Global migration patterns in the fungal wheat pathogen Phaeosphaeria nodorum

The global migration patterns of the fungal wheat pathogen Phaeosphaeria nodorum were analysed using 12 microsatellite loci. Analysis of 693 isolates from nine populations indicated that the population structure of P. nodorum is characterized by high levels of genetic diversity and a low degree of subdivision between continents. To determine whether genetic similarity of populations was a result of recent divergence or extensive gene flow, the microsatellite data were analysed using an isolation-with-migration model. We found that the continental P. nodorum populations diverged recently, but that enough migration occurred to reduce population differentiation. The migration patterns of the pathogen indicate that immigrants originated mainly from populations in Europe, China and North America.     

Isolation and characterization of EST‐derived microsatellite loci from the fungal wheat pathogen Phaeosphaeria nodorum

Eleven polymorphic microsatellite loci and one minisatellite locus originating from expressed sequence tag (EST) libraries of Phaeosphaeria (syn. Stagonospora) nodorum were isolated and characterized. The satellite markers were used to genotype isolates from field populations collected in China, North America and South Africa. The number of alleles per locus ranged from two to 15. Genotype diversity ranged from 87.5 to 95.3 and gene diversity from 0.1 to 0.8. The variable levels of polymorphism within and among populations of P. nodorum renders these 12 satellite loci ideal markers for population genetic analysis of P. nodorum.     

Long-term relationships between environment and abundance in wheat of Phaeosphaeria nodorum and Mycosphaerella graminicola

Relationships between weather, agronomic factors and wheat disease abundance were examined to determine possible causes of variability on century time scales. In archived samples of wheat grain and leaves obtained from the Rothamsted Broadbalk experiment archive (1844-2003), amounts of wheat, Phaeosphaeria nodorum and Mycosphaerella graminicola DNA were determined by quantitative polymerase chain reaction (PCR). Relationships between amounts of pathogens and environmental and agronomic factors were examined by multiple regression. Wheat DNA decayed at approx. 1% yr(-1) in stored grain. No M. graminicola DNA was detected in grain samples. Fluctuations in amounts of P. nodorum in grain were related to changes in spring rainfall, summer temperature and national SO(2) emission. Differences in amounts of P. nodorum between grain and leaf were related to summer temperature and spring rainfall. In leaves, annual variation in spring rainfall affected both pathogens similarly, but SO(2) had opposite effects. Previous summer temperature had a highly significant effect on M. graminicola. Cultivar effects were significant only at P = 0.1. Long-term variation in P. nodorum and M. graminicola DNA in leaf and grain over the period 1844-2003 was dominated by factors related to national SO(2) emissions. Annual variability was dominated by weather factors occurring over a period longer than the growing season.     

Contrasting patterns of nonneutral evolution in proteins encoded in nuclear and mitochondrial genomes

We report that patterns of nonneutral DNA sequence evolution among published nuclear and mitochondrially encoded protein-coding loci differ significantly in animals. Whereas an apparent excess of amino acid polymorphism is seen in most (25/31) mitochondrial genes, this pattern is seen in fewer than half (15/36) of the nuclear data sets. This differentiation is even greater among data sets with significant departures from neutrality (14/15 vs. 1/6). Using forward simulations, we examined patterns of nonneutral evolution using parameters chosen to mimic the differences between mitochondrial and nuclear genetics (we varied recombination rate, population size, mutation rate, selective dominance, and intensity of germ line bottleneck). Patterns of evolution were correlated only with effective population size and strength of selection, and no single genetic factor explains the empirical contrast in patterns. We further report that in Arabidopsis thaliana, a highly self-fertilizing plant with effectively low recombination, five of six published nuclear data sets also exhibit an excess of amino acid polymorphism. We suggest that the contrast between nuclear and mitochondrial nonneutrality in animals stems from differences in rates of recombination in conjunction with a distribution of selective effects. If the majority of mutations segregating in populations are deleterious, high linkage may hinder the spread of the occasional beneficial mutation.     

Mating Types of Phaeosphaeria nodorum (anamorph Stagonospora nodorum) from Central Asia

The distribution was examined of the mating type idiomorphs (MAT‐1 and ‐2) of Phaeosphaeria (anamorph Stagonospora) nodorum using DNA from 49 isolates collected from commercial and experimental fields in 2003 and 2004 in Central Asia. MAT‐1 and ‐2 isolates were present in the Kazakh and Russian origins of P. nodorum, but no MAT‐2 isolates were found in Tajikistan. The possibility of a skewed Tajik population cannot be excluded, considering that the sampled region in Tajikistan was geographically isolated from Kazakhstan and Russia.     

Interaction of nuclear and mitochondrial genomes in yeasts

The review is focused on the bulk of information on the interaction between nuclear and mitochondrial genes gained in studies of nuclear and mitochondrial mutations interaction. This approach is the most adequate for identification of nuclear genes controlling the mitochondrial genomes' expression. It also permits to study the effects of mitochondrial genome state on expression of nuclear genes.     

A metabolomic approach to dissecting osmotic stress in the wheat pathogen Stagonospora nodorum

A non-targeted metabolomics approach was used to identify significant changes in metabolism upon exposure of the wheat pathogen Stagonospora nodorum to 0.5M NaCl. The polyol arabitol, and to a lesser extent glycerol, was found to accumulate in response to the osmotic stress treatment. Amino acid synthesis was strongly down-regulated whilst mannitol levels were unaffected. A reverse genetic approach was undertaken to dissect the role of arabitol metabolism during salt stress. Strains of S. nodorum lacking a gene encoding an l-arabitol dehydrogenase (abd1), a xylitol dehydrogenase (xdh1) and a double-mutant lacking both genes (abd1xdh1) were exposed to salt and the intracellular metabolites analysed. Arabitol levels were significantly up-regulated upon salt stress in the xdh1 strains but were significantly lower than the wild-type. Arabitol was not significantly different in either the abd1 or the abd1xdh1 strains during osmotic stress but the concentration of glycerol was significantly higher indicating a compensatory mechanism in operation. Genome sequence analysis identified a second possible enzyme capable of synthesizing arabitol explaining the basal level of arabitol present in the abd1xdh1 strains. This study identified that arabitol is the primary compatible solute in S. nodorum but in-built levels of redundancy are present allowing the fungus to tolerate osmotic stress.     

Photochemical characterization of a novel fungal rhodopsin from Phaeosphaeria nodorum

Eukaryotic microbial rhodopsins are widespread bacteriorhodopsin-like proteins found in many lower eukaryotic groups including fungi. Many fungi contain multiple rhodopsins, some significantly diverged from the original bacteriorhodopsin template. Although few fungal rhodopsins have been studied biophysically, both fast-cycling light-driven proton pumps and slow-cycling photosensors have been found. The purpose of this study was to characterize photochemically a new subgroup of fungal rhodopsins, the so-called auxiliary group. The study used the two known rhodopsin genes from the fungal wheat pathogen, Phaeosphaeria nodorum. One of the genes is a member of the auxiliary group while the other is highly similar to previously characterized proton-pumping Leptosphaeria rhodopsin. Auxiliary rhodopsin genes from a range of species form a distinct group with a unique primary structure and are located in carotenoid biosynthesis gene cluster. Amino acid conservation pattern suggests that auxiliary rhodopsins retain the transmembrane core of bacteriorhodopsins, including all residues important for proton transport, but have unique polar intramembrane residues. Spectroscopic characterization of the two yeast-expressed Phaeosphaeria rhodopsins showed many similarities: absorption spectra, conformation of the retinal chromophore, fast photocycling, and carboxylic acid protonation changes. It is likely that both Phaeosphaeria rhodopsins are proton-pumping, at least in vitro. We suggest that auxiliary rhodopsins have separated from their ancestors fairly recently and have acquired the ability to interact with as yet unidentified transducers, performing a photosensory function without changing their spectral properties and basic photochemistry.     

Identity and conservation of mating type genes in geographically diverse isolates of Phaeosphaeria nodorum

Mating type idiomorphs (MAT1-1 and MAT1-2) were identified from the heterothallic loculoascomycete Phaeosphaeria nodorum (wheat biotype) using DNA from a pair of isolates from Poland and Georgia, USA that are known to mate. MAT predicted proteins of P. nodorum are similar in sequence and in phylogenetic relationship to those described for other loculoascomycetes such as Cochliobolus spp., Alternaria alternata, and Didymella zeae-maydis. The organization of the MAT locus of the P. nodorum differs from these species in that its idiomorph begins within an adjacent upstream conserved ORF of unknown function. MAT-specific primers were used to identify isolates of both mating types in field populations, demonstrating that an absence of either mating type is not the reason that the teleomorph has not been found in New York. Portions of MAT1-1 and MAT1-2 were sequenced from geographically diverse isolates, including those from regions where the teleomorph has been reported. MAT was highly conserved and no significant differences in sequence were found.     

Parallel evolution of the genetic code in arthropod mitochondrial genomes

The genetic code provides the translation table necessary to transform the information contained in DNA into the language of proteins. In this table, a correspondence between each codon and each amino acid is established: tRNA is the main adaptor that links the two. Although the genetic code is nearly universal, several variants of this code have been described in a wide range of nuclear and organellar systems, especially in metazoan mitochondria. These variants are generally found by searching for conserved positions that consistently code for a specific alternative amino acid in a new species. We have devised an accurate computational method to automate these comparisons, and have tested it with 626 metazoan mitochondrial genomes. Our results indicate that several arthropods have a new genetic code and translate the codon AGG as lysine instead of serine (as in the invertebrate mitochondrial genetic code) or arginine (as in the standard genetic code). We have investigated the evolution of the genetic code in the arthropods and found several events of parallel evolution in which the AGG codon was reassigned between serine and lysine. Our analyses also revealed correlated evolution between the arthropod genetic codes and the tRNA-Lys/-Ser, which show specific point mutations at the anticodons. These rather simple mutations, together with a low usage of the AGG codon, might explain the recurrence of the AGG reassignments.     

Comparative rates of evolution in endosymbiotic nuclear genomes

Background  

The nucleomorphs associated with secondary plastids of cryptomonads and chlorarachniophytes are the sole examples of organelles with eukaryotic nuclear genomes. Although not as widespread as their prokaryotic equivalents in mitochondria and plastids, nucleomorph genomes share similarities in terms of reduction and compaction. They also differ in several aspects, not least in that they encode proteins that target to the plastid, and so function in a different compartment from that in which they are encoded.     

The structure and evolution of angiosperm nuclear genomes

Despite several decades of investigation, the organization of angiosperm genomes remained largely unknown until very recently. Data describing the sequence composition of large segments of genomes, covering hundreds of kilobases of contiguous sequence, have only become available in the past two years. Recent results indicate commonalities in the characteristics of many plant genomes, including in the structure of chromosomal components like telomeres and centromeres, and in the order and content of genes. Major differences between angiosperms have been associated mainly with repetitive DNAs, both gene families and mobile elements. Intriguing new studies have begun to characterize the dynamic three-dimensional structures of chromosomes and chromatin, and the relationship between genome structure and co-ordinated gene function.     

Onchocerca volvulus: limited heterogeneity in the nuclear and mitochondrial genomes

West African populations of Onchocerca volvulus endemic to the rain forest and savanna bioclimes of West Africa differ in their ability to induce ocular disease in infected individuals. In recent years, both clinical- and animal-model-based studies have implicated particular parasite antigens in the development of ocular onchocerciasis. To test the hypothesis that the difference in pathogenic potential of blinding and nonblinding parasites might be reflected in qualitative differences in antigens that have been implicated in the development of ocular onchocerciasis, we compared the sequences of two parasite antigens implicated in the development of ocular disease in blinding- and nonblinding-strain parasites. The results demonstrated a high level of homogeneity between the parasite strains in these genes. The study was extended to include additional nuclear genes encoding antigens that are commonly recognized by individuals infected with O. volvulus and to the mitochondrial genome of the parasite. The results demonstrate a high degree of homogeneity in both the nuclear and the mitochondrial genomes among O. volvulus isolates collected from several different sites in Africa and in the Americas. This high degree of genetic homogeneity may reflect the passage of the parasite through a recent genetic bottleneck.     

Mannitol 1-phosphate metabolism is required for sporulation in planta of the wheat pathogen Stagonospora nodorum

An expressed sequence tag encoding a putative mannitol 1-phosphate dehydrogenase (Mpd1) has been characterized from the fungal wheat pathogen Stagonospora nodorum. Mpd1 was disrupted by insertional mutagenesis, and the resulting mpd1 strains lacked all detectable NAD-linked mannitol 1-phosphate dehydrogenase activity (EC 1.1.1.17). The growth rates, sporulation, and spore viability of the mutant strains in vitro were not significantly different from the wild type. The viability of the mpd1 spores when subjected to heat stress was comparable to wild type. Characterization of the sugar alcohol content by nuclear magnetic resonance spectroscopy revealed that, when grown on glucose, the mutant strains contained significantly less mannitol, less arabitol, but more trehalose than the wild-type strains. The mannitol content of fructose-grown cultures was normal. No secreted mannitol could be detected in wild type or mutants. Pathogenicity assays revealed the disruption of Mpd1 did not affect lesion development, however the mutants were unable to sporulate. These results throw new light on the role of mannitol in fungal plant interactions, suggesting a role in metabolic and redox regulation during the critical process of sporulation on senescing leaf material.     

Dynamic evolution of eukaryotic mitochondrial and nuclear genomes: a case study in the gourmet pine mushroom Tricholoma matsutake

Fungi, as eukaryotic organisms, contain two genomes, the mitochondrial genome and the nuclear genome, in their cells. How the two genomes evolve and correlate to each other is debated. Herein, taking the gourmet pine mushroom Tricholoma matsutake as an example, we performed comparative mitogenomic analysis using samples collected from diverse locations and compared the evolution of the two genomes. The T. matsutake mitogenome encodes 49 genes and is rich of repetitive and non-coding DNAs. Six genes were invaded by up to 11 group I introns, with one cox1 intron cox1P372 showing presence/absence dynamics among different samples. Bioinformatic analyses suggested limited or no evidence of mitochondrial heteroplasmy. Interestingly, hundreds of mitochondrial DNA fragments were found in the nuclear genome, with several larger than 500 nt confirmed by PCR assays and read count comparisons, indicating clear evidence of transfer of mitochondrial DNA into the nuclear genome. Nuclear DNA of T. matsutake showed a higher mutation rate than mitochondrial DNA. Furthermore, we found evidence of incongruence between phylogenetic trees derived from mitogenome and nuclear DNA sequences. Together, our results reveal the dynamic genome evolution of the gourmet pine mushroom.     

Homologies to plastid DNA in the nuclear and mitochondrial genomes of potato

Summary Potato plastid DNA clones, representing onefourth of the potato plastome complexity and containing sequences of the 16SrRNA, rps16, atpA, atpE, psaA, psaB, trnK, trnV, and trnG genes, were used as hybridization probes on nuclear- and mitochondrial-enriched DNAs. Each probe hybridized to multiple nuclear restriction fragments distinct from the plastid cleavage products generated by the same endonucleases. The nuclear hybridizable fragments are highly methylated at their Hpall target sequences (C/CGG). In some instances, the transfer seemed to involve plastid regions of several kilobase pairs, as reflected by the co-integration in the nucleus of restriction sites that are distant in the plastome. Three clones hybridized additionally to distinct mitochondrial fragments. These results indicate that extensive DNA transfers did occur between plastids and other organelles in potato.     

Mannitol is required for asexual sporulation in the wheat pathogen Stagonospora nodorum (glume blotch)

The physiological role of the mannitol cycle in the wheat pathogen Stagonospora nodorum (glume blotch) has been investigated by reverse genetics and metabolite profiling. A putative mannitol 2-dehydrogenase gene (Mdh1) was cloned by degenerate PCR and disrupted. The resulting mutated mdh1 strains lacked all detectable NADPH-dependent mannitol dehydrogenase activity. The mdh1 strains were unaffected for mannitol production but, surprisingly, were still able to utilize mannitol as a sole carbon source, suggesting a hitherto unknown mechanism for mannitol catabolism. The mutant strains were not compromised in their ability to cause disease or sporulate. To further our understanding of mannitol metabolism, a previously developed mannitol-1-phosphate dehydrogenase (gene mpd1) disruption construct [Solomon, Tan and Oliver (2005) Mol. Plant-Microbe Interact. 18, 110-115] was introduced into the mutated mdh1 background, resulting in a strain lacking both enzyme activities. The mpd1mdh1 strains were unable to grow on mannitol and produced only trace levels of mannitol. The double-mutant strains were unable to sporulate in vitro when grown on minimal medium for extended periods. Deficiency in sporulation was correlated with the depletion of intracellular mannitol pools. Significantly sporulation could be restored with the addition of mannitol. Pathogenicity of the double mutant was not compromised, although, like the previously characterized mpd1 mutants, the strains were unable to sporulate in planta. These findings not only question the currently hypothesized pathways of mannitol metabolism, but also identify for the first time that mannitol is required for sporulation of a filamentous fungus.