness of Metarhizium anisopliae IP 46 conidia mixed with soil was tested against Aedes aegypti eggs. Mycelium and new conidia developed first on eggs between 4.8 and 15 days respectively after incubation of fungus-treated soils at 3.3 × 10(3) up to 3.3 × 10(5) conidia/g soil at 25°C and relative humidities close to saturation. After 15-day incubation, 53.3% of the eggs exposed to soil with 3.3 × 10(5) conidia/g showed external development of mycelium and conidia. Fungus-inoculated soils (but not untreated controls) showed some mycelial growth and sporulation apart from the eggs. Some eggs on treated soils hatched; those larvae died and eventually showed fungal development on their bodies. The cumulative relative eclosion of larvae after submersion of treated eggs in water decreased from 52.2% at 3.3 × 10(3) conidia/g to 25.3% at 3.3 × 10(5) conidia/g. These findings clearly showed that A. aegypti eggs can be infected by M. anisopliae when deposited on fungus-contaminated soil. The effectiveness of M. anisopliae against gravid females, larvae, and also eggs of A. aegypti underscored the possible usefulness of this fungus as a mycoinsecticide, whether naturally occurring or artificially applied, in the breeding sites of this mosquito.
Assuntos
Aedes/microbiologia , Metarhizium/crescimento & desenvolvimento , Metarhizium/patogenicidade , Óvulo/microbiologia , Controle Biológico de Vetores/métodos , Animais , Feminino , Larva/microbiologia , Micélio/crescimento & desenvolvimento , Micélio/patogenicidade , Microbiologia do Solo , Esporos Fúngicos/patogenicidadeRESUMO
BACKGROUND: Sporotrichosis caused by the dimorphic fungus Sporothrix schenckii can presents in a variety of clinical forms. Routine diagnosis is made by mycology and serology studies. Few investigations have been focused on the evaluation of the molecular diagnosis. AIM: To determine the value of the nested PCR technique for the diagnosis of experimental sporotrichosis in organs of mice, and to compare the results with the established laboratory diagnostic procedures. METHODS: BALB/c mice were inoculated with growing concentrations of the 2 morphological phases of the fungus. The infected animals were sacrificed one month later and specimens from liver, spleen, lung and testicle were obtained to perform wet mount, culture and molecular diagnosis by the nested PCR technique. Blood samples were obtained for determination of specific antibodies against S. schenckii by the double immunodiffusion procedure. RESULTS: The pathogenicity observed with the different concentrations of the fungus inoculated and its isolation by culture, showed scarce differences in the study of specimens from organs infected with the 2 morphological phases of S. schenckii. Specimens from organs of mice inoculated with the mycelial phase when studied by wet mount and culture, showed a higher positivity (100 and 37.5%) than those from mice inoculated with the yeast phase (73 and 2%). However, diagnosis by the nested PCR molecular technique applied to the latter specimens showed a higher percentage of positivity (75%) and 43% of positive results coming from animals infected with the mycelial phase. Specific antibody detection was positive in 100% all groups of infected mice. CONCLUSIONS: In the study of experimental sporotrichosis in mice, the culture, as well as the antibody detection, was an effective diagnostic procedure, while the nested PCR and microscopic studies had a lower diagnostic value.
Assuntos
DNA Fúngico/análise , Reação em Cadeia da Polimerase/métodos , Sporothrix/isolamento & purificação , Esporotricose/diagnóstico , Animais , Anticorpos Antifúngicos/sangue , Biópsia , Fígado/microbiologia , Pulmão/microbiologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Micélio/patogenicidade , Micologia/métodos , Baço/microbiologia , Sporothrix/genética , Sporothrix/crescimento & desenvolvimento , Sporothrix/imunologia , Sporothrix/patogenicidade , Testículo/microbiologia , VirulênciaRESUMO
The pathogenicity of seven strains of Fusarium equiseti isolated from seabed soil was evaluated on different host plants showing pre and post emergence damage. Radial growth of 27 strains was measured on culture media previously adjusted to different osmotic potentials with either KCl or NaCl (-1.50 to - 144.54 bars) at 15º, 25º and 35º C. Significant differences and interactive effects were observed in the response of mycelia to osmotic potential and temperature.
Assuntos
Cloreto de Potássio/análise , Cloreto de Sódio/análise , Microbiologia Ambiental , Fusarium/isolamento & purificação , Fusarium/patogenicidade , Meios de Cultura/isolamento & purificação , Micélio/crescimento & desenvolvimento , Micélio/patogenicidade , Microbiologia do Solo , Solos Salitrosos/análise , Métodos , Pressão Osmótica , Plantas , MétodosRESUMO
This study aimed to determine the antifungal activity of leaf aqueous and hydroethanolic extracts of 10 plants from the Brazilian Cerrado on Colletotrichum gloeosporioides and Corynespora cassiicola. Antifungal activity was measured through the incorporation of each extract in a culture media or spore suspension, at 50 percent concentration relative to the volume, determining respectively the mycelial growth and the spore germination. Then, the percentages of mycelial growth inhibition and spore germination inhibition were obtained based on the comparison with the control. The extracts had a variable action on the phytopathogens, from mycelial growth stimulation for Aristolochia esperanzae and Byrsonima verbascifolia extracts to complete inhibition of mycelial growth and spore germination for Myracrodruon urundeuva and Lafoensia pacari extracts. M. urundeuva, L. pacari and Caryocar brasiliense leaf extracts had antifungal activity against Colletotrichum gloeosporioides and Corynespora cassiicola; the hydroethanolic extracts presented more antifungal activity than the aqueous extracts, and spore germination of both phytopathogens was more affected than their mycelial growth.
O objetivo deste trabalho foi determinar a atividade antifúngica de extratos aquosos e extratos hidroetanólicos de folhas de 10 plantas do Cerrado brasileiro sobre Colletotrichum gloeosporioides e Corynespora cassiicola. A determinação da atividade antifúngica foi realizada pela incorporação do extrato em meio de cultura ou na suspensão de esporos, na concentração de 50 por cento em relação ao volume, determinando-se, respectivamente, o crescimento micelial e a germinação de esporos. Em seguida, pela comparação com a testemunha, foram obtidas as percentagens de inibição do crescimento micelial e da germinação dos esporos. Foi constatado comportamento variável dos extratos sobre os fitopatógenos, desde o estímulo no crescimento micelial para os extratos de Aristolochia esperanzae e Byrsonima verbascifolia, até a inibição completa do crescimento micelial e dagerminação dos esporos para os extratos de Myracrodruon urundeuva e Lafoensia pacari. Extratos de folhas de L. pacari, de M. urundeuva e de Caryocar brasiliense apresentaram atividade antifúngica sobre Colletotrichum gloeosporioides e Corynespora cassiicola; os extratos hidroetanólicos proporcionaram mais atividade antifúngica que os extratos aquosos, e a germinação de esporos de ambos os fitopatógenos foi mais afetada que o crescimento micelial.
Assuntos
Antifúngicos/análise , Colletotrichum , Extratos Vegetais/análise , Esporos Fúngicos/crescimento & desenvolvimento , Esporos Fúngicos/patogenicidade , Micélio/crescimento & desenvolvimento , Micélio/patogenicidadeRESUMO
Catalases and peroxidases are the most important enzymes that degrade hydrogen peroxide into water and oxygen. These enzymes and superoxide dismutase are the first lines of cell defense against reactive oxygen species. Metarhizium anisopliae displays an increase in catalase-peroxidase activity during germination and growth. To determine the importance of catalase during the invasion process of M. anisopliae, we isolated the cat1 gene. cat1 cDNA expression in Escherichia coli and the subsequent purification of the protein confirmed that the cat1 gene codes for a monofunctional catalase. Expression analysis of this gene by RT-PCR from RNA isolated from fungus grown in liquid cultures showed a decrease in the expression level of the cat1 gene during germination and an increase during mycelium growth. The expression of this gene in the fungus during the infection process of the larvae of Plutella xylostella also showed a significant increase during invasive growth. Transgenic strains overexpressing the cat1 gene had twice the catalase activity of the wild-type strain. This increase in catalase activity was accompanied by a higher level of resistance to exogenous hydrogen peroxide and a reduction in the germination time. This improvement was also observed during the infection of P. xylostella larvae. M. anisopliae transgenic strains overexpressing the cat1 gene grew and spread faster in the soft tissue of the insect, reducing the time to death of the insect by 25% and the dose required to kill 50% of the population 14-fold.