First Report of Sclerotium rolfsii in Greater Plantain in Paraná, Brazil.

Greater plantain or common plantain (Plantago major L.) is an herbaceous plant native to most of Europe, northern and central Asia, and it has adapted well to tropical regions where it is used as a medicinal plant. Between November 2011 and April 2012, greater plantains cultivated in the Medicinal Plant Garden at the Umuarama Advanced Campus of the State University of Maringa (UEM-CAU) suddenly died off. A visual examination revealed the presence of white mycelium and sclerotia on the lower third of the plant. These sclerotia were collected and deinfested by immersion in 70% alcohol and 0.5% sodium hypochlorite for 45 s, and in sterilized water for 1 min. Next, the sclerotia were placed on 10 petri dishes containing potato dextrose agar (PDA) culture medium and incubated at 29°C. After 7 days, the culture medium was entirely coated with a cottony white mycelial growth and, 15 days after isolation, sclerotia began to form. Healthy seedlings were transplanted individually into pots containing autoclaved soil (120°C/2 h). After 10 days, eight seedlings were inoculated with 8-mm mycelia disks deposited on the base of the plant, and eight seedlings inoculated with fungus-free PDA disks (control). The plants were irrigated and the pots placed in with plastic bags and kept at an average temperature of 28°C. Three days after inoculation, we observed a cottony white mycelial growth and symptoms of rot on the plants. The plastic bags were then removed and the plants kept under the same temperature, relative humidity of 80% and 12 h of light. Seven days after inoculation, the plants treated with the fungus died, whereas the plants treated with PDA developed normally. The fungus was re-isolated from the symptomatic plants and slides evaluated under a light microscope, revealing that the mycelium was thick, septate, and hyaline. The sclerotia formed were spherical, initially white or light brown, becoming dark brown or black, with diameters ranging from 0.5 to 1.5 mm. The fungus was subjected to DNA analysis using ribosomal region oligonucleotides ITS4 (5'-TCCTCCGCTTATTGATAT-3') and ITS5 (5'-GGAAGTAAAAGTCGTAACAAGG-3') (1) to amplify the target region. The segment including the 5.8S gene and rDNa regions ITS1 and ITS2 was 630 bp long. DNA analysis revealed that it was 99% identical to Athelia rolfsii (Curzi) Tu and Kimbr (anamorph: S. rolfsii) (GenBank Accession No. HM222638.1). The isolate was deposited in the fungus collection at the UEM-CAU Phytopathology Laboratory under code F-Sr-01-UMU. Reference: (1) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications, M. A. Innis et al., eds. Academic Press, San Diego, CA, 1990.

First Report of Leaf Spot Caused by Cladosporium perangustum on Syagrus oleracea in Brazil.

Syagrus oleracea (Mart.) Becc. (bitter coconut), a palm tree species that is native to central Brazil, has been increasingly cultivated in this country for heart-of-palm production. Epidemics of a necrotic leaf spot of unknown etiology have been recorded on bitter coconut plants in transplant nurseries and plantation since 2008. The first symptoms appear as small, yellow, hydrotic flecks on young or mature leaves that evolve to necrotic brown streaks that run parallel to the leaf veins. Usually, yellow halos occur around the lesions and hydrosis is common during lesion expansion. Necrotic lesions can reach up to 40 mm in length and 10 mm in width, and the lesions often coalesce, causing extensive tissue damage. During a survey in a 3-year-old bitter coconut plantation in Maringá County (coordinates: 23°23'51.25″ S, 51°57'02.09″ W; elevation: 507 m) in the state of Parana, a dozen symptomatic leaves were collected with the aim of elucidating the etiology of this disease. Conidia and conidiophores typical of Cladosporium were frequently observed on the diseased leaf tissue under natural field conditions as well on the surfaces of disinfected leaf tissues kept in a humid chamber for 48 h at 25 ± 2°C with a 12-h photoperiod. Five monoconidial cultures growing on potato dextrose agar (PDA) medium were obtained from different leaves showing leaf spot symptoms. The cultures were grown on PDA to induce sporulation. At 7 days after incubation at 25 ± 2°C and a 12-h photoperiod, gray to gray-olive colonies were observed. The conidiophores were macronematous, erect, oblong, branched, 1 to 5 septate, and 75.0 to 120.0 × 1.90 to 3.20 µm. The ramoconidia were cylindrical or oblong, 0 to 2 septate, and 28.0 to 40.0 × 2.8 to 3.6 µm, with a truncate base of 1.9 to 2.2 µm; secondary ramoconidia were cylindrical or oblong, 0 to 2 septate, 8.0 to 31.0 × 2.2 to 3.1 µm, with 3 to 5 distal conidial hila; intercalary 1-septate conidia were 5.5 to 17.0 × 2.1 to 3.4 µm, with 1 to 3 distal conidial hila; terminal 1-septate conidia were catenulate and 2.2 to 4.2 × 1.8 to 3.1 µm. Species identification was performed based on morphology and DNA sequence data (1). Portions of the elongation factor 1α (551 bp; TEF) and actin (213 bp; ACT) genes were amplified by PCR. A BLAST search of the GenBank database revealed that the TEF (KC484658 to KC484662) and ACT (KC484663 to KC484667) sequence fragments from isolates Gua1, Gua2, Gua3, Gua4, and Gua5 had 100% identity with the accessions HM148616 and HM148371 of Cladosporium perangustum (1). Isolates were tested for pathogenicity against bitter coconut. Ten potted plants with 4 to 6 fully expanded leaves were inoculated with each isolate by spraying a suspension of conidia (105 spores per ml) onto leaves until runoff using a handheld spray bottle. Non-inoculated controls (10 plants) were sprayed with distilled water. The plants were kept in a humid plastic chamber at 20 to 26°C. All examined isolates were pathogenic to bitter coconut, causing symptoms identical to those described above 12 days after inoculation. All inoculated tissues were plated onto PDA to confirm the presence of the pathogen. Live cultures are being maintained at the Micoteca/URM/UFPE ( www.ufpe.br/micoteca ), Brazil. To our knowledge, this is the first report of a disease caused by C. perangustum on S. oleracea worldwide, and the study provides valuable plant disease diagnostic information for the palm hearth industry in Latin America. Reference: (1) K. Bensch et al. Stud Mycol. 67:1, 2010.

Fusarium species and fumonisins associated with maize kernels produced in Rio Grande do Sul State for the 2008/09 and 2009/10 growing seasons.

Ear rots caused by Fusarium spp. are among the main fungal diseases that contribute to poor quality and the contamination of maize grains with mycotoxins. This study aimed to determine the visual incidence of fungal-damaged kernels (FDKs), the incidence of two main Gibberella (a teleomorph of Fusarium) complexes (G. fujikuroi and G. zeae) associated with maize using a seed health blotter test, and the fumonisin levels, using high performance liquid chromatography, in samples of maize grains grown across 23 municipalities during the 2008/09 and 2009/10 growing seasons. Additionally, 104 strains that were representative of all of the analysed samples were identified to species using PCR assays. The mean FDK was seven per cent, and only six of the samples had levels greater than six per cent. Fusarium spp. of the G. fujikuroi complex were present in 96% of the samples, and G. zeae was present in 18% of the samples (5/27). The mean incidence of G. fujikuroi was 58%, and the incidence of G. zeae varied from 2 to 6%. FB1 was found in 58.6%, FB2 in 37.9%, and both toxins in 37.9% of the samples. The FB1 and FB2 levels were below the quantification limits for 41.3% of the samples, and the mean FB1 levels (0.66 µg/g) were higher than the mean FB2 levels (0.42 µg/g). The PCR identification separated the 104 isolates into three of the G. fujikuroi complex: F. verticillioides (76%), F. subglutinans (4%) and F. proliferatum (2%); and G. zeae (anamorph = F. graminearum) (18%). Our results confirmed the dominance of F. verticillioides, similar to other regions of Brazil, but they differed due to the relatively higher incidence of F. graminearum. Total fumonisin levels were below the maximum limit determined by current Brazilian regulations.

Fusarium species and fumonisins associated with maize kernels produced in Rio Grande do Sul State for the 2008/09 and 2009/10 growing seasons

Ear rots caused by Fusarium spp. are among the main fungal diseases that contribute to poor quality and the contamination of maize grains with mycotoxins. This study aimed to determine the visual incidence of fungal-damaged kernels (FDKs), the incidence of two main Gibberella (a teleomorph of Fusarium) complexes (G. fujikuroi and G. zeae) associated with maize using a seed health blotter test, and the fumonisin levels, using high performance liquid chromatography, in samples of maize grains grown across 23 municipalities during the 2008/09 and 2009/10 growing seasons. Additionally, 104 strains that were representative of all of the analysed samples were identified to species using PCR assays. The mean FDK was seven per cent, and only six of the samples had levels greater than six per cent. Fusarium spp. of the G. fujikuroi complex were present in 96% of the samples, and G. zeae was present in 18% of the samples (5/27). The mean incidence of G. fujikuroi was 58%, and the incidence of G. zeae varied from 2 to 6%. FB1 was found in 58.6%, FB2 in 37.9%, and both toxins in 37.9% of the samples. The FB1 and FB2 levels were below the quantification limits for 41.3% of the samples, and the mean FB1 levels (0.66 µg/g) were higher than the mean FB2 levels (0.42 µg/g). The PCR identification separated the 104 isolates into three of the G. fujikuroi complex: F. verticillioides (76%), F. subglutinans (4%) and F. proliferatum (2%); and G. zeae (anamorph = F. graminearum) (18%). Our results confirmed the dominance of F. verticillioides, similar to other regions of Brazil, but they differed due to the relatively higher incidence of F. graminearum. Total fumonisin levels were below the maximum limit determined by current Brazilian regulations.

Molecular survey of trichothecene genotypes of Fusarium graminearum species complex from barley in southern Brazil.

Fusarium head blight is a disease of primary concern to small-grain cereals of Brazil, including barley. Its main causal agent, Fusarium graminearum species complex (Fg complex)¸ is able to produce mycotoxins, especially deoxynivalenol (DON) and nivalenol (NIV), that usually contaminate grain. Strains that produce DON may also produce its acetylated derivatives: 3-acetyl-DON (3-ADON) and 15-acetyl-DON (15-ADON). Ninety two isolates were obtained from samplings of barley grain during three years (2007, 2008 and 2009) from several fields in both southern and northern production regions of Rio Grande do Sul state, Brazil. These isolates were examined for polymerase chain-reaction-based (PCR) trichothecene genotype based on the amplification of portions of Tri3 and Tri12. There was no effect of year or region on the proportion of trichothecene genotypes. Overall, 66% of the strains (61/92) were 15-ADON, 4.4% (4/92) were 3-ADON and 29.3% (27/92) were NIV. The overall NIV/DON ratio estimated (0.41) was five times higher than that found in previous studies with strains from wheat grown in the same region. Species identification of nine strains representing the trichothecene genotypes, based on comparisons of DNA sequences of portions of the PHO, RED and URA genes with sequences from curated reference isolates of Fusarium from GenBank, revealed that they belong to F. graminearum sensu stricto (four 15-ADON and one 3-ADON strain), F. meridionale (three NIV strains) and F. austroamericanum (one 3-ADON strain). These results add to the current regional knowledge of trichothecene genotypes and species within the Fg complex affecting barley in the region.

Diallel analysis of leaf disease resistance in inbred Brazilian popcorn cultivars.

We estimated general and specific combining abilities and examined resistance to northern leaf blight (Exserohilum turcicum) and to gray leaf spot (Cercospora zeae-maydis) in a set of nine inbred popcorn lines. These inbreds were crossed in a complete diallel scheme without reciprocals, which produced 36 F(1) hybrids. Two experiments with a square lattice design and three replications were conducted during the 2008/2009 crop season, in Maringá, PR, Brazil. The severity of northern leaf blight and gray leaf spot was assessed under natural infestation conditions. Data were examined by individual and joint analysis of variance. Individual and joint Griffing's diallel analyses were carried out for adjusted means. General combining ability and specific combining ability were significant (P < 0.10) by the F-test for northern leaf blight and gray leaf spot infestation levels. This denotes that additive and non-additive gene effects both contributed to resistance to these diseases, but that the additive gene effects were more important. Among the inbred lines, P(8) and P(9) gave the highest resistance to northern leaf blight, and P(3) and P(4.3) gave the highest resistance to gray leaf spot. The hybrids P(7.4) x P(8) and P(4.3) x P(9) could be exploited by reciprocal recurrent selection to provide genotypes with both northern leaf blight and gray leaf spot resistance. Significant interaction between general combining ability and crop season (P < 0.10) denotes the importance of environment, even though the disease levels in the hybrids were quite consistent.