Detection of high-phosphorus tolerant VAM-fungi colonizing hops and peppermint

Summary In a continuing search for field sources of endomycorrhizal fungi that tolerate high levels of available phosphorus in soil, samples were obtained from long-established liberally fertilized hops (Humulus lupulus L.) and peppermint (Mentha piperita L.) fields. In hops fields, where extractable P (Bray) levels ranged from 21–196 ppm, spores of twoAcaulospora spp. and sixGlomus spp. were isolated. From peppermint fields with P levels of 44 to 244 ppm, spores of oneAcaulospora sp., fourGlomus spp. and oneGigaspora sp. were obtained. Vesicular-arbuscular mycorrhizal (VAM) fungi were found colonizing most roots to some extent, despite the very high fertility of several sites. Although percentages of colonization in hops tended to be low, ranging from 0 to 9.3%, colonizations in peppermint roots were considerably higher averaging 26.5% between the sampled fields, notwithstanding P levels as high as 244 ppm. Curiously, considerable external VAM hyphae were found adherent to or entangling roots in many cases, even where percentage of root length with internal VAM structures was low. The functional benefit, neutrality, or detriment to plants of such high-P tolerant species remains to be determined.Contribution from the USDA-ARS, Horticultural Crops Research Laboratory and the Dept. of Soil Science, Oregon State University, in cooperation with the Oregon State University Agricultural Experiment Station, Corvallis, OR 97331. Technical paper no. 6979 of the latter     

Above and belowground community development in a marine sand dune ecosystem

Transects across undisturbed marine sand dunes sequentially traversed the following plant communities: littoral, foredunes, intradune complex of ridges and hollows, deflation plain, myrtle forest, and mature conifer forest. Organic carbon levels were low in the littoral zone and increased across the dune ecosystem landward to the forest communities. The highest percentage of nutrients was isolated from the heavy fraction of soil residues. Soil microfloral populations responded to vegetation, physical dune characteristics, and seasonal moisture patterns. Populations of bacteria and actinomycetes were higher in winter than during summer sampling periods in all communities. Populations of microscopic fungi were higher in winter in all the communities except the hollows. The distribution of vesicular-arbuscular mycorrhizal fungi responded to vegetation and sand dune succession but did not display seasonality. Species of Gigaspora and Acaulospora were the most commonly isolated VAM fungi.Sand aggregation increased along the dune transect and was correlated to plant community succession: the most highly aggregated soil was found in the two forest communities. With scanning electron microscopy, sand grains and organic residues were observed entangled by strands of filamentous mciroorganisms. Many of the filaments were of the dimensions of VAM fungi, which may be important for the release of nutrients associated with the cementing agents of sand aggregates and for the survival of early pioneer plants of sand dunes.     

Effect of salinity on the association between root symbionts and Acacia cyanophylla Lind.: growth and nutrition

The behaviour of Acacia cyanophylla Lind. plants submitted to salinity stress was followed in the greenhouse. The plants were associated with indigenous symbiotic microorganisms isolated from the coastal dunes of the Souss-Massa region. A two months period of salinity had a large negative impact on plant growth and acquisition of macro nutrients. However, the study underlined the role of the microbial inoculum for the plant in the achievement of salt tolerance. An isolate of Bradyrhizobium sp., RCM6 (R1), originating from the Massa dunes, was highly efficient in improving growth and nutrition of the A. cyanophylla. Double inoculation with the rhizobia and an endomycorrhizal complex, isolated from the Lamzar dunes had a clear additional positive effect, i.e. the fungi further increased the tolerance of the A. cyanophylla plants to salinity. This revised version was published online in June 2006 with corrections to the Cover Date.     

Plugging into the network: belowground connections between germlings and extraradical mycelium of arbuscular mycorrhizal fungi

Arbuscular mycorrhizal fungi (AMF) are obligate biotrophs; nevertheless their spores can germinate in the absence of host plants. Such inconsistent behavior is balanced by diverse survival strategies. The ability of AM fungal hyphae to fuse might represent a fundamental survival strategy because germlings could plug into compatible mycorrhizal networks, thus gaining access to plant-derived carbon before asymbiotic growth arrest. An in vivo experimental system was used to grow extraradical mycelium produced by Glomus mosseae colonizing three different plant species and germlings of the same isolate. After symbiotic and asymbiotic mycelia came into contact we showed that germling hyphae fused with symbiotic network hyphae and established protoplasm connections with nuclei occurring in fusion bridges. The frequency of anastomoses between germling and symbiotic hyphae was 4.9-23.9%. Prefusion and postfusion incompatible responses, with protoplasm withdrawal in interacting hyphae, were evident in some hyphal contacts. Given the multigenomic nature of AMF, the mingling of germling nuclei with those of the mycorrhizal network through perfect fusions might represent a means for the maintenance of genetic diversity in the absence of sexual recombination.     

At the Root of the Wood Wide Web: Self Recognition and Non-Self Incompatibility in Mycorrhizal Networks

Arbuscular mycorrhizal (AM) fungi are mutualistic symbionts living in the roots of 80% of land plant species, and developing extensive, below-ground extraradical hyphae fundamental for the uptake of soil nutrients and their transfer to host plants. Since AM fungi have a wide host range, they are able to colonize and interconnect contiguous plants by means of hyphae extending from one root system to another. Such hyphae may fuse due to the widespread occurrence of anastomoses, whose formation depends on a highly regulated mechanism of self recognition. Here, we examine evidences of self recognition and non-self incompatibility in hyphal networks formed by AM fungi and discuss recent results showing that the root systems of plants belonging to different species, genera and families may be connected by means of anastomosis formation between extraradical mycorrhizal networks, which can create indefinitely large numbers of belowground fungal linkages within plant communities.Key Words: arbuscular mycorrhizal symbiosis, extraradical mycelium, anastomosis, plant interconnectedness, self recognition, non-self incompatibility, mycorrhizal networks     

Laboratory and field methods for measurement of hyphal uptake of nutrients in soil

Experimental systems for measuring nutrient transport by arbuscular mycorrhizal (AM) fungi in soil are described. The systems generally include two soil compartments that are separated by fine nylon mesh. Both roots and root-external hyphae grow in one compartment, but only hyphae are fine enough to grow through the mesh into the other compartment. Application of tracer isotopes to the soil of this hyphal compartment can be used to measure nutrient uptake by plants via AM fungal hyphae. Use of compartmented systems is discussed with particular reference to phosphorus, which is the mineral nutrient transported in the largest quantity by AM fungi. Laboratory and field applications of the compartmentation methodology are presented with emphasis on the functioning of native AM fungal communities. Advantages and limitations of the method are considered and future important research directions are discussed in this context. This revised version was published online in June 2006 with corrections to the Cover Date.     

Plant infection and the establishment of fungal biotrophy

To exploit plants as living substrates, biotrophic fungi have evolved remarkable variations of their tubular cells, the hyphae. They form infection structures such as appressoria, penetration hyphae and infection hyphae to invade the plant with minimal damage to host cells. To establish compatibility with the host, controlled secretory activity and distinct interface layers appear to be essential. Colletotrichum species switch from initial biotrophic to necrotrophic growth and are amenable to mutant analysis and molecular studies. Obligate biotrophic rust fungi can form the most specialized hypha: the haustorium. Gene expression and immunocytological studies with rust fungi support the idea that the haustorium is a transfer apparatus for the long-term absorption of host nutrients.     

FACILITATION OF SUCCESSION BY THE NONMYCOTROPHIC COLONIZER SALSOLA KALI (CHENOPODIACEAE) ON A HARSH SITE: EFFECTS OF MYCORRHIZAL FUNGI

The effects of inoculation with vesicular-arbuscular mycorrhizal fungi on nonmycotrophic (plants which do not form a mycorrhizal association) colonizing annuals and on mycotrophic perennials were investigated in southwestern Wyoming. A subsoil containing no initial mycorrhizal inoculum was used, and seeds of the later successional perennials were planted. The annuals were removed from one-half of the sampling quadrats to test for the interaction of competition and mycorrhizae on the perennials. During the third and fourth years of succession, the density and percent cover of Salsola kali, the most abundant nonmycotrophic annual, decreased by one-half to one-third with inoculation. Mycorrhizal hyphae and spores were found in the rhizosphere of S. kali, with only occasional (1–2% of the root length) penetration of hyphae into the cortex. There were no differences in tissue phosphorus and nitrogen concentrations or water relations of inoculated vs. uninoculated S. kali. The planted grasses, all Agropyron species, had no significant increase in density or percent cover with inoculation. The percent root infection of A. smithii was 5–30%. Grass density and percent cover was greatest where S. kali was present, suggesting facilitation of grass establishment by annuals on this harsh, windy site. However, grass density was lower where 5. kali density was lower following inoculation. Mycorrhizal fungi were hypothesized to hasten the rate of succession on other sites. On this site where facilitation is an important process, inoculation of early seral plants and their subsequent decline may slow primary succession in the early years.     

Collimonas fungivorans, an unpredicted in vitro but efficient in vivo biocontrol agent for the suppression of tomato foot and root rot

Although bacteria from the genus Collimonas have demonstrated in vitro antifungal activity against many different fungi, they appeared inactive against the plant-pathogenic fungus Fusarium oxysporum f.sp. radicis-lycopersici (Forl), the causal agent of tomato foot and root rot (TFRR). Visualization studies using fluorescently labelled organisms showed that bacterial cells attached extensively to the fungal hyphae under nutrient-poor conditions but not in glucose-rich Armstrong medium. Collimonas fungivorans was shown to be as efficient in colonizing tomato root tips as the excellent colonizer Pseudomonas fluorescens strain WCS365. Furthermore, it appeared to colonize the same sites on the root as did the phytopathogenic fungus. Under greenhouse conditions in potting soil, C. fungivorans performed as well in biocontrol of TFRR as the well-established biocontrol strains P. fluorescens WCS365 and Pseudomonas chlororaphis PCL1391. Moreover, under biocontrol conditions, C. fungivorans did not attach to Forl hyphae colonizing plant roots. Based on these observations, we hypothesize that C. fungivorans mainly controls TFRR through a mechanism of competition for nutrients and niches rather than through its reported mycophagous properties, for which attachment of the bacteria to the fungal hyphae is assumed to be important.     

Mycorrhizae and soil phosphorus affect growth of <Emphasis Type="Italic">Celastrus orbiculatus</Emphasis>

Celastrus orbiculatus is an exotic liana that exploits disturbed areas in the eastern United States and once established, can invade into relatively undisturbed forest ecosystems. Mechanisms facilitating its invasion are not fully understood, including whether associations with mycorrhizal fungi are related to its invasion success. We grew C. orbiculatus in a greenhouse and compared growth responses when mycorrhizal fungi or a fungistatic were added to growth media that was either phosphorus-limiting or non-limiting. Results indicated C. orbiculatus forms association with native endomycorrhizal but not with native ectomycorrhizal fungi regardless of phosphorus level. Plants grown with sufficient phosphorus had significantly higher above-ground morphological and physiological traits but significantly lower root biomass compared to plants grown in low phosphorus conditions. Although above-ground traits did not vary significantly between mycorrhizal versus fungistatic added treatments, root biomass was significantly less in plants inoculated with mycorrhizae compared to plants receiving fungistatic. Under low phosphorus conditions, mycorrhizae appeared to be beneficial to the plant although being mycorrhizal did not fully compensate for insufficient phosphorous in the greenhouse situation where pot size limited soil exploration. Our results suggest that in the presence of mycorrhizae or sufficient phosphorus, C. orbiculatus can respond by preferentially allocating energy to above-ground growth, thus supporting its liana growth form onto trees and allowing the exotic to outcompete native species for light resources. If mycorrhizal fungi aid in acquisition of phosphorus, this association could be related to the invasion success of C. orbiculatus.     

Arbuscular mycorrhizal fungal communities change among three stages of primary sand dune succession but do not alter plant growth

Plant interactions with soil biota could have a significant impact on plant successional trajectory by benefiting plants in a particular successional stage over others. The influence of soil mutualists such as mycorrhizal fungi is thought to be an important feedback component, yet they have shown benefits to both early and late successional plants that could either retard or accelerate succession. Here we first determine if arbuscular mycorrhizal (AM) fungi differ among three stages of primary sand dune succession and then if they alter growth of plants from particular successional stages. We isolated AM fungal inoculum from early, intermediate or late stages of a primary dune succession and compared them using cloning and sequencing. We then grew eight plant species that dominate within each of these successional stages with each AM fungal inoculum. We measured fungal growth to assess potential AM functional differences and plant growth to determine if AM fungi positively or negatively affect plants. AM fungi isolated from early succession were more phylogenetically diverse relative to intermediate and late succession while late successional fungi consistently produced more soil hyphae and arbuscules. Despite these differences, inocula from different successional stages had similar effects on the growth of all plant species. Host plant biomass was not affected by mycorrhizal inoculation relative to un‐inoculated controls. Although mycorrhizal communities differ among primary dune successional stages and formed different fungal structures, these differences did not directly affect the growth of plants from different dune successional stages in our experiment and therefore may be less likely to directly contribute to plant succession in sand dunes.     

The diversity of ant-associated black yeasts: insights into a newly discovered world of symbiotic interactions

Based on pure culture studies and DNA phylogenetic analyses, black yeasts (Chaetothyriales, Ascomycota) are shown to be widely distributed and important components of numerous plant-ant-fungus networks, independently acquired by several ant lineages in the Old and New World. Data from ITS and LSU nu rDNA demonstrate that a high biodiversity of fungal species is involved. There are two common ant-fungus symbioses involving black yeasts: (1) on the carton walls of ant nests and galleries, and (2) the fungal mats growing within non-pathogenic naturally hollow structures (so-called domatia) provided by myrmecophytic plants as nesting space for ants (ant-plant symbiosis). Most carton- and domatia-inhabiting fungi stem from different phylogenetic lineages within Chaetothyriales, and almost all of the fungi isolated are still undescribed. Despite being closely related, carton and domatia fungi are shown to differ markedly in their morphology and ecology, indicating that they play different roles in these associations. The carton fungi appear to improve the stability of the carton, and several species are commonly observed to co-occur on the same carton. Carton fungi commonly have dark-walled monilioid hyphae, colouring the carton blackish and apparently preventing other fungi from invading the carton. Despite the simultaneous presence of usually several species of fungi, forming complex associations on the carton, little?overlap is observed between carton fungi from different ant species, even those that co-occur in nature, indicating at least some host specificity of fungi. Most fungi present on carton belong to Chaetothyriales, but in a few samples, Capnodiales are also an important component. Carton fungi are difficult to assign to anamorph genera, as most lack conidiation. The domatia fungi are more specific. In domatia, usually only one or two fungal species co-occur, producing a dense layer on living host plant tissue in domatia. They have hyaline or light brown thin-walled hyphae, and are commonly sporulating. In both carton and domatia, the fungal species seem to be specific to each ant-plant symbiosis. Representative examples of carton and domatia ant-fungus symbioses are illustrated. We discuss hypotheses on the ecological significance of the Chaetothyriales associated with ants.     

Availability and function of arbuscular mycorrhizal and ectomycorrhizal fungi during revegetation of dewatered reservoirs left after dam removal

Revegetation following dam removal projects may depend on recovery of arbuscular mycorrhizal (AM) and ectomycorrhizal (EM) fungal communities, which perform valuable ecosystem functions. This study assessed the availability and function of AM and EM fungi for plants colonizing dewatered reservoirs following a dam removal project on the Elwha River, Olympic Peninsula, Washington, United States. Availability was assessed via AM fungal spore density in soils and EM root tip colonization of Salix sitchensis (Sitka willow) in an observational field study. The effect of mycorrhizal fungi from 4 sources (reservoir soils, commercial inoculum, and 2 mature plant community soils) on growth and nutrient status of S. sitchensis was quantified in a greenhouse study. AM fungal spores and EM root tips were present in all field samples. In the greenhouse, plants receiving reservoir soil inoculum had only incipient mantle formation, while plants receiving inoculum from mature plant communities had fully formed EM root tips. EM formation corresponded with alleviation of phosphorus stress in plants (lower shoot nitrogen:phosphorus). Thus, revegetating plants have access to AM and EM fungi following dam removal, and EM formation may be especially important for plant P uptake in reservoir soils. However, availability of mycorrhizal fungi declines with distance from established plant communities. Furthermore, EM fungal communities in recently dewatered reservoirs may not be as effective at forming beneficial mycorrhizae as those from mature plant communities. Whole soil inoculum from mature plant communities may be important for the success of revegetating plants and recovery of mycorrhizal fungal communities.     

In vitro degradation of the moss Hylocomium splendens by three pleosporalean fungi

Three darkly pigmented species of conidial fungi of the family Pleosporaceae isolated from plants colonizing the Saskatchewan Glacier forefield were examined for potential roles in the degradation of moss gametophytes. Curvularia inaequalis and Ulocladium atrum isolated from bryophytes Ditrichum flexicaule and Tortella tortuosa , respectively, and Chalastospora gossypii from Saxifraga oppositifolia were inoculated onto autoclaved gametophytes of the moss Hylocomium splendens. All three species of fungi caused mass losses of the moss gametophytes. In vitro enzymatic tests revealed that all three fungi degraded cellulose, while none degraded insoluble polyphenols. When this material was examined by scanning electron microscopy, it was evident that the fungi had eroded the outer wall layer of the moss leaf cells to some extent but not the inner layer containing more lignin-like compounds. Once the outer wall layer was removed, the cells easily disarticulated. It is proposed that accumulations of these phenolics-rich leaf fragments subsequently ameliorate the rooting environment for vascular plants and have the potential to support the growth of basidiomycetes and other fungi, potentially mycorrhizal with pioneer vascular plants.     

High turnover of fungal hyphae in incubation experiments

Soil biological studies are often conducted on sieved soils without the presence of plants. However, soil fungi build delicate mycelial networks, often symbiotically associated with plant roots (mycorrhizal fungi). We hypothesized that as a result of sieving and incubating without plants, the total fungal biomass decreases. To test this, we conducted three incubation experiments. We expected total and arbuscular mycorrhizal (AM) fungal biomass to be higher in less fertilized soils than in fertilized soils, and thus to decrease more during incubation. Indeed, we found that fungal biomass decreased rapidly in the less fertilized soils. A shift towards thicker hyphae occurred, and the fraction of septate hyphae increased. However, analyses of phospholipid fatty acids (PLFAs) and neutral lipid fatty acids could not clarify which fungal groups were decreasing. We propose that in our soils, there was a fraction of fungal biomass that was sensitive to fertilization and disturbance (sieving, followed by incubation without plants) with a very high turnover (possibly composed of fine hyphae of AM and saprotrophic fungi), and a fraction that was much less vulnerable with a low turnover (composed of saprotrophic fungi and runner hyphae of AMF). Furthermore, PLFAs might not be as sensitive in detecting changes in fungal biomass as previously thought.     

Conidial anastomosis tubes in Colletotrichum

We describe the occurrence of special kinds of hyphae that create anastomoses directly between conidia. They can be found both in the laboratory and on infected plants. They first appear within asexual fruiting bodies approximately 15 days after conidiation has begun leading to the appearance of chains of connected conidia. Coincident with this we demonstrate in Colletotrichum lindemuthianum nuclear dynamics, including fragmentation, with cytoplasmic flow and passage of nuclei and organelles between conidia through the anastomosis tubes. We propose that conidial anastomosis tubes play an important role in the life cycle of these fungi.     

Gravisusception by buoyancy: a mechanism ubiquitous among fungi?

Gravitropism is ubiquitous among the fungal taxa; however, the mechanism(s) of gravisusception have overall remained obscure so far. In the vegetative sporangiophore of the zygomycete Phycomyces blakesleeanus some 200 large lipid globules form a conspicuous spherical complex which is positioned in a dense mesh of filamentous actin about 100 microm below the growing tip of the apex. Experimental suppression of that complex by transient growth at low temperature greatly diminishes the gravitropic response of the sporangiophore. With respect to size and abundance of the globules, the complex of lipid globules meets basic physical criteria for a possible function of gravisusception. Accumulations of similar lipid globules of critical size are documented in the apex of gravitropically growing hyphae of the endomycorrhizal fungus Gigaspora margarita (Glomeromycota) and have been described in the hyphal apices of members of various fungal phyla. We suppose that--in contrast to plants which use starch as a carbon storage and amyloplasts as statoliths--the fungi utilise the buoyancy of carbon-storing oil droplets for gravisusception.     

Mesophilic Actinomycetes in the Natural and Reconstructed Sand Dune Vegetation Zones of Fraser Island, Australia

The natural coastal habitat of Fraser Island located in the State of Queensland, Australia, has been disturbed in the past for mining of the mineral sand ilmenite. Currently, there is no information available on whether these past mining disturbances have affected the distribution, diversity, and survival of beneficial soil microorganisms in the sand dunes of the island. This in turn could deleteriously affect the success of the natural regeneration, plant growth, and establishment on the sand dunes. To support ongoing restoration efforts at sites like these mesophilic actinomycetes were isolated using conventional techniques, with particular emphasis on the taxa previously reported to produce plant-growth-promoting substances and providing support to mycorrhizal fungi, were studied at disturbed sites and compared with natural sites. In the natural sites, foredunes contained higher densities of micromonosporae replaced by increasing numbers of streptomycete species in the successional dune and finally leading to complex actinomycete communities in the mature hind dunes. Whereas in the disturbed zones affected by previous mining activities, which are currently being rehabilitated, no culturable actinomycete communities were detected. These findings suggest that the paucity of beneficial microflora in the rehabilitated sand dunes may be limiting the successful colonization by pioneer plant species. Failure to establish a cover of plant species would result in the mature hind dune plants being exposed to harsh salt and climatic conditions. This could exacerbate the incidence of wind erosion, resulting in the destabilization of well-defined and vegetated successional dunal zones.