Morphological and molecular characterization of Curvularia and related species associated with leaf spot disease of rice in Peninsular Malaysia

Curvularia species are important phytopathogens reported worldwide. They are closely related; consist of major destructive pathogens mainly for grasses and cereal plants including rice (Oryza sativa). A leaf spot symptom of rice is one of the common symptoms found in the rice field and caused reduction of rice yield. However, there are no reports on Curvularia species associated with rice leaves showing spot symptoms. The objectives are to isolate and characterize Curvularia and related species from leaf spot of rice by using morphological and molecular characterization and to determine the phylogenetic relationship between the isolated fungi. Fungal isolation was done from diseased rice leaves showing leaf spot symptoms collected throughout Peninsular Malaysia. Thirty-three isolates were recovered and identified based on their morphological characteristics such as conidia morphology, colony appearance, pigmentation and growth rate for species delimitation. Internal transcribed spacer (ITS) region was amplified to confirm the species identification. The 33 isolates were identified as Bipolaris sorokiniana (10 isolates), Curvularia hawaiiensis (8 isolates), C. geniculata (6 isolates), C. eragrostidis (6 isolates), C. aeria (2 isolates) and C. lunata (1 isolate). A phylogenetic tree was constructed based on ITS sequences using neighbour-joining method. The tree grouped members of Curvularia and Bipolaris into different clades. The phylogenetic tree indicated that the presence of two groups of fungi species; highly virulent and mild pathogens. In conclusion, Curvularia species and Bipolaris sorokiniana were present in rice field in Malaysia and associated with leaf spot of rice

Mating type idiomorphs of Pyrenophora teres in Turkey

Pyrenophora teres f. maculata (Ptm) and Pyrenophora teres f. teres (Ptt) causes spot form and net form of net blotch diseases of barley, respectively. Although both forms of P. teres are morphologically similar, their symptoms and genetic background differ. In this study, 175 single spore (109 Ptm and 66 Ptt) isolates obtained from different regions of Turkey were evaluated for their mating type distribution and prevalence. Fungal isolates of both forms were verified using species-speci.c polymerase chain reaction (PCR) primers. For mating type determination studies, duplex PCR was performed using MAT-specific single nucleotide polymorphism primers. Sixty and 49 of 109 Ptm isolates were found as MAT1-1 and MAT1-2 types, respectively and 43 and 23 of 66 Ptt isolates were found as MAT1-1 and MAT1-2 types, respectively. These results show the possibility of sexual reproduction among the Ptm isolates in Turkey and Ptt population of Central Anatolia, Turkey. However, the overall pattern of Ptt isolates did not support the sexual reproduction hypothesis in Turkey. Sexual reproduction in the life cycle of P. teres is important since it could lead to genetic and pathogenic variation. As a result of new sexual combinations more virulent pathotypes of P. teres may occur

Development of Expressed Sequence Tag (EST)–based Markers for Genomic Analysis of a Barley 6H Region Harboring Multiple Net Form Net Blotch Resistance Genes

We previously identified a region near the centromere of barley ( L.) chromosome 6H that harbors two closely linked net form net blotch resistance genes ( and ). In this study, we mined barley expressed sequence tag (EST) databases and developed EST-derived markers for further mapping of this region in a population derived from ‘Rika’ and ‘Kombar’. Additionally, we did a comparative analysis of this region to other grass species. Fifteen newly developed markers were added to the previous map, and most were distributed within a 27.0-cM interval spanning and . The two resistance loci were delineated to a 1.6 cM genetic interval. Comparison of mapped ESTs along chromosome 6H to wheat bin-mapped ESTs suggests that and are most likely located on the long arm of chromosome 6H. Comparative analysis revealed that a 12.6-cM region containing the two resistance loci is colinear with a region of rice chromosome 2 between 23.7 and 29.9 Mb with only a few rearrangements, and nearly the same level of colinearity was observed between barley and in this region. This work refines the genetic and physical location of and and provides an initial step toward the map-based cloning of these two genes

A first genome assembly of the barley fungal pathogen Pyrenophora teres f. teres

Background: Pyrenophora teres f. teres is a necrotrophic fungal pathogen and the cause of one of barley’s most important diseases, net form of net blotch. Here we report the first genome assembly for this species based solely on short Solexa sequencing reads of isolate 0-1. The assembly was validated by comparison to BAC sequences, ESTs, orthologous genes and by PCR, and complemented by cytogenetic karyotyping and the first genome-wide genetic map for P. teres f. teres. Results: The total assembly was 41.95 Mbp and contains 11,799 gene models of 50 amino acids or more. Comparison against two sequenced BACs showed that complex regions with a high GC content assembled effectively. Electrophoretic karyotyping showed distinct chromosomal polymorphisms between isolates 0-1 and 15A, and cytological karyotyping confirmed the presence of at least nine chromosomes. The genetic map spans 2477.7 cM and is composed of 243 markers in 25 linkage groups, and incorporates SSR markers developed from the assembly. Among predicted genes, non-ribosomal peptide synthetases and efflux pumps in particular appear to have undergone a P. teres f. teres-specific expansion of non-orthologous gene families. Conclusions: This study demonstrates that paired-end Solexa sequencing can successfully capture coding regions of a filamentous fungal genome. The assembly contains a plethora of predicted genes that have been implicated in a necrotrophic lifestyle and pathogenicity and presents a significant resource for examining the bases for P. teres f. teres pathogenicity