A workflow for low-cost automated image analysis of myxomycete spore numbers, size and shape

Measuring spore size is a standard method for the description of fungal taxa, but in manual microscopic analyses the number of spores that can be measured and information on their morphological traits are typically limited. To overcome this weakness we present a method to analyze the size and shape of large numbers of spherical bodies, such as spores or pollen, by using inexpensive equipment. A spore suspension mounted on a slide is treated with a low-cost, high-vibration device to distribute spores uniformly in a single layer without overlap. Subsequently, 10,000 to 50,000 objects per slide are measured by automated image analysis. The workflow involves (1) slide preparation, (2) automated image acquisition by light microscopy, (3) filtering to separate high-density clusters, (4) image segmentation by applying a machine learning software, Waikato Environment for Knowledge Analysis (WEKA), and (5) statistical evaluation of the results. The technique produced consistent results and compared favorably with manual measurements in terms of precision. Moreover, measuring spore size distribution yields information not obtained by manual microscopic analyses, as shown for the myxomycete Physarum albescens. The exact size distribution of spores revealed irregularities in spore formation resulting from the influence of environmental conditions on spore maturation. A comparison of the spore size distribution within and between sporocarp colonies showed large environmental and likely genetic variation. In addition, the comparison identified specimens with spores roughly twice the normal size. The successful implementation of the presented method for analyzing myxomycete spores also suggests potential for other applications.

A new epiphytic species, Symphytocarpus macrosporus (Myxomycetes) from Western Siberia, Russia

A new species <em>Symphytocarpus macrosporus</em> is described based on collections made in the Khanty- Mansi Autonomous Area – Yugra, Russia. Thirty- two specimens of the new species were isolated from moist chambers with <em>Picea obovata</em> and <em>Abies siberica</em> bark. The new species is characterized by the presence of pseudoaethalia, without cortex, with peridium remaining as fragments. This new species clearly differs from previously described species of the genus by both morphological and molecular characters. It has large spores, (14)15–17(18) μm diam. with irregular ornamentation of large warts. Such a spore size seems to be the largest for the genus. The holotype specimen of <em>Symphytocarpus macrosporus</em> is stored in the M.G. Popov Herbarium (NSK), Novosibirsk, Russia. It is the first new species described within the genus <em>Symphytocarpus</em> since 1984.

Ascospore size declines with elevation in two tropical parmelioid lichens

Spore size and shape are biometric parameters frequently used in lichen taxonomy, especially in species characterization. However, the influence of environmental factors on the intraspecific variability of these characters remains very little investigated in lichenology. The elevational variation in spore length, width, volume and shape (length/ width ratio) of two species of the genus Hypotrachyna (H. aff. damaziana et H. altorum) occurring on Réunion Island (Indian Ocean) were studied. Spore length, width and volume significantly decrease with elevation in H. aff. damaziana, and spore width and volume also significantly decrease with elevation in H. altorum. There is no relation between spore shape and elevation in either of the two species. A significant correlation was further observed between the intra-individual variability in spore size of H. aff. damaziana and elevation. For this species, inter-individual variability in spore volume is also correlated with mean annual temperature and mean annual precipitation of the sampling locations, and spore width and length are correlated with mean annual temperature.

Fungal artillery of zombie flies: infectious spore dispersal using a soft water cannon

Dead sporulating female fly cadavers infected by the house fly-pathogenic fungus Entomophthora muscae are attractive to healthy male flies, which by their physical inspection may mechanically trigger spore release and by their movement create whirlwind airflows that covers them in infectious conidia. The fungal artillery of E. muscae protrudes outward from the fly cadaver, and consists of a plethora of micrometric stalks that each uses a liquid-based turgor pressure build-up to eject a jet of protoplasm and the initially attached spore. The biophysical processes that regulate the release and range of spores, however, are unknown. To study the physics of ejection, we design a biomimetic ‘soft cannon’ that consists of a millimetric elastomeric barrel filled with fluid and plugged with a projectile. We precisely control the maximum pressure leading up to the ejection, and study the cannon efficiency as a function of its geometry and wall elasticity. In particular, we predict that ejection velocity decreases with spore size. The calculated flight trajectories under aerodynamic drag predict that the minimum spore size required to traverse a quiescent layer of a few millimetres around the fly cadaver is approximately 10 µm. This corroborates with the natural size of E. muscae conidia (approx. 27 µm) being large enough to traverse the boundary layer but small enough (less than 40 µm) to be lifted by air currents. Based on this understanding, we show how the fungal spores are able to reach a new host.

Quantifying and interpreting nestedness of bryophytes in the Zhoushan Archipelago, China

Background: Detecting nestedness distribution and understanding their origin are important for biological conservation. Although previous studies on nestedness covered a wide range of taxa, discrepancies still exist on the prevalence of nestedness among biotas, and on the mechanisms to form nested distribution. Because bryophytes are poikilohydric, highly sensitive to habitats, and have a long-distance dispersal capacity, together with the fact that continental islands highly varied in area, elevation and habitat types, bryophytes on continental archipelago are thus an ideal system for the settlement of relevant disputes on nested distribution. However, few studies have been conducted on nestedness of bryophytes. To quantify nestedness level of different bryophyte categories on continental islands and possible mechanisms, we complied a presence/absence matrix of 446 species of bryophytes in 66 islands of the Zhoushan Archipelago, China. We also recorded spore sizes of 319 moss species based on available literature. By using the matrix temperature and the NODF metrics, as well as a conservative null model, we evaluated the nestedness levels of 18 bryophyte categories. We further explored possible mechanisms to form nested distribution by using partial Spearman rank correlations based on island area, elevation, habitat types and spore size of moss species. Results : We found that all 18 categories of bryophytes exhibited a high degree of nestedness. Nestedness level was higher for mosses than liverworts, higher for acrocarpous mosses than pleurocarpous mosses，and varied among different bryophyte 13 families. The nested distribution of bryophytes in the archipelago was not due to passive sampling, but mainly due to nested habitats and selective-extinction. Spore size might be a factor linked to selective-colonization accounting for nestedness of some moss categories. Conclusions : Nested habitats, selective-extinction, and partial selective-colonization resulted in a high level of nestedness for bryophytes in continental islands. Although the nestedness of bryophyte distribution in the Zhoushan Archipelago was statistically significant, such a nestedness was not perfect. Conservation of bryophytes should not only focus on the species-richest islands.