Hanne N. Rasmussen

Abundance and distribution of Corallorhiza odontorhiza reflect variations in climate and ectomycorrhizae

The abundance and reproductive activity of orchids have been linked to variations in weather conditions, but few investigators have examined the relationships between orchid flowering dynamics and the distribution and abundance of mycorrhizal fungi. We quantified the abundance of flowering individuals of Corallorhiza odontorhiza, a mycoheterotrophic orchid, over a 14-year period and mapped the distribution of individuals in six of the 14 years. For two seasons, we conducted intensive and extensive studies of the mycorrhizal fungi that were associated with C. odontorhiza. The annual abundance of flowering plants was statistically related to growing-season precipitation and winter temperature, and the distribution of individuals within the study plot was related to the abundance and distribution of appropriate host fungi. We used DNA sequencing to identify ectomycorrhizal root tips that hosted Tomentella fungi that could potentially support C. odontorhiza. We found that Tomentella spp. were distributed thro...

Germination and seedling establishment in orchids: a complex of requirements

BACKGROUND Seedling recruitment is essential to the sustainability of any plant population. Due to the minute nature of seeds and early-stage seedlings, orchid germination in situ was for a long time practically impossible to observe, creating an obstacle towards understanding seedling site requirements and fluctuations in orchid populations. The introduction of seed packet techniques for sowing and retrieval in natural sites has brought with it important insights, but many aspects of orchid seed and germination biology remain largely unexplored. KEY CONSIDERATIONS The germination niche for orchids is extremely complex, because it is defined by requirements not only for seed lodging and germination, but also for presence of a fungal host and its substrate. A mycobiont that the seedling can parasitize is considered an essential element, and a great diversity of Basidiomycota and Ascomycota have now been identified for their role in orchid seed germination, with fungi identifiable as imperfect Rhizoctonia species predominating. Specificity patterns vary from orchid species employing a single fungal lineage to species associating individually with a limited selection of distantly related fungi. A suitable organic carbon source for the mycobiont constitutes another key requirement. Orchid germination also relies on factors that generally influence the success of plant seeds, both abiotic, such as light/shade, moisture, substrate chemistry and texture, and biotic, such as competitors and antagonists. Complexity is furthermore increased when these factors influence seeds/seedling, fungi and fungal substrate differentially. CONCLUSIONS A better understanding of germination and seedling establishment is needed for conservation of orchid populations. Due to the obligate association with a mycobiont, the germination niches in orchid species are extremely complex and varied. Microsites suitable for germination can be small and transient, and direct observation is difficult. An experimental approach using several levels of environmental manipulation/control is recommended.

Genetic diversity, compatibility patterns and seed quality in isolated populations of Cypripedium calceolus (Orchidaceae)

Cypripedium calceolus has suffered an alarming decline, and today mainly occurs in small and isolated populations. In Denmark there are only two populations, close to each other and situated far from other European stands. One population is stagnant or in slow decline, whereas the other is in rapid increase. We examined the levels of genetic diversity and compatibility and seed quality following experimental crosses. No genetic variation could be detected in plastid and nuclear markers within or between the two populations—in contrast to results previously reported from other European populations of C. calceolus. This may indicate a founder effect in both populations, but it could also be the outcome of prolonged inbreeding or reflect a genetic bottleneck after the populations were established. According to fruit dimensions and frequency of fully developed seeds there was full self-compatibility in the stagnant population, and partial late-acting self-incompatibility in the proliferating population. In combination with previous reports from other countries, this suggests that several self-incompatibility systems may occur in C. calceolus. Seeds from the older and stagnant population performed more poorly in germination tests in vitro than seeds from the thriving population. The difference needs not be genetically based, but could be due to environmental differences during seed maturation, producing different seed quality or dormancy characteristics. However, low level of genetic diversity within the populations may affect their ability to adapt and the possibility of inbreeding depression should be investigated.

Seedlings of Neuwiedia (Orchidaceae subfamily Apostasioideae) have typical orchidaceous mycotrophic protocorms

Naturally occurring seedlings of Neuwiedia veratrifolia were found in three localities in Sabah, Borneo, Malaysia. Seedlings consisted of an irregular oblong protocorm and a terminal leafy rooted shoot. Protocorms contained mycotrophic tissue of the kind typical of orchid mycorrhiza (tolypophagy). This finding demonstrates an important synapomorphy between Neuwiedia and other orchids and strongly supports the monophyly of Orchidaceae in the broad sense, including apostasiod orchids.

Composition of Cypripedium calceolus (Orchidaceae) seeds analyzed by attenuated total reflectance IR spectroscopy: In search of understanding longevity in the ground

UNLABELLED PREMISE OF THE STUDY Orchid seeds are minute and covered with a thin coat, yet they often have a long life after dispersal. They are notorious for low and irregular germination, in nature as well as in vitro. Since orchids are often rare species of conservational and commercial interest, reproduction by seeds is an important concern. The purpose of this study was to learn more about the resilience of these highly specialized seeds and stimulatory processes toward germination. • METHODS We studied testa and embryos of Cypripedium calceolus to identify natural components in intact seeds and the impact of 7 yr in soil in its natural habitat. We also analyzed the effects of Ca(OCl)2, used technically to enhance germination for cultivation in vitro. For the first time with this kind of plant material, we used attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy, an ideal method for minute sample sizes and surface selectivity. Thus, we could link treatments with changes in seed surface chemistry. • KEY RESULTS A lignin-like polymer is an essential testa component that undergoes degradation by soil or hypochlorite processes. In both cases, we found a build-up of CaCO3 on the testa, which could interact with lignin to enhance germination. Very minor changes occurred in embryo reserve nutrient content after a long sojourn underground, which supports their continued viability. • CONCLUSIONS We suggest that degradation of lignin and enrichment of the testa surface with CaCO3 are important stimulants of germination both in the habitat and during laboratory sowing.

Crown architecture and dynamics in Abies procera as influenced by cutting for greenery

Greenery production from 13 to 17 years old Abies procera was studied in three localities differing in soil and tree provenance. Two alternative clipping strategies for greenery were applied during 8 years. Subsequently, destructive crown analyses were carried out on the trees sampled from these and traditionally cut plots. Regeneration occurred from stubs or bases of cut primary branches. In cases when stubs were removed, proliferation took place around the scar; these branches compensated in numbers but not in biomass for the absence of stub branches. More severe cutting regimes resulted in an overall biomass reduction, most notably with shorter and lighter whorl branches. Interwhorl branches and branches issued from the base of primary branches were able to extend progressively in response to severe cutting. Base branches were apparently recruited along most of the trunk, the latency of growth points thus being up to at least 10 years. Competition among interwhorl branches, as well as more complicated interactions between whorl branches and secondary branches arising from their base, are indicated by the data. Reiteration directly from the stem could not be provoked and the results suggest that optimum greenery yield depends on careful stub management.

Cypripedium calceolus germination in situ: seed longevity, and dormancy breakage by long incubation and cold winters

A successful in situ germination experiment with Cypripedium calceolus , the European Lady’s slipper, is reported here for the first time. The seeds originated from controlled pollinations within and between two closely related Danish populations. The seeds were sown ripe in seed packets in proximity of mother plants. Germination was first observed after 4.5 y in the ground, following two successive cold and snowy winters, and only in one population. Seedlings expanded through the sides of the broken testa and were hair-less. A corresponding set of seeds, germinated in vitro as asymbiotic controls, responded positively to repeated cold stratifications after long incubation, suggesting that time (leaching?) and chilling are dormancy breakage factors.

Ontogeny in terminal buds of Abies nordmanniana (Pinaceae) characterized by ubiquitin

Meristematic activity in the bud meristem of Abies nordmanniana was visualized by ubiquitin immunohistochemical localization from before bud break and throughout shoot expansion. Ubiquitin was detected in meristematic cells either in the cytosol or nucleus, or both, depending on tissue type and developmental stage. During winter dormancy, ubiquitin was only observed in the protodermal/hypodermal layers, but at bud break in mid May, the signal expanded to the entire shoot tip. At the end of May, a clear zonation in ubiquitin localization appeared that lasted about one month. Throughout this period, ubiquitin was barely detectable in a central group of cells that might indicate an organizing center with stem cells. At the end of June, coinciding with the transition from scale leaf to needle primordia production, ubiquitin again was more prevalent in the peripheral cell layers. During shoot expansion, a strong ubiquitin signal developed in the axil of all needles. Most of these signals later disappeared, except for those few axils where buds actually developed. A strong ubiquitin signal was also observed in cells lining the young resin ducts. Our data showed that ubiquitin may be used as a marker for metabolic activity associated with seasonal development in the apical meristem.