Efrén Remesal

Mycelial compatibility groups and pathogenic diversity in Sclerotium rolfsii populations from sugar beet crops in Mediterranean-type climate regions

The population structure of Sclerotium rolfsii from autumn‐sown sugar beet crops in Mediterranean‐type climate regions of Chile, Italy, Portugal and Spain was determined by analyses of mycelial compatibility groups (MCGs) and pathogenicity to 11 economically important plant species. Twelve MCGs (i–xii) were identified among 459 S. rolfsii isolates. MCG iii was the most prevalent group in all countries except Italy. MCG i, the most abundant group (64·7% of isolates) was identified in Portugal and Spain. The remaining MCGs were restricted to various regions within one country (ii, vi, ix) or different countries (v), or to specific localities (iv, vii, viii, x, xi, xii). MCGs iv, vii and x each comprised one isolate. Fields extensively sampled in southern Spain were infected with one to three MCGs. Plant species differed in susceptibility to MCG tester isolates with a MCG by species interaction. Cluster analyses allowed selection into five MCG groupings and grouped plant species into species‐groups 1 (broccoli, chickpea, sunflower, tomato) and 2 (cotton, pepper, sugar beet, watermelon). MCG groupings 1 (i, ix), 2 (ii, iii, vi, viii) and 5 (x, xii) were moderately virulent to species‐group 1 and mildly virulent to species‐group 2. MCG groupings 3 (iv, v, xi) and 4 (vii) were mildly virulent to both species‐groups. Across MCG groups, species were rated highly susceptible (chickpea, sunflower), susceptible (cotton, pepper, tomato, watermelon), moderately resistant (broccoli, melon, sugar beet) and resistant (corn, wheat). Establishing the MCG population structure and virulence variability among S. rolfsii isolates should help in the management of sclerotium root rot diseases.

Sequence variation in two protein-coding genes correlates with mycelial compatibility groupings in Sclerotium rolfsii.

Populations of Sclerotium rolfsii, the causal organism of Sclerotium root-rot on a wide range of hosts, can be placed into mycelial compatibility groups (MCGs). In this study, we evaluated three different molecular approaches to unequivocally identify each of 12 previously identified MCGs. These included restriction fragment length polymorphism (RFLP) patterns of the internal transcribed spacer (ITS) region of nuclear ribosomal DNA (rDNA) and sequence analysis of two protein-coding genes: translation elongation factor 1α (EF1α) and RNA polymerase II subunit two (RPB2). A collection of 238 single-sclerotial isolates representing 12 MCGs of S. rolfsii were obtained from diseased sugar beet plants from Chile, Italy, Portugal, and Spain. ITS-RFLP analysis using four restriction enzymes (AluI, HpaII, RsaI, and MboI) displayed a low degree of variability among MCGs. Only three different restriction profiles were identified among S. rolfsii isolates, with no correlation to MCG or to geographic origin. Based on nucleotide polymorphisms, the RPB2 gene was more variable among MCGs compared with the EF1α gene. Thus, 10 of 12 MCGs could be characterized utilizing the RPB2 region only, while the EF1α region resolved 7 MCGs. However, the analysis of combined partial sequences of EF1α and RPB2 genes allowed discrimination among each of the 12 MCGs. All isolates belonging to the same MCG showed identical nucleotide sequences that differed by at least in one nucleotide from a different MCG. The consistency of our results to identify the MCG of a given S. rolfsii isolate using the combined sequences of EF1α and RPB2 genes was confirmed using blind trials. Our study demonstrates that sequence variation in the protein-coding genes EF1α and RPB2 may be exploited as a diagnostic tool for MCG typing in S. rolfsii as well as to identify previously undescribed MCGs.

First report of southern blight of pepper caused by Sclerotium rolfsii in southern Spain

In May 2009, a stem rot of pepper (Capsicum annuum L.) occurred in a 20-ha field in Hacienda de Tarazona, Seville, in southern Spain. Affected plants appeared singly or were grouped in circular patches as much as 8 to 10 m in diameter. Early symptoms consisted of water-soaked lesions on crown and lower stem tissue in contact with the soil. Plant foliage became pale green and wilted, followed by a complete collapse of the plant. A dense white mycelial mat formed on the lower stem and crown with 1- to 2-mm-diameter, spherical, tan-to-dark brown sclerotia. Lower stem pieces of 12 plants with early disease symptoms were surface sterilized in 0.5% NaOCl, dried, transferred to acidified potato dextrose agar, and incubated at 25 ± 1°C in the dark. Fast-growing fungal colonies with white mycelium and abundant sclerotia developed after 6 to 10 days of incubation. On the basis of morphological characters, the fungus was identified as Sclerotium rolfsii Sacc. (2). To confirm the identity of the pathogen, the ribosomal DNA internal transcribed spacer was amplified and sequenced for two isolates (one of the two exact sequences was deposited as GenBank Accession No. GU080230). The sequence was 99% similar to sequences of Athelia rolfsii (S. rolfsii) in GenBank. Pathogenicity of two isolates was determined by placing two oat seeds colonized by each isolate 0.5 to 1 cm from the stem of 2-week-old pepper plants cv. Cristal (one plant per pot, eight replicates). Plants were incubated in a growth chamber maintained at 28 ± 1°C with a 14-h photoperiod of 360 μE·m-2·s-1 and 60 to 90% relative humidity for 10 days. By the sixth day, discoloration and blight of the foliage and stem was observed. Sclerotia formed around the crown and 88% of the plants died 7 days after inoculation. S. rolfsii was recovered from all affected pepper plants. Noninoculated control plants did not develop symptoms. In southern Spain, S. rolfsii is widely distributed in areas of sugar beet production (1). Because of the wide host range of the pathogen, southern blight could become an important disease of vegetable production in southern Spain. References: (1) R. Jordán-Ramírez et al. IOBC/WPRS Bull. 42:101. 2009. (2) J. E. M. Mordue. CMI Descriptions of Pathogenic Fungi and Bacteria. No. 410, 1974.