Genetic Structure of the Population of Alternaria solani in Brazil

Understanding the genetic structure of the population of Alternaria solani (AS) is an important component of epidemiological studies of early blight, a severe disease that affects potato (Po) and tomato (To) worldwide. Up to 150 isolates obtained from both hosts were analysed with RAPD and AFLP markers to estimate the amount and distribution of genetic variability of AS in Brazil. Using RAPD, gene diversity (h = 0.20) and scaled indices of diversity of Shannon (H′ = 0.66) and Stoddart and Taylor’s (G = 0.31) for the Po population were higher than those of the To (h = 0.07, H′ = 0.34, G = 0.17). For AFLP, the statistics for the Po (h = 0.17, H′ = 0.86, G = 0.49) and To (h = 0.17, H′ = 0.85, G = 0.36) populations were similar. For each RAPD and AFLP locus, the allele frequency for the overall population ranged from 0.006 to 0.988, and 0.007 to 0.993, respectively. Genetic differentiation was high (G_ST = 0.41 and θ = 0.59) and moderately high (G_ST = 0.23 and θ = 0.37) when estimated with RAPD and AFLP, respectively. Based on cluster analyses, there was strong evidence of association of pathogen haplotypes with host species. The null hypothesis of random association of alleles was rejected in the analysis of both RAPD (I_A = 13.1, P < 0.001) and AFLP (I_A = 2.2, P < 0.001) markers. The average number of migrants was estimated to be around one and two individuals per generation, using RAPD and AFLP, respectively. There was no correlation between genetic distance and geographical origin of AS haplotypes for RAPD (r = ?0.07, P = 0.84) and AFLP (r = ?0.03, P = 0.70). The AS population is clonal with high genetic variability, and there is genetic differentiation between the populations that affect To and Po.     

Bipolaris euphorbiae as a Biological Control Agent for Wild Poinsettia (Euphorbia heterophylla): Host-Specificity and Variability in Pathogen and Host Populations

Bipolaris euphorbiae (anamorphic Ascomycota: Pleosporaceae) has been studied since the early 1980s as a potential biocontrol agent of wild poinsettia (Euphorbia heterophylla), a noxious invader of soybean fields in Brazil. A new isolate (KLN05) was arbitrarily selected after the aggressiveness of six isolates of the fungus obtained from different sites was evaluated on wild poinsettias grown from seeds obtained from six different geographic origins in Brazil: Viçosa-state of Minas Gerais; Itabuna-state of Bahia; Niterói-state of Rio de Janeiro; Nova Laranjeira, Londrina and Foz do Iguaçú-state of Paraná. Bipolaris euphorbiae grew between 10 and 30 °C, with the optimum for colony growth at 25 °C. The best temperature range for conidial germination was between 20 and 30 °C. Conidial germination was not inhibited by mixing conidia with imazethapyr (106 g of active ingredient [a.i.]/l), glyphosate (720 g of a.i./kg), fomesafen (250 g of a.i./l), carfentrazone (400 g of a.i./l), atrazine (500 g of a.i./l) and glyphosate+carfentrazone (720 g of a.i/kg+400 g of a.i/l). Bipolaris euphorbiae was inoculated on plants belonging to 32 species of 12 different plant families and was only capable of infecting E. heterophylla.     

Genetics of metalaxyl resistance in Phytophthora infestans.

Several sexual crosses involving isolates of Phytophthora infestans of diverse sensitivities to metalaxyl were studied. Metalaxyl sensitivity was determined by comparing the growth of an isolate on metalaxyl-amended agar medium (5 microg/ml) with growth on medium containing no metalaxyl. When both parents had the same phenotype for metalaxyl sensitivity (both resistant or both sensitive), all F1 progeny had the parental phenotype. In two crosses (75 and 76) each involving one sensitive and one resistant parent, however, the progeny segregated 1:1, suggesting that the common resistant parent (Bg8) was heterozygous for metalaxyl sensitivity. When an F2 progeny was constructed from resistant F1 isolates in cross 76, the progeny segregated 1:3 (sensitive:resistant), indicating that metalaxyl resistance in Bg8 is conferred by a single dominant gene. Variation in the progeny sensitivity appears to involve minor genes. A correlation study between metalaxyl resistance and fitness components did not reveal any association.     

Effects of vitreousness and particle size of maize grain on ruminal and intestinal in sacco degradation of dry matter,starch and nitrogen

We assessed the effects of vitreousness and particle size of maize grain on ruminal and intestinal in sacco degradation of dry matter, starch and nitrogen. Six maize grain (Zea mays) genotypes characterized by differing vitreousness (proportion of vitreous in total endosperm) were ground (3-mm screen; Gr, ground particles, mean particle size (MPS): 526 μm) and cracked with a roller mill using two gap width settings (CS, cracked small particles, MPS: 1360 μm; CL, cracked large particles, MPS: 2380 μm). The ruminal and intestinal in sacco degradation of dry matter, starch and nitrogen was measured on three dry Holstein cows, fitted with rumen, proximal duodenum and terminal ileum cannulas, fed maize silage ad libitum twice daily. The ruminal starch degradability and intestinal digestibility differed among genotypes (P<0.001) and decreased as particle size increased (P<0.001). For the same particle size, starch ruminal degradability decreased (P<0.05) and intestinal digestibility decreased (P<0.002) with vitreousness. Particle size and vitreousness of maize grain are efficient factors for manipulating the amount of starch escaping rumen degradation, but may be limiting for the amount of starch digested in the small intestine.     

Novel and highly diverse fungal endophytes in soybean revealed by the consortium of two different techniques

Fungal endophytes were isolated from the leaves of soybean cultivars in Brazil using two different isolation techniques — fragment plating and the innovative dilution-to-extinction culturing — to increase the species richness, frequency of isolates and diversity. A total of 241 morphospecies were obtained corresponding to 62 taxa that were identified by analysis of the internal transcribed spacer (ITS) of the ribosomal DNA (rDNA). The Phylum Ascomycota predominated, representing 99% and 95.2% of isolates in the Monsoy and Conquista cultivars, respectively, whereas the Phylum Basidiomycota represented 1% and 4.8% of isolates, respectively. The genera Ampelomyces, Annulohypoxylon, Guignardia, Leptospora, Magnaporthe, Ophiognomonia, Paraconiothyrium, Phaeosphaeriopsis, Rhodotorula, Sporobolomyces, and Xylaria for the first time were isolated from soybean; this suggests that soybean harbours novel and highly diverse fungi. The yeasts genera Rhodotorula and Sporobolomyces (subphylum Pucciniomycotina) represent the Phylum Basidiomycota. The species richness was greater when both isolation techniques were used. The diversity of fungal endophytes was similar in both cultivars when the same isolation technique was used except for Hill’s index, N₁. The use of ITS region sequences allowed the isolates to be grouped according to Order, Class and Phylum. Ampelomyces, Chaetomium, and Phoma glomerata are endophytic species that may play potential roles in the biological control of soybean pathogens. This study is one of the first to apply extinction-culturing to isolate fungal endophytes in plant leaves, thus contributing to the development and improvement of this technique for future studies.     

Infection Process of Cercospora coffeicola on Coffee Leaf

Brown eye spot, caused by Cercospora coffeicola, is an important disease of coffee. Both adaxial and abaxial leaf surfaces were inoculated with a conidial suspension of C. coffeicola. Samples were collected from 4 to 168 h after inoculation and then again at 35 days. Germinated conidia showed positive tropism to stomata where attempted penetrations occurred. Appressoria were not observed. After penetration, C. coffeicola colonized the lacunous parenchyma both inter and intracellularly. Sporulation occurred through or around the stomata. Results from this study provide new insights into the infection process of C. coffeicola on coffee leaf.     

Genetic diversity analysis of isolates of the fungal bean pathogen Pseudocercospora griseola from central and southern Brazil

Planting resistant varieties is the most effective control measure against the angular leaf spot of dry beans, a fungal disease caused by Pseudocercospora griseola. However, dry bean varieties with durable resistance are not easily obtained. Knowledge about the genetic variability of the pathogen population is key for the success of dry bean breeding programs aimed at developing resistant materials, but finding suitable operationally simple and genetically accurate markers is not an easy task. The aim of this study was to assess the suitability of the ISSR-PCR technique to quantify the genetic variability of P. griseola isolates. Total DNA of 27 P. griseola isolates from Goiás, Minas Gerais, Espírito Santo, and Paraná States was extracted and amplified using specific primers for ISSR. Using cluster analysis, 27 genotypes were identified. The ISSR-PCR technique was suitable for assessing intraspecific variability of P. griseola. The ISSR-PCR marker was found to be highly sensitive to genetic variation and can aid in elucidating the genetic structure of the population of this plant pathogen as a support tool for the dry bean breeding programs.     

Biological control by Clonostachys rosea as a key component in the integrated management of strawberry gray mold

Gray mold, caused by Botrytis cinerea, is an important strawberry disease. As gray mold control is difficult, there is a need to evaluate integrated methods to successfully manage the disease. The efficiency of integrating Clonostachys rosea sprays, fungicide sprays, and crop debris removal to manage gray mold was evaluated in field experiments conducted in 2006 and 2007. Leaf colonization by C. rosea (LAC), average number of B. cinerea conidiophores (ANC), gray mold incidence in both flowers (Iflower) and fruits (Ifruit), and yield were evaluated weekly. In both years, LAC was higher in the treatments with no fungicide. When compared to the check, ANC, Iflower and Ifruit were most reduced in treatments that included C. rosea sprays. Maximal reductions were achieved with the combination of C. rosea sprays, fungicide sprays and debris removal (96.62%, 86.54% and 65.33% reductions of ANC, Iflower and Ifruit, respectively). Otherwise, maximal yield (103.14% increase as compared to the check) was achieved with the combination of the three treatments. With just C. rosea sprays, ANC, Iflower and Ifruit were reduced by 92.01%, 68.48% and 65.33%, respectively, whereas yield was increased by 75.15%. Considering the individual effects, application of C. rosea was the most efficient treatment. Chemical control was effective only in plots without debris removal. Elimination of crop debris was the least effective method in reducing gray mold incidence in both flowers and fruits. The integrated control approach enhanced the efficacy of the individual methods of gray mold control and provided high strawberry yield. An important component of this integrated approach it the biological control with C. rosea.