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Fungicide injection is regularly performed to prevent and manage Dutch elm disease (DED) of American elm (Ulmus americana). In an effort to better understand the effects of long-term fungicide injection on tree health, sonic tomography (SoT) and electricalresistance tomography (ERT) were used to nondestructively determine the incidence and severity of internal decay in the lower trunk of American elms in suburban and urban settings. Overall, 253 sonic and electricalresistance tomograms were generated from 210 American elms. Sampled trees were partitioned into two fungicide injection groups: (1) regular injection; and (2) irregular injection or no known history of injection. Among all American elms, the incidence of internal decay in the lower trunk was 30% (63/210) with a mean percent decay, as determined by SoT, of 39%. Based on Chi-square analysis, there were no significant differences in the frequency of elms with decay by injection history (P = 0.799). Mean percent decay was significantly different by dbh class (P = 0.005) and while linear regression demonstrated a positive correlation between percent decay and dbh, most of the variability went unexplained (R2 = 0.182). For elms with decay, there was a significantly higher frequency of trees in the lowest decay class (< 25% of the cross section) compared to the highest decay class (> 75% of the cross section). The results suggest that the wounding associated with regular fungicide injection does not increase the likelihood of internal decay and that American elms exhibit a low frequency and severity of internal decay. [ABSTRACT FROM AUTHOR]

The population structure of Armillaria gallica, an important pathogen of Quercus spp., was investigated from mixed oak forests in central Massachusetts, encompassing a sampling area over 500 km². From 16 plots at four sites a total of 153 isolates (34-40 isolates per site) was analyzed with amplified fragment length polymorphisms (AFLPs). Analyses of 204 polymorphic loci detected 38 AFLP genotypes from a sample area of 4.51 hectares (ha). Genets ranged in distribution from five to 33 genets per hectare (GPH), with a mean of eight GPH and the average A. gallica genet occupying 0.13 ha. Allele frequencies produced an unbiased expected heterozygosity (HE) value of 0.112 (SE = 0.006) and a Nei's expected heterozygosity (Hj) value of 0.190 (SE = 0.009), indicating low genetic diversity within the population. Analysis of molecular variation (ΦPT = 0.301; P < 0.001) indicates high genetic differentiation, with 70% of the molecular variation explained at the site-level within A. gallica subpopulations. However, results of the Mantel test, used to assess the isolation-by-distance hypothesis, were inconclusive in determining whether the subpopulations were truly isolated by distance. A neighbor-joining tree constructed from a genetic distance matrix grouped genotypes from the same site (subpopulation) together, but from three of four sites genotypes were randomly clustered at the plot level. The results suggest that basidiospore dispersal is an important means of new genet formation at linear distances up to 2000 m. [ABSTRACT FROM AUTHOR]

Sudden vegetation dieback (SVD) is the loss of smooth cordgrass (Spartina alterniflora) along intertidal creeks in salt marshes of the Atlantic and Gulf states. The underlying cause of SVD remains unclear, but earlier work suggested a contributing role for Fusarium spp. in Louisiana. This report investigated whether these or other Fusarium species were associated with S. alterniflora dieback in mid- to north-Atlantic states. Isolations from seven SVD sites yielded 192 isolates of Fusarium spp., with more than 75% isolated from aboveground tissue. Most isolates (88%) fell into two undescribed morphospecies (MS) distinguished from each other by macroconidial shape, phialide ontogeny and growth rates. Pathogenicity tests on wound-inoculated S. alterniflora steins and seedling roots revealed that isolates in MS1 were more virulent than those in MS2 but no single isolate caused plant mortality. No matches to known species of Fusarium were revealed by DNA sequence queries of translation elongation factor 1-α (tef1) sequences. A phylogenetic analysis of partial sequences of three genes, β-tubulin (β-tub), calmodulin (cal) and tef1, was conducted on representative isolates from MSI (n = 20) and MS2 (n = 18); it provided strong evidence that the MS1 isolates form a clade that represents a heretofore undescribed species, which we designate Fusarium palustre sp. nov. Isolates from the more variable MS2 clustered with the F. incarnatum-equiseti species complex as F. cf. incarnatum. Although a strong association exists between both species and declining S. alterniflora in SVD sites, neither appears to play a primary causal role in SVD. However, our findings suggest that F. palustre might play an important secondary role in the ecological disruption of the salt marshes. [ABSTRACT FROM AUTHOR]

Neonectria ditissima is a fungal pathogen native to eastern North America that causes disfiguring cankers on numerous tree species, particularly birches ( Betula spp.). In order to develop control strategies, fundamental knowledge of the pathogen's reproductive and dispersal dynamics is necessary. To undertake these studies, we developed genomic libraries enriched for clones containing microsatellites, and then designed primers flanking each locus. Of 34 loci that were screened, 11 were polymorphic among 38 isolates obtained from a single population. Gene diversities ranged from 0.05 to 0.862 with a mean of 0.409. These markers will be invaluable in population studies of this fungus. [ABSTRACT FROM AUTHOR]

The study of quantitative traits provides a window on the interactions between multiple unlinked genetic loci. The interaction between hosts and pathogenic microbes, such as fungi, involves aspects of quantitative genetics for both partners in this dynamic equilibrium. One important pathogenic fungus is Cryptococcus neoformans, a basidiomycete yeast that can infect the human brain and whose mating system has two mating type alleles, a and α. The α mating-type allele has previously been linked to increased virulence potential. Here congenic C. neoformans strains were generated in the two well-characterized genetic backgrounds B3501α and NIH433a to examine the potential influence of genes outside of the mating-type locus on the virulence potential of mating type. The congenic nature of these new strain pairs was established by karyotyping, amplified fragment length polymorphism genotyping, and whole-genome molecular allele mapping (congenicity mapping). Virulence studies revealed that virulence was equivalent between the B3501 a and α congenic strains but the a strain was more virulent than its a counterpart in the NIH433 genetic background. These results demonstrate that genomic regions outside the mating type locus contribute to differences in virulence between a and α cells. The congenic strains described here provide a foundation upon which to elucidate at genetic and molecular levels how mating-type and other unlinked loci interact to enable microbial pathogenesis. [ABSTRACT FROM AUTHOR]

To construct a genetic linkage map of the heterothallic yeast, Cryptococcus neoformans (Filobasidiella neoformans), we crossed two mating-compatible strains and analyzed 94 progeny for the segregation of 301 polymorphic markers, consisting of 228 restriction site polymorphisms, 63 microsatellites, two indels, and eight mating-type (MAT)-associated markers. All but six markers showed no significant (P< 0.05) segregation distortion. At a minimum LOD score of 6.0 and a maximum recombination frequency of 0.30, 20 linkage groups were resolved, resulting in a map length of ∼1500 cM. Average marker density is 5.4 cM (range 1-28.7 cM). Hybridization of selected markers to blots of electrophoretic karyotypes unambiguously assigned all linkage groups to chromosomes and led us to conclude that the C. neoformans genome is ∼20.2 Mb, comprising 14 chromosomes ranging in size from 0.8 to 2.3 Mb, with a ratio of ∼13.2 kb/cM averaged across the genome. However, only 2 of 12 ungrouped markers hybridized to chromosome 10. The hybridizations revealed at least one possible reciprocal translocation involving chromosomes 8, 9, and 12. This map has been critical to genome sequence assembly and will be essential for future studies of quantitative trait inheritance. [ABSTRACT FROM AUTHOR]

Selfing in the chestnut blight fungus, Cryphonectria parasitica, occurs by two different genetic mechanisms. Most self-fertile isolates of C. parasitica are heterokaryotic for mating type, and the progeny from selfing segregate for mating type. Further, we resolved mating-type (MAT) heterokaryons into homokaryons of both mating types by isolating uninucleate asexual spores (conidia). However, because ascospore progeny, with rare exceptions, are not MAT heterokaryons, C. parasitica must lack a regular mechanism to maintain heterokaryosis by selfing. We hypothesize that heterokaryon formation may occur either because of recurrent biparental inbreeding, or by mating-type switching, possibly one involving some kind of parasexual process. The second mechanism found for selfing in C. parasitica occurred less frequently. Three single-conidial isolates (MAT-1 and MAT-2) selfed and produced progeny that did not segregate for mating type. It is currently not known if meiosis occurs during ascospore formation by this mechanism.Index descriptors: Cryphonectria parasitica, Heterokaryosis, MAT, Mating type, Self-fertility, Self-incompatibility, Self-sterility [Copyright &y& Elsevier]

Although the genetic components of mating systems in fungi are well understood as laboratory phenomena, surprisingly little is known about their function in nature or about their role in determining mating patterns and population genetic structure. Our study of the mating system of the haploid ascomycete fungus, Cryphonectria parasitica, resulted in the following. (1) Laboratory crosses among 20 isolates, chosen randomly from North America and China, resolved into two incompatibility groups (occurring on both continents), confirming that C. parasitica has a diallelic, bipolar sexual self-incompatibility system, typical of other self-incompatible Ascomycetes, in which mating is only successful between isolates of opposite mating type. (2) PCR-based markers for mating-type alleles correlated perfectly with mating-type phenotypes of individual isolates. (3) Three genotypes, isolated from natural populations in Virginia and West Virginia, were inoculated onto chestnut trees in two sites in West Virginia and were confirmed to have self-fertilized and outcrossed in both sites. (4) Ten isolates, of a total of over 200 assayed, were confirmed to have self-fertilized in the laboratory, albeit at very low frequency. Five of these 10 isolates were ramets of a single genet, suggesting a genetic basis underlying the proclivity to self-fertilize in the laboratory. (5) Self-fertilization could not be induced in the laboratory with exudates (ostensibly containing pheromones) from isolates of opposite mating type. These results demonstrate that, a sexual self-incompatibility system notwithstanding, self-fertilization occurs under both laboratory and field conditions in C. parasitica. The disparity between observations of frequent selfing in nature and rare selfing in the laboratory suggests that the mating system is under ecological as well as genetic control. [ABSTRACT FROM AUTHOR]

Boxwood blight, a fungal disease of ornamental plants (Buxus spp.), is caused by two sister species, Calonectria pseudonaviculata (Cps) and C. henricotiae (Che). Compared to Cps, Che is documented to display reduced sensitivity to fungicides, including the azole class of antifungals, which block synthesis of a key fungal membrane component, ergosterol. A previous study reported an ergosterol biosynthesis gene in Cps, CYP51A, to be a pseudogene, and RNA-Seq data confirm that a functional CYP51A is expressed only in Che. The lack of additional ergosterol biosynthesis genes showing significant differential expression suggests that the functional CYP51A in Che could contribute to reduced azole sensitivity when compared to Cps. RNA-Seq and bioinformatic analyses found that following azole treatment, 55 genes in Cps, belonging to diverse pathways, displayed a significant decrease in expression. Putative xenobiotic detoxification genes overexpressed in tetraconazole-treated Che encoded predicted monooxygenase and oxidoreductase enzymes. In summary, expression of a functional CYP51A gene and overexpression of predicted xenobiotic detoxification genes appear likely to contribute to differential fungicide sensitivity in these two sister taxa. [ABSTRACT FROM AUTHOR]

Calonectria pseudonaviculata and C. henricotiae are two closely related fungal species responsible for boxwood blight disease of ornamental shrubs (Buxus spp.) in the U.S. and Europe. A previous study has shown isolates of the latter species, which is restricted to Europe, to be less sensitive to tetraconazole, an azole fungicide. In this study, we have analyzed the CYP51 paralogs for polymorphism in 26 genomes, representing geographically disparate populations of C. pseudonaviculata (n = 19) and C. henricotiae (n = 7), from the U.S., Europe, Asia, and New Zealand. The presence of a CYP51A pseudogene and lack of a functional CYP51A paralog in all C. pseudonaviculata genomes examined is a novel discovery for fungi and could have implications for the evolution of resistance to antifungal chemicals. [ABSTRACT FROM AUTHOR]

Functional toxicology has enabled the identification of genes involved in conferring tolerance and sensitivity to engineered nanomaterial (ENM) exposure in the model plant Arabidopsis thaliana (L.) Heynh. Several genes were found to be involved in metabolic functions, stress response, transport, protein synthesis, and DNA repair. Consequently, analysis of physiological parameters, metal content (through ICP-MS quantification), and gene expression (by RT-qPCR) of A. thaliana orthologue genes were performed across different plant species of agronomic interest to highlight putative biomarkers of exposure and effect related to ENMs. This approach led to the identification of molecular markers in Solanum lycopersicum L. and Cucurbita pepo L. (tomato and zucchini) that might not only indicate exposure to ENMs (CuO, CeO2, and La2O3) but also provide mechanistic insight into response to these materials. Through Gene Ontology (GO) analysis, the target genes were mapped in complex interatomic networks representing molecular pathways, cellular components, and biological processes involved in ENM response. The transcriptional response of 38 (out of 204) candidate genes studied varied according to particle type, size, and plant species. Importantly, some of the genes studied showed potential as biomarkers of ENM exposure and effect and may be useful for risk assessment in foods and in the environment. [ABSTRACT FROM AUTHOR]