Developmental aspects of Gigaspora rosea and Glomus etunicatum, alone or in association with Alnus glutinosa

Attempts were conducted to establish in vitro vesicular-arbuscular mycorrhizal (VAM) associations under gnotobiotic conditions. Alnus glutinosa (L.) Gaertn. served as the host in the form of whole plants, root organs, and undifferentiated callus. Spores of Gigaspora rosea Nicolson and Schenck or Glomus etunicatum Becker and Gerdemann were utilized as VAM fungus (VAMF) inoculum. Limited VAM associations were attained within certain in vitro culture systems that contained intact seedlings or root organs. In addition, whole seedling cultures exhibited occasional sporulation by Gl. etunicatum;The influences of chloramphenicol (150, 75, and 35 ppm), gentamicin sulfate (100 and 50 ppm), streptomycin sulfate (50 and 25 ppm), and penicillin G potassium (50,000 and 25,000 units/L) upon spore germination and hyphal growth by the two VAMF species also were evaluated in 1% water agar. Germination of Gl. etunicatum chlamydospores was inhibited by gentamicin, and hyphal growth was sensitive to higher concentrations of chloramphenicol and gentamicin. Higher levels of chloramphenicol also reduced germination and hyphal growth of Gi. rosea. Effects of other antibiotics were dependent upon their concentration and VAMF species. No significant correlation was found between spore diameter and hyphal growth for either VAMF species. Hyphal growth by Gi. rosea was, however, highly correlated with the number of extra-matrical vesicle clusters that were produced upon each hypha. When extra-matrical vesicle clusters became separated by a septum from the parental hypha, they occasionally generated a new radial mass of thin hyphae. Chlamydospores of Gl. etunicatum occasionally germinated directly through the spore wall, and hyphae from separate spores in close proximity frequently anastomosed;Light and scanning electron microscopy were utilized to observe various anatomical aspects in the development of Gi. rosea and Gl. etunicatum. These VA mycobionts were examined in an independent state and in association with roots of A. glutinosa

Molecular Identification of \u3ci\u3eArmillaria gallica\u3c/i\u3e from the Niobrara Valley Preserve in Nebraska

Armillaria isolates were collected from a unique forest ecosystem in the Niobrara Valley Preserve in Nebraska, USA, which comprises a glacial and early postglacial refugium in the central plains of North America. The isolates were collected from diverse forest trees representing a unique mixture of forest types. Combined methods of rDNA sequencing and flow cytometric measurements of nuclear DNA content determined that all Armillaria isolates collected from the site were A. gallica

Use of flow cytometry, fluorescence microscopy, and PCR-based techniques to assess intraspecific and interspecific matings of \u3ci\u3eArmillaria\u3c/i\u3e species

For assessments of intraspecific mating using flow cytometry and fluorescence microscopy, two compatible basidiospore-derived isolates were selected from each of four parental basidiomata of North American Biological Species (NABS) X. The nuclear status in NABS X varied with basidiospore-derived isolates. Nuclei within basidiospore-derived isolates existed as haploids, diploids (doubled haploids), or a mixture of haploids and diploids (doubled haploids). Depending on the nuclear status of the basidiospore-derived lines of NABS X, intraspecifically mated cultures can exist as diploids or tetraploids, and possibly triploids or aneuploids under in vitro conditions. Based on previous in vitro mating studies, seven basidiospore isolates were specifically selected to assess rare, interspecific mating among Armillaria cepistipes, A. sinapina, NABS X, and NABS XI. Cultures from basidiospore-derived isolates were paired to produce four interspecifically paired cultures, and matings were assessed using flow cytometry and restriction fragment length polymorphism (RFLP) analyses. Based on flow cytometric analysis, the A. cepistipes isolate exhibited compatibility with a NABS X isolate, and the A. sinapina isolate exhibited compatibility with a NABS X isolate, and the A. sinapina isolates were individually compatible with isolates of NABS X and NABS XI. Mean fluorescence intensities of A. cepistipes x NABS X, A. sinapina x NABS X, and A. sinapina x NABS XI mated cultures revealed a triploid or tetraploid nuclear status compared to the haploid or diploid (doubled haploid) nuclear status of initial basidiospore-derived isolates. Polymerase chain reaction (PCR) and RFLP of the intergenic spacer (IGS) region generated banding patterns for basidiospore-derived isolates and mated cultures. Four species-specific RFLP banding patterns were observed in basidiospore-derived isolates of A. cepistipes, A. sinapina, NABS X, and NABS XI. PCR-RFLP analysis showed combined banding patterns from mated cultures. Flow cytometry and PCR-RFLP analysis are effective tools to assess matings of Armillaria species

Molecular Characterization of \u3ci\u3eFusarhm oxysporum\u3c/i\u3e and \u3ci\u3eFusarium commune\u3c/i\u3e Isolates from a Conifer Nursery

Fusarium species can cause severe root disease and damping-off in conifer nurseries. Fusarium inoculum is commonly found in most container and bareroot nurseries on healthy and diseased seedlings, in nursery soils, and on conifer seeds. Isolates of Fusarium spp. can differ in virulence; however, virulence and colony morphology are not correlated. Forty-one isolates of Fusarium spp., morphologically indistinguishable from F. oxysporum, were collected from nursery samples (soils, healthy seedlings, and diseased seedlings). These isolates were characterized by amplified fragment length polymorphism (AFLP) and DNA sequencing of nuclear rDNA (internal transcribed spacer including 5.8s rDNA), mitochondrial rDNA (small subunit [mtSSU]), and nuclear translation elongation factor I-alpha. Each isolate had a unique AFLP phenotype. Out of 121 loci, 11 1 (92%) were polymorphic; 30 alleles were unique to only highly virulent isolates and 33 alleles were unique to only isolates nonpathogenic on conifers. Maximum parsimony and Bayesian analyses of DNA sequences from all three regions and the combined data set showed that all highly virulent isolates clearly separated into a common clade that contained F. commune, which was recently distinguished from its sister taxon, F. oxysporum. Interestingly, all but one of the nonpathogenic isolates grouped into a common clade and were genetically similar to F. oxysporum. The AFLP cladograms had similar topologies when compared with the DNA-based phylograms. Although all tested isolates were morphologically indistinguishable from F. oxysporum based on currently available monographs, some morphological traits can be plastic and unreliable for identification of Fusarium spp. We consider the highly virulent isolates to be F. commune based on strong genetic evidence. To our knowledge, this is the first reported evidence that shows F. commune is a cause of Fusarium disease (root rot and damping-off) on Douglas-fir seedlings. Furthermore, several AFLP genetic markers and mtSSU sequences offer potential for development of molecular markers that could be used to detect and distinguish isolates of F. oxysporum nonpathogenic to conifers and highly virulent isolates of F. commune in forest nurseries

<i>Armillaria altimontana</i> in North America: Biology and Ecology

Armillaria altimontana is a fungus (Basidiomycota, Agaricomycetes, Agaricales, and Physalacriaceae) that is generally considered as a weak/opportunistic pathogen or saprophyte on many tree hosts. It widely occurs across the northwestern USA to southern British Columbia, Canada, but relatively little is known about its ecological role in the diverse forest ecosystems where it occurs. This review summarizes the biology and ecology of A. altimontana, including its identification, life cycle, distribution, host associations, and bioclimatic models under climate change

Silvopasture: An Agroforestry Practice

Although some form of silvopasture management has been practiced for centuries, silvopasture as an agroforestry practice is specifically designed and managed for the production of trees, tree products, forage, and livestock. Silvopasture results when forage crops are deliberately introduced or enhanced in a timber production system, or timber crops are deliberately introduced or enhanced in a forage production system. As a silvopasture, timber and pasture are managed as a single integrated system. Silvopastoral systems are designed to produce a high-value timber component, while providing short-term cash flow from the livestock component. The interactions among timber, forage, and livestock are managed intensively to simultaneously produce timber commodities, a high quality forage resource, and efficient livestock production. Overall, silvopastures can provide economic returns while creating a sustainable system with many environmental benefits. Well-managed silvopastures offer a diversified marketing opportunity that can stimulate rural economic development

Molecular Characterization of Fusarium oxysporum and Fusarium commune Isolates from a Conifer Nursery

Fusarium species can cause severe root disease and damping-off in conifer nurseries. Fusarium inoculum is commonly found in most container and bareroot nurseries on healthy and diseased seedlings, in nursery soils, and on conifer seeds. Isolates of Fusarium spp. can differ in virulence; however, virulence and colony morphology are not correlated. Forty-one isolates of Fusarium spp., morphologically indistinguishable from F. oxysporum, were collected from nursery samples (soils, healthy seedlings, and diseased seedlings). These isolates were characterized by amplified fragment length polymorphism (AFLP) and DNA sequencing of nuclear rDNA (internal transcribed spacer including 5.8S rDNA), mitochon-drial rDNA (small subunit [mtSSU]), and nuclear translation elongation factor 1-alpha. Each isolate had a unique AFLP phenotype. Out of 121 loci, 111 (92%) were polymorphic; 30 alleles were unique to only highly virulent isolates and 33 alleles were unique to only isolates nonpathogenic on conifers. Maximum parsimony and Bayesian analyses of DNA sequences from all three regions and the combined data set showed that all highly virulent isolates clearly separated into a common clade that contained F. commune, which was recently distinguished from its sister taxon, F. oxysporum. Interestingly, all but one of the nonpathogenic isolates grouped into a common clade and were genetically similar to F. oxysporum. The AFLP cladograms had similar topologies when compared with the DNA-based phylograms. Although all tested isolates were morphologically indistinguishable from F. oxysporum based on currently available monographs, some morphological traits can be plastic and unreliable for identification of Fusarium spp. We consider the highly virulent isolates to be F. commune based on strong genetic evidence. To our knowledge, this is the first reported evidence that shows F. commune is a cause of Fusarium disease (root rot and dampingoff) on Douglas-fir seedlings. Furthermore, several AFLP genetic markers and mtSSU sequences offer potential for development of molecular markers that could be used to detect and distinguish isolates of F. oxysporum nonpathogenic to conifers and highly virulent isolates of F. commune in forest nurseries