George C. Carroll

Infection studies of Discula quercina, an endophyte of Quercus garryana

Discula quercina is a ubiquitous endophyte in Oregon white oak (Quercus garryana). We determined which tree parts were infected with this endophyte, censused how infection levels in leaves varied o...

Fluorescein diacetate hydrolysis as an estimator of microbial biomass on coniferous needle surfaces

Estimating microbial standing crops and microbial production in natural habitats has been difficult for microbial ecologists. The present paper describes a simple spectrophotometric assay based on the hydrolysis of fluorescein diacetate which estimates well the standing crops of microbial cells on coniferous needles and twigs. A technique is also presented for correlating optical density readings with actual dry weights of microbial cells epiphytic on needles, and thus for standardizing the assay. The assay shows promise of broad applicability to other microbial habitats.

PATTERNS OF SUBSTRATE UTILIZATION BY SOME FUNGAL ENDOPHYTES FROM CONIFEROUS FOLIAGE

Fifty-six strains of endophytic fungi isolated from coniferous foliage in Europe and in Oregon, U.S.A., were tested for their ability to utilize various complex substrates including cellulose, xylan, mannan, several other hemicelluloses, pectin, and two lignin derivatives. Cultures were also assayed for lipolytic activity, laccase production, growth on gallic acid, and production of darkly pigmented reaction products on gallic acid. A majority of isolates utilized xylan and pectin, showed lipolytic activity, and grew on gallic acid. Fewer isolates (54%) utilized cellulose, and only a small proportion utilized hemicelluloses other than xylan. Among isolates from conifers with distinct needles, fungi occurring only in the petioles showed a broad range of substrate utilization capabilities, while those from the needle blades were more restricted in their abilities. Isolates from cuppressaceous foliage utilized xylan but varied greatly with respect to other capabilities. Different isolates of the same fungus often varied in their substrate utilization capacities. Substrate utilization patterns of all the common endophytes from a single host suggested that several endophytes may co-exist within a single leaf by biochemical partitioning of resources.

The internal mycobiota of Juncus spp.: microscopic and cultural observations of infection patterns

Differences in species composition, infection frequencies and fungal colonization were compared in asymptomatic leaves and culms of one annual and three perennial Juncus species in western Oregon. Stagonospora innumerosa was the principal fungus species isolated from culms of the annual, J. bufonius , and two of the perennial, J. effusus and J. patens , host species; Phaeosphaeria juncicola and Drechslera sp. were isolated from leaves of J. bolanderi . Characteristic mechanisms of penetration and colonization corresponding to individual endophyte species were observed in host tissue. Infections limited to a single host epidermal cell were characteristic of S. innumerosa, Drechslera sp. and an unidentified endophyte of J. bufonius . Infections by P. junicola originate in the substomatal cavity followed by limited intercellular colonization of the mesophyll. Cladosporium cladosporioides and Alternaria alternata were isolated at low frequencies from J. effusus, J. patens and J. bolanderi . Infections by C. cladosporioides were restricted to the substomatal chambers, and this species did not colonize internal leaf tissue. Infections of A. alternata were also localized in the substomatal chamber, frequently with an intercellular colonization of the mesophyll tissue.

AVOIDANCE OF HIGH-ENDOPHYTE SPACE BY GALL-FORMING INSECTS

Endophytic fungi and phytophagous insects derive energy from shared host plants, and so are likely to interact with one another. Several authors have proposed that fungal endophytes of woody perennials, like endophytes of grasses, have a mutualistic association with their host and provide enhanced protection from herbivory. However, there have been few tests of this hypothesis, and most studies have focused on correlating the presence of endophytes and herbivores. We investigated a system where an endophyte was known to cause significant mortality on one species of gall-forming insect (Besbicus mi- rabilis), but not another (Bassettia ligni) that occupies the same plant host. We predicted that gall insects should avoid high-endophyte space and that mortality would be lowest in low-endophyte space if endophytes protect plants from herbivory. We used the spatial location, temporal infection patterns, and within-leaf growth activity of the endophyte as elements of endophyte space. Besbicus mirabilis avoided high-endophyte space at the within-leaf scale by occupying part of the leaf where endophyte infection and endophyte-caused mortality were predictably low. However, the within-leaf position was under opposing selection pressure from mortality caused by grazing herbivores and developmental constraints, so the gall did not occupy the region of the leaf where both infection and endophyte-caused mortality were the lowest.- B. ligni was spatially correlated within leaves with the presence of the endophyte but suffered almost no endophyte-caused mortality. However, B. ligni occupies the leaf lamina, where the endophyte has no growth activity, in contrast to the leaf midrib, where B. mirabilis galls are found. Further, B. ligni emerges from the gall just as endophyte infection levels reach a peak, so this species temporally avoids prolonged contact with highly infected tissue. However, the between-leaf distributions of both insects were independent of en- dophyte infection. Fungal endophytes influence the spatial patterns of insect distributions, but at different scales. Growth activity of the endophyte may also play a role, and simply examining presence/absence correlations of insects may be misleading.

GENOTYPIC DIVERSITY IN POPULATIONS OF A FUNGAL ENDOPHYTE FROM DOUGLAS FIR

Rhabdocline parkeri, an endophyte of Douglas fir, forms symptomless infections within single epidermal cells of healthy needles. Rhabdocline parkeri strains were isolated from trees growing in various habitats ranging from virgin old growth forests to a single young tree growing isolated in a meadow. Different genotypes of R. parkeri were distinguished by using the Random Amplified Polymorphic DNA (RAPD) technique. A significantly lower number of R. parkeri genotypes per unit foliage were isolated from trees within a 20-year-old managed stand and from an isolated tree than from old growth trees. Foliage from young trees (20 years old) in a stand directly adjacent to the sampled old growth trees showed levels of genotypic endophyte diversity comparable to that in the old growth trees. Genotypic diversity is estimated to be at least three orders of magnitude greater in foliage of the old growth trees and adjacent young trees than in foliage from the managed stand and from an isolated tree. This difference is ascribed to differences in tree age and access to inoculum.

Fungal Succession on Needles and Young Twigs of Old-Growth Douglas Fir

The pattern of presence and abundance of fungi on needles and twigs of old-growth Douglas fir (Pseudotsuga menciesii) in the Oregon Cascades exhibits a well-defined successional sequence which was documented by counting thalli and fruiting bodies under a dissecting microscope. Detailed information on the distribution of fungi in this habitat suggests that their mode of nutrition has yet to be elucidated.

SYNAPTONEMAL COMPLEXES AND MEIOSIS IN DIDYMIUM IRIDIS: A REINVESTIGATION

SUMMARY The ultrastructure of meiotic events has been studied in three isolates of Didymium iridis, including the one used by Carroll and Dykstra in an earlier study. In all three isolates a precleavage mitotic division occurs. About 9-12 hr after spore cleavage, synaptonemal complexes appear in virtually all spore nuclei, indicating the onset of meiotic prophase. Subsequent division stages have also been observed. Attempts to change the time of meiosis by manipulation of the time of onset of illumination were unsuccessful. Re-examination of the original blocks used by Carroll and Dykstra revealed that the sporangium containing the synaptonemal complexes studied by them represented an incompletely cleaved, aberrant development and was in reality a stage about 12-18 hr after an abnormal cleavage.

Isozyme polymorphism in six endophytic Phyllosticta species

Isozyme variation of 70 endophytic Phyllosticta isolates representing four species from coniferous hosts and two species from angiospermous hosts was examined using starch gel electrophoresis of ten enzymes. All enzymes were polymorphic among species and each electromorph was restricted to a single host. Strains of Phyllosticta pseudotsugae were most diverse and separated into seven phenotypes. In Ph. cryptomeriae five phenotypes and in Ph. abietis two phenotypes were detected. The remaining species (Ph. multicorniculata, Ph. ampelicida, Ph. pyrolae) were monomorphic. Interspecific variation was always very high, with differences in at least 70% of the electromorphs. Within some species the same amount of variation was present, suggesting the possibility of splitting these taxa in subspecific entities. Cluster and principal components analysis of phenotype frequencies revealed two clearly separated groups that corresponded to species from different continents. The two species from angiospermous hosts were also distinct from the other species.

Testing assumptions of the enemy release hypothesis: generalist versus specialist enemies of the grass Brachypodium sylvaticum

The enemy release hypothesis (ERH) suggests greater success of species in an invaded range due to release from natural enemies. The ERH assumes there will be more specialist enemies in the native range and that generalists will have an equal effect in both ranges. We tested these assumptions with the grass Brachypodium sylvaticum in the native range (Switzerland) and invaded range (Oregon, USA). We assessed all the kinds of damage present (caused by fungi, insects, mollusk and deer) on both leaves and seeds at 10 sites in each range and correlated damage with host fitness. Only two of the 20 fungi found on leaves were specialist pathogens, and these were more frequent in the native range. Conversely there was more insect herbivory on leaves in the invaded range. All fungi and insects found on seeds were generalists. More species of fungi were found on seeds in the native range, and a higher proportion of them were pathogenic than in the invaded range. There were more kinds of enemies in the native range, where the plants had lower fitness, in accordance with the ERH. However, contrary to assumptions of the ERH, generalists appear to be equally or more important than specialists in reducing host fitness.