Genetic diversity of Rhizoctonia solani AG-3 from potato and tobacco in North Carolina

Anastomosis group 3 (AG-3) of Rhizoctonia solani (teleomorph = Thanatephorus cucumeris) is frequently associated with diseases of potato (AG-3 PT) and tobacco (AG-3 TB). Although isolates of R. solani AG-3 from these two Solanaceous hosts are somatically related based on anastomosis reaction and taxonomically related based on fatty acid, isozyme and DNA characters, considerable differences are evident in their biology, ecology, and epidemiology. However, genetic diversity among field populations of R. solani AG-3 PT and TB has not been documented. In this study, the genetic diversity of field populations of R. solani AG-3 PT and AG-3 TB in North Carolina was examined using somatic compatibility and amplified fragment length polymorphism (AFLP) criteria. A sample of 32 isolates from potato and 36 isolates from tobacco were paired in all possible combinations on PDA plus activated charcoal and examined for their resulting somatic interactions. Twenty-eight and eight distinct somatic compatibility groups (SCG) were identified in the AG-3 PT and AG-3 TB samples, respectively. AFLP analyses indicated that each of the 32 AG-3 PT isolates had a distinct AFLP phenotype, whereas 28 AFLP phenotypes were found among the 36 isolates of AG-3 TB. None of the AG-3 PT isolates were somatically compatible or shared a common AFLP phenotype with any AG-3 TB isolate. Clones (i.e., cases where two or more isolates were somatically compatible and shared the same AFLP phenotype) were identified only in the AG-3 TB population. Four clones from tobacco represented 22% of the total population. All eight SCG from tobacco were associated with more than one AFLP phenotype. Compatible somatic interactions between AG-3 PT isolates occurred only between certain isolates from the same field (two isolates in each of four different fields), and when this occurred AFLP phenotypes were similar but not identical.     

Genetic structure of populations of Rhizoctonia solani AG-3 on potato in eastern North Carolina

A polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method was developed to identify and differentiate genotypes of Rhizoctonia solani anastomosis group 3 subgroup PT (AG-3 PT), a fungal pathogen of potato. Polymorphic co-dominant single-locus PCR-RFLP markers were identified after sequencing of clones from a genomic library and digestion with restriction enzymes. Multilocus genotypes were determined by a combination of PCR product and digestion with a specific restriction enzyme for each of seven loci. A sample of 104 isolates from one commercial field in each of five counties in eastern North Carolina was analyzed, and evidence for high levels of gene flow between populations was revealed. When data were clone-corrected and samples pooled into one single North Carolina population, random associations of alleles were found for all loci or pairs of loci, indicating random mating. However, when all genotypes were analyzed, the observed genotypic diversity deviated from panmixia and alleles within and between loci were not randomly associated. These findings support a model of population structure for R. solani AG-3 PT on potato that includes both recombination and clonality.     

Development and Characterization of a Genetic Linkage Map of Cryptococcus neoformans var. neoformans Using Amplified Fragment Length Polymorphisms and Other Markers

A segregating population of single basidiospore isolates from a sexual cross was used to generate the first moderately dense genetic linkage map of Cryptococcus neoformans var. neoformans (Serotype D). Polymorphic DNA markers were developed using amplified fragment length polymorphisms, random amplified polymorphic DNA, and gene-encoding sequences. These markers were used to analyze 100 meiotic progeny. All markers were tested for distorted segregation with a goodness of fit test. Of the total of 181 markers, 148 showed balanced (1:1) segregation ratios. Segregation distortion was observed for 33 markers. Based on all the markers, a linkage map was generated that consists of 14 major linkage groups with 127 markers, several small linkage groups, and 2 linkage groups that consist only of highly skewed markers. The genetic distance of the linkage map is 1356.3 cM. The estimated total haploid genome size for C. neoformans var. neoformans was calculated using Hulberts method and yielded a map size of 1917 cM. The number of major linkage groups correlates well with the proposed number of 13 chromosomes for C. neoformans var. neoformans. Several genes, including CAP64, CnLAC, and the mating-type locus, were mapped, and their associations were consistent with published data. To date, 6 linkage groups have been assigned to their corresponding chromosomes. This linkage map should provide a framework for the ongoing genome sequencing project and will be a useful tool for studying the genetics and pathogenicity of this important medical yeast.     

Improved resolution of major clades within Tuber and taxonomy of species within the Tuber gibbosum complex

Tuber gibbosum Harkn., described from northern California, originally was thought to be a single, variable species that fruited from autumn through winter to spring. It has become popular as a culinary truffle in northwestern USA, where it is commercially harvested. Morphological studies suggested it might be a complex that includes at least two species. We conducted morphological and phylogenetic studies of the complex to determine how many species it might contain and how they differed morphologically, geographically and seasonally. We also provide the first LSU phylogeny for the genus Tuber. Phylogenetic analyses resolve nine major clades in the genus with high bootstrap support and distinguish the Gibbosum clade from the Aestivum, Excavatum, Macrosporum, Magnatum, Melanosporum, Puberulum, Rufum and Spinoreticulatum clades. Further analyses of ITS and LSU regions revealed four distinct species in the Gibbosum complex. Although morphologically similar the four species differ in spore size and shape and in peridial anatomy. These species share the synapomorphy of having suprapellis hyphae with distinctive, irregular wall swellings at maturity; we have not seen this hyphal type in any other Tuber spp. worldwide. The three new species are named and described as T. bellisporum Bonito & Trappe, T. castellanoi Bonito & Trappe and T. oregonense Trappe, Bonito & Rawlinson.     

Phylogeography of the Solanaceae-infecting Basidiomycota fungus <Emphasis Type="Italic">Rhizoctonia solani</Emphasis> AG-3 based on sequence analysis of two nuclear DNA loci

Background  

The soil fungus Rhizoctonia solani anastomosis group 3 (AG-3) is an important pathogen of cultivated plants in the family Solanaceae. Isolates of R. solani AG-3 are taxonomically related based on the composition of cellular fatty acids, phylogenetic analysis of nuclear ribosomal DNA (rDNA) and beta-tubulin gene sequences, and somatic hyphal interactions. Despite the close genetic relationship among isolates of R. solani AG-3, field populations from potato and tobacco exhibit comparative differences in their disease biology, dispersal ecology, host specialization, genetic diversity and population structure. However, little information is available on how field populations of R. solani AG-3 on potato and tobacco are shaped by population genetic processes. In this study, two field populations of R. solani AG-3 from potato in North Carolina (NC) and the Northern USA; and two field populations from tobacco in NC and Southern Brazil were examined using sequence analysis of two cloned regions of nuclear DNA (pP42F and pP89).     

New species and distribution records for Clavulina (Cantharellales,Basidiomycota) from the Guiana Shield,with a key to the lowland neotropical taxa

Three new and one previously described species of Clavulina (Clavulinaceae, Cantharellales, Basidiomycota) are reported from the central Guiana Shield region from tropical rainforests dominated by ectomycorrhizal trees of the leguminous genus Dicymbe (Fabaceae subfam. Caesalpinioideae). We provide morphological, DNA sequence, habitat, and fruiting occurrence data for each species. The new species conform to a generic concept of Clavulina that includes coralloid, branched basidiomata with amphigenous hymenia, basidia with two or 2−4 incurved sterigmata and postpartal septa present or absent, and smooth, hyaline, guttulate basidiospores. Placements of the new species in Clavulina were corroborated with DNA sequence data from the internal transcribed spacer and large subunit of the nuclear ribosomal repeat, and their infrageneric relationships were examined with phylogenetic analyses based on DNA from the region coding for the second largest subunit of DNA-dependent RNA polymerase II (rpb2). To facilitate future studies of the genus in the neotropics, a key is provided for all Clavulina species described from the lowland neotropics.