Adaptation of the Spore Discharge Mechanism in the Basidiomycota

Background

Spore discharge in the majority of the 30,000 described species of Basidiomycota is powered by the rapid motion of a fluid droplet, called Buller''s drop, over the spore surface. In basidiomycete yeasts, and phytopathogenic rusts and smuts, spores are discharged directly into the airflow around the fungal colony. Maximum discharge distances of 1–2 mm have been reported for these fungi. In mushroom-forming species, however, spores are propelled over much shorter ranges. In gilled mushrooms, for example, discharge distances of <0.1 mm ensure that spores do not collide with opposing gill surfaces. The way in which the range of the mechanism is controlled has not been studied previously.

Methodology/Principal Findings

In this study, we report high-speed video analysis of spore discharge in selected basidiomycetes ranging from yeasts to wood-decay fungi with poroid fruiting bodies. Analysis of these video data and mathematical modeling show that discharge distance is determined by both spore size and the size of the Buller''s drop. Furthermore, because the size of Buller''s drop is controlled by spore shape, these experiments suggest that seemingly minor changes in spore morphology exert major effects upon discharge distance.

Conclusions/Significance

This biomechanical analysis of spore discharge mechanisms in mushroom-forming fungi and their relatives is the first of its kind and provides a novel view of the incredible variety of spore morphology that has been catalogued by traditional taxonomists for more than 200 years. Rather than representing non-selected variations in micromorphology, the new experiments show that changes in spore architecture have adaptive significance because they control the distance that the spores are shot through air. For this reason, evolutionary modifications to fruiting body architecture, including changes in gill separation and tube diameter in mushrooms, must be tightly linked to alterations in spore morphology.     

Descriptions and phylogenetic systematics of Myxobolus spp. from cyprinids in Algonquin Park, Ontario

Eight species of Myxobolus were collected from four species of cyprinids in Algonquin Park, Ontario. On the basis of spore morphology, five of these species are described as new and two are redescribed. The evolutionary relationships among these eight species were studied using partial small subunit ribosomal DNA (ssu-rDNA) sequence data. The resulting cladograms, which were highly resolved and with strongly supported relationships, allowed for the evaluation of spore morphology, host specificity, and tissue tropism, criteria traditionally used in species identification. These criteria, recently criticized for creating artificial rather than natural taxonomic groupings, were evaluated for their reliability in the systematics of the species examined. The data showed that distantly related species often infect the same host and tissue, and that closely related species often occur in different hosts. Morphologically similar species are more closely related to each other and the taxonomy based on spore morphology is consistent with the relationships depicted in the phylogenies. These results suggest that spore morphology is better than host specificity and tissue tropism as a species character, as well as for determining evolutionary relationships among the species of Myxobolus examined.     

A basic helix-loop-helix protein with similarity to the fungal morphological regulators, Phd1p, Efg1p and StuA, controls conidiation but not dimorphic growth in Penicillium marneffei

Members of the APSES protein group are basic helix-loop-helix (bHLH) proteins that regulate processes such as mating, asexual sporulation and dimorphic growth in fungi. Penicillium marneffei is a human pathogen and is the only member of its genus to display a dimorphic growth transition. At 25 degrees C, P. marneffei grows with a filamentous morphology and produces asexual spores from multicellular con-idiophores. At 37 degrees C, the filamentous morphology is replaced by yeast cells that reproduce by fission. We have cloned and characterized an APSES protein-encoding gene from P. marneffei that has a high degree of similarity to Aspergillus nidulans stuA. Deletion of stuA in P. marneffei showed that it is required for metula and phialide formation during conidiation but is not required for dimorphic growth. This suggests that APSES proteins may control processes that require budding (formation of the metulae and phialides, pseudohyphal growth in Saccharomyces cerevisiae and dimorphic growth in Candida albicans) but not those that require fission (dimorphic growth in P. marneffei). The A. nidulans DeltastuA mutant has defects in both conidiation and mating. The P. marneffei stuA gene was capable of complementing the conidiation defect but could only inefficiently complement the sexual defects of the A. nidulans mutant. This suggests that the P. marneffei gene, which comes from an asexual species, has diverged significantly from the A. nidulans gene with respect to sexual but not asexual development.     

Symbiotic status alters fungal eco-evolutionary offspring trajectories

Despite host-fungal symbiotic interactions being ubiquitous in all ecosystems, understanding how symbiosis has shaped the ecology and evolution of fungal spores that are involved in dispersal and colonization of their hosts has been ignored in life-history studies. We assembled a spore morphology database covering over 26,000 species of free-living to symbiotic fungi of plants, insects and humans and found more than eight orders of variation in spore size. Evolutionary transitions in symbiotic status correlated with shifts in spore size, but the strength of this effect varied widely among phyla. Symbiotic status explained more variation than climatic variables in the current distribution of spore sizes of plant-associated fungi at a global scale while the dispersal potential of their spores is more restricted compared to free-living fungi. Our work advances life-history theory by highlighting how the interaction between symbiosis and offspring morphology shapes the reproductive and dispersal strategies among living forms.     

Phylogeny of certain biocontrol agents with special reference to nematophagous fungi based on RAPd

A number of phylogenetic studies have been carried out on biocontrol agents having similar biological control activity. However, no work has been carried out to determine the phylogenetic relationship amongst various groups of biological control agents with varied biocontrol properties. Our aim was to derive a phylogenetic relationship between diverse biocontrol agents belonging to the deuteromycetes and determine its correlation with their spore morphology and their biocontrol activity. RAPD was used to assess genomic variability in fungi used as biological control agents which included ten isolates of nematophagous fungi such as Arthrobotrys sp., Duddingtonia sp., Paecilomyces sp. and Verticillium sp., along with two isolates of fungal biocontrol agents such as Trichoderma sp. and two isolates of entomopathogenic fungi including Beauveria sp. A plant pathogenic fungus, Verticillium alboatrum was also included to increase the diversity of Deuteromycetes used. A similarity matrix was created using Jaccard's similarity coefficient & clustering was done using unweighted pair group arithmetic mean method (UPGMA). The final dendogram was created using a combination of two programs, Freetree and TreeExplorer. The phylogenetic tree constructed from the RAPD data showed marked genetic variability among different strains of the same species. The spore morphologies of all these fungi were also studied. The phylogenetic pattern could be correlated with the conidial and conidiophore morphology, a criterion commonly used for the classification of fungi in general and Deuteromycetes in particular. Interestingly, the inferred phylogeny showed no significant grouping based on either their biological control properties or the trapping structures amongst the nematophagous fungi as reported earlier by other workers. The phylogenetic pattern was also similar to the tree obtained by comparing the 18S rRNA sequences from the database. The result clearly indicates that the classical method of classification of these deuteromycete members on the basis of their spore morphology is reliable and could be used for identification of these fungi at species level. The PCR fragment pattern polymorphism exhibited by the various species of a genus and different strains of a species indicates that construction of probes from one or more of these fragments will prove to be useful as a rapid tool for identification of species and strains of nematophagous fungi in future.     

Fungal spore swelling and germination are restricted by the macrophage phagolysosome

Many species of medically important fungi are prolific in the formation of asexual spores. Spores undergo a process of active swelling and cell wall remodelling before a germ tube is formed and filamentous growth ensues. Highly elongated germ tubes are known to be difficult to phagocytose and pose particular challenges for immune phagocytes. However, the significance of the earliest stages of spore germination during immune cell interactions has not been investigated and yet this is likely to be important for defence against sporogenous fungal pathogens. We show here that macrophages restrict the early phases of the spore germination process of Aspergillus fumigatus and Mucor circinelloides including the initial phase of spore swelling, spore germination and early polarised growth. Macrophages are therefore adept at retarding germination as well as subsequent vegetative growth which is likely to be critical for immune surveillance and protection against sporulating fungi.     

Characterizing and handling different kinds of AM fungal spores in the rhizosphere

Spores are important propagules as well as the most reliable species-distinguishing traits of arbuscular mycorrhizal (AM) fungi. During surveys of AM fungal communities, spore enumeration and spore identification are frequently conducted, but generally little attention is given to the age and viability of the spores. In this study, AM fungal spores in the rhizosphere were characterized as live or dead by vital staining and by performing a germination assay. A considerable proportion of the spores in the rhizosphere were dead despite their intact appearance. Furthermore, morphological and molecular analyses of spores to determine species identity revealed that both viable spores and dead spores with contents were identified. The accurate identification of spores at different developmental stages on the basis of morphology requires considerable experience. Our findings suggest that surveys of AM fungal communities based on spore enumeration and morphological and molecular identification are likely to be inaccurate, primarily because of the large proportion of dead spores in the rhizosphere. A viability check is recommended prior to spore molecular identification, and the use of trap cultures would give more reliable morphological identification results. We show that the abundance and activity of AM fungi in the rhizosphere can be determined by calculating the density of viable spores and the density of spores that could germinate. The adoption of these methods should provide a more reliable basis for further AM fungal community analysis.     

线虫内寄生真菌资源及生防应用研究进展

线虫内寄生真菌是一类能通过产生各种特殊孢子寄生游离线虫的食线虫真菌,对维持自然界线虫种群及其数量起到了关键作用,是线虫的重要天敌,也是研究物种多样性的重要材料,因此具有特殊的研究意义和经济价值.目前许多学者主要研究捕食线虫真菌对线虫的侵染力.这种食线虫真菌的侵染力主要针对二龄幼虫,而线虫内寄生真菌对线虫的各个时期都有侵染力.本文综述了线虫内寄生真菌物种资源现状,孢子形态多样性、侵染方式多样性、生活史多样性和寄主多样性,以及具有生防潜力和已商品化的生防菌的生防应用,探讨了在物种资源的发掘、物种分类鉴定和生防制剂商品化存在的问题及其对策,并展望了研究前景.     

Mycophagy in small mammals: A comparison of the occurrence and diversity of hypogeal fungi in the diet of the long-nosed potoroo Potorous tridactylus and the bush rat Rattus fuscipes from southwestern Victoria,Australia

Abstract Three broad dietary categories—fungus, plant and arthropod—were identified from faecal samples of two species of small terrestrial mammal in forest vegetation in southwestern Victoria. Fungal material formed the major component of the diet of the long-nosed potoroo Potorous tridactylus throughout the year and of the bush rat Rattus fuscipes during autumn and winter. Fungal material was most abundant for both species during autumn and winter and significantly less common in spring and summer. These results confirm previous studies which found P. tridactylus to be highly mycophagous throughout the year and R. fuscipes to be strongly mycophagous seasonally. Particular consideration was given to the composition of fungi in the diet. Fungal spores in faecal material were assigned to spore classes, which represent one or more fungal species that have similar spore morphology. Twenty-four fungal spore classes were recorded, but for both animal species most of the fungi consumed were from seven major spore classes. The proportions of major spore classes in the diet of both animals were generally similar, even though the composition of spore classes differed markedly across seasons. Minor differences between species in the fungi consumed may be related to differences in selectivity, foraging, or microhabitat use. If fungal resources are limiting, competition for such resources may be important in this and other small mammal communities. The amount and diversity of hypogeal fungi consumed by the two animal species makes them both important spore dispersal agents in forest ecosystems. The capacity of R. fuscipes and other seasonally mycophagous mammals in this role may be more important than previously recognized, especially in habitats where species of the Potoroidae are absent.     

Relationships and taxonomic status of Alternaria radicina, A. carotiincultae, and A. petroselini based upon morphological, biochemical, and molecular characteristics

Alternaria radicina, A. carotiincultae, and A. petroselini are closely related pathogens of umbelliferous crops. Relationships among these fungi were determined based on growth rate, spore morphology, cultural characteristics, toxin production, and host range. Random amplified polymorphic DNA (RAPD) analysis of these species, other species of Alternaria, and closely related fungi was also performed. A. petroselini was readily differentiated from A. radicina and A. carotiincultae on the basis of spore morphology, production of microsclerotia, host range, and RAPD analysis. Alternaria radicina and A. carotiincultae were considerably more similar to each other than to A. petroselini, but could be differentiated on the basis of growth rate, spore morphology, colony morphology, and, to a limited extent, RAPD analysis. When grown on media having a high nutritional content, A. radicina produced a diffusible yellow pigment and crystals of the fungal metabolite radicinin. In contrast, A. carotiincultae produced little or no radicinin. However, when A. carotiincultae was grown on the same medium amended with radicinin, growth rate and colony and conidial morphology were more similar to those of A. radicina. These results suggest that the morphological differences between A. radicina and A. carotiincultae are due, at least in part, to radicinin production, and that these fungi are conspecific. Therefore, we propose that A. carotiincultae be considered a synonym of A. radicina.     

Ultrastructural Studies on the Yeastlike and Mycelial Phases of Sporotrichum schenckii

Fine details of the internal and external morphology of the yeastlike and mycelial phases of the dimorphic fungal pathogen Sporotrichum schenckii as seen in ultrathin sections are described and illustrated by electronphotomicrography. Comparisons of yeastlike phase ultrastructure were made using two different methods of fixation and embedding. The internal morphology of the two forms of yeastlike S. schenckii was in many ways similar to that of similarly dimorphic fungi and yeasts studied by other authors. However, the use of the glutaraldehyde-osmium in agar fixation technique suggests the presence of an electron transparent capsular or slime layer with associated electron dense microfibrils to be present external to the cell wall of the yeastlike phase but not the mycelial phase. Mycelial phase S. schenckii was found to contain many of the internal microstructures reported for other filamentous dimorphic fungi. Conidia production in agitated liquid culture was found to be restricted since only rare sessile conidia were observed.     

The evolution of spore size in Agarics: do big mushrooms have big spores?

As a first attempt to investigate evolutionary patterns of spore size in Agarics, I tested whether this trait was correlated to the size of the fruit-body (basidiocarp). Based on phylogenetically independent contrasts, it was shown that big mushroom species had on average 9% longer, 9% wider and 33% more voluminous spores (all with P < 0.05, one-tailed tests) than small congeneric species (a three-fold difference in cap diameter was used to discriminate big and small mushrooms). It is argued that larger spore size does not consistently confer higher fitness in fungi, owing to aerodynamic constraints. Surprisingly, the cap–spore correlation was strongly lineage-specific. Thus, spore volume correlated significantly with cap diameter in five of 16 large genera (four positive and one negative correlation). Positive cap–spore correlations are interpreted in terms of developmental constraints, mediated by hyphal swelling during cap expansion. The possible mechanisms which can account for the breakdown of this constraint in the majority of genera investigated are discussed.     

An ultrastructural study of spore wall development and septal pores in species of the <Emphasis Type="Italic">Pachyphlodes</Emphasis> (Pezizaceae,Pezizales) lineage,with a description of the new species <Emphasis Type="Italic">Pachyphlodes annagardnerae</Emphasis>

Pachyphlodes (Pezizaceae) is a genus of truffle-like fungi that is distributed across the Northern Hemisphere. These fungi form ectomycorrhizae primarily with trees in the Fagaceae family, and occasionally with other host plants. The genus Plicariella (= Scabropezia) is phylogenetically inferred as an ally of, or within, the Pachyphlodes lineage. Despite molecular phylogenetic analyses that show the close relationships of species in these two genera, morphological differences in ascomata shape and color, spore ornamentation, and ascus shape are profound. Here, we studied spore wall development to better understand affinities within the Pachyphlodes–Plicariella lineages. Electron microscopy studies indicate that the initial spore wall development is similar across six Pachyphlodes species and a Plicariella species, despite striking differences in mature spore ornamentation among species. Ultrastructural analyses reveal that differences in spore ornamentation among Pachyphlodes species are due to unique developmental events at the final stages of spore wall deposition. Septal pore ultrastructure in Pachyphlodes species is similar to other Pezizaceae that have been studied. Molecular analyses of the five species studied indicate that four of them have not been previously described. The new species Pachyphlodes annagardnerae is here described, and the ultrastructural features of species of Pachyphlodes, Plicariella, and other Pezizales are compared and discussed.     

Small genets of Lactarius xanthogalactus, Russula cremoricolor and Amanita francheti in late-stage ectomycorrhizal successions

We determined the size of genets of late-stage ectomycorrhizal fungi in field sites in coastal Northern California. Basidiocarps were collected, mapped and subjected to genetic fingerprinting using amplified fragment length polymorphisms (AFLPs). The minimum size estimates for the largest genets of Amanita francheti, Lactarius xanthogalactus and Russula cremoricolor were 1.5, 9.3 and 1.1 m2, respectively. The molecular markers also showed that R. cremoricolor is dimorphic, with red- and white-capped morphotypes of this species forming a continuous population. Our results suggest that spore propagation plays a much more important role in the life history of the Russulaceae in undisturbed forest settings than previously recognized. Fungi appearing late in the succession sequence and systems without obvious disturbance therefore do not necessarily colonize primarily by mycelium.     

Calicioid lichens and fungi in amber – Tracing extant lineages back to the Paleogene

Calicioid lichens and fungi are a polyphyletic grouping of tiny ascomycetes that accumulate a persistent spore mass (mazaedium) on top of their usually well-stalked ascomata (‘mazaediate fungi’). In addition to extant forms, six fossils of the group were previously known from European Paleogene amber. Here we report nine new fossils and analyze the preserved features of all fossils to assess their applicability for dating molecular phylogenies. Many fossils are extremely well preserved, allowing detailed comparisons with modern taxa. SEM investigation reveals that even fine details of ascospore wall ultrastructure correspond to those seen in extant specimens. All fossils can confidently be assigned to modern genera: three to Calicium (Caliciaceae, Lecanoromycetes), five to Chaenotheca (Coniocybaceae, Coniocybomycetes), six to Chaenothecopsis (Mycocaliciaceae, Eurotiales), and one to Phaeocalicium (Mycocaliciaceae, Eurotiales). Several Calicium and Chaenotheca fossils are assignable to specific lineages within their genera, while the Chaenothecopsis fossils demonstrate the extent of intraspecific variation within one such lineage. Some features in the morphology of Chaenotheca succina nov. sp. seem to be ancestral as they have not been reported from modern species of the genus.     

Entomophthora species with E. muscae-like primary spores on two new insect orders, Coleoptera and Hymenoptera

Adults of Cantharis livlda (Coleoptera: Cantharidae) and of Torymus druparum (Hymenoptera: Torymidae) were found naturally infected by fungi from the Entomophthorales in Denmark. The morphology of the primary spores of the two fungi clearly showed that they belong to the genus Entomophthora s.str. No species from this genus has been reported so far from these insect orders. With respect to spore size and number of nuclei per spore, the fungi fall within the range of species from the E. muscae complex, known only from Diptera. A transfer of the fungus from T. druparum to Psila rosae (Diptera: Psilidae) was however possible. The findings thus confirm a significant widening of the host range of species within the E. muscae complex.     

祖先性状重建法揭示铁线蕨属植物孢子表面纹饰的形态多样性及其演化

蕨类植物孢子表面的纹饰形态复杂多样, 在分类学和古生物学研究中具有重要意义, 目前探索祖先性状演化常用形态学与系统发育学结合的方法。本文选取一回羽叶铁线蕨类植物为材料, 用扫描电子显微镜观察其孢子形态, 用5个叶绿体基因序列构建系统树, 用最大似然法和最大简约法来分析孢子性状的演化。结果表明: 一回羽叶铁线蕨类的孢子纹饰有5种类型, 即光滑、粗糙、颗粒、疣状及瘤状; 这一类群的祖先孢子纹饰有很大的可能是瘤状纹饰; 疣状纹饰可能是由瘤状纹饰演化而来; 孢子纹饰由简单向复杂演化, 越进化的物种其孢子纹饰越复杂。     

Morphogenesis of bacillus spore surfaces

Spores produced by bacilli are encased in a proteinaceous multilayered coat and, in some species (including Bacillus anthracis), further surrounded by a glycoprotein-containing exosporium. To characterize bacillus spore surface morphology and to identify proteins that direct formation of coat surface features, we used atomic-force microscopy (AFM) to image the surfaces of wild-type and mutant spores of Bacillus subtilis, as well as the spore surfaces of Bacillus cereus 569 and the Sterne strain of Bacillus anthracis. This analysis revealed that the coat surfaces in these strains are populated by a series of bumps ranging between 7 and 40 nm in diameter, depending on the species. Furthermore, a series of ridges encircled the spore, most of which were oriented along the long axis of the spore. The structures of these ridges differ sufficiently between species to permit species-specific identification. We propose that ridges are formed early in spore formation, when the spore volume likely decreases, and that when the spore swells during germination the ridges unfold. AFM analysis of a set of B. subtilis coat protein gene mutants revealed three coat proteins with roles in coat surface morphology: CotA, CotB, and CotE. Our data indicate novel roles for CotA and CotB in ridge pattern formation. Taken together, these results are consistent with the view that the coat is not inert. Rather, the coat is a dynamic structure that accommodates changes in spore volume.     

Spore ontogeny of the arbuscular mycorrhizal fungus<Emphasis Type="Italic"> Archaeospora trappei</Emphasis> (Ames &; Linderman) Morton &; Redecker (<Emphasis Type="Italic">Archaeosporaceae</Emphasis>)

Arbuscular mycorrhizal (AM) fungi in the genus Archaeospora (family Archaeosporaceae) contain both monomorphic and dimorphic species. The synanamorphism is often hard to discern without ontogenetic observations. Here, the spore ontogeny of Ar. trappei is reported from single species pot culture studies. The sporogenous hypha swelled up to a terminal sporiferous saccule and produced a lateral spore primordium on its neck. The saccule expanded fully before the spore primordium emerged. The saccule transferred its contents into the expanding spore and collapsed while wall differentiation continued inside the spore. The spore wall of Ar. trappei differentiated sequentially, in discrete steps, as in Acaulosporaceae members. In contrast, Ar. trappei produced a simplified spore wall in which the components differed in chemical and physical characteristics from those of the Acaulosporaceae members. Ontogenetic studies confirmed Ar. trappei to be monomorphic and producing acaulosporoid spores. The fungus is a new record to New Zealand.     

Identities and distributions of the co-invading ectomycorrhizal fungal symbionts of exotic pines in the Hawaiian Islands

Pine species have become invasive throughout the globe and threaten to replace native biota. The threat of pine invasion is particularly pressing in parts of the tropics where there are no native pines. The factors that govern pine invasion are not often well understood. However, key to pine survival is an obligate and mutualistic interaction with ectomycorrhizal fungi. Thus for pines to successfully invade new habitats compatible ectomycorrhizal fungi must already be present, or be co-introduced. The purpose of this study was to examine the community structure of non-native ectomycorrhizal fungi associated with pine invasions in the Hawaiian Islands. To accomplish this we executed a field and greenhouse study and used a molecular ecology approach to identify the fungi associating with invasive pines in Hawai‘i. We show that: (1) ectomycorrhizal fungal species richness in non-native pine plantations is far less than what is found in pine’s native range, (2) there was a significant decrease in average ectomycorrhizal fungal species richness as distance from pine plantations increased and, (3) Suillus species were the dominant fungi colonizing pines outside plantations. The keystone ectomycorrhizal fungal taxa responsible for pine establishment in Hawai‘i are within genera commonly associated with pine invasions throughout the globe. We surmise that these fungi share functional traits such as the ability for long-distance dispersal from plantations and host tree colonization via spore that lead to their success when introduced to new habitats.