Life Cycles of Myxogastria Stemonitopsis typhina and Stemonitis fusca on Agar Culture

Myxogastria is a group of protozoa characterized by cellular uninucleate amoeboflagellates (myxamoebae and flagellated swarm cell), acellular multinucleate plasmodia, and stationary spore‐bearing sporocarps. The Stemonitales is a large order in the Myxogastria and contains approximately 230 species, but only 13 species have their completed life cycles observed so far. Here, we described the life cycles of two species in Stemonitales, Stemonitopsis typhina and Stemonitis fusca by culturing in water agar medium and observing the morphogenesis of their spore germination, plasmodium, and sporocarp development. The spore‐to‐spore life cycles of Ste. typhina and S. fusca were completed in approximately 67 and 12 d, respectively. Both species possessed an aphanoplasmodium. However, the spores of Ste. typhina and S. fusca germinated by the V‐shape split and pore methods, respectively. Unlike S. fusca with an evanescent peridium, Ste. typhina produced a shiny persistent peridium which was continuous with the membrane surrounding its stalk. The information will contribute to a better understanding of their taxonomy and phylogeny.     

Spore-to-spore agar culture of the myxomycete Physarum globuliferum

The ontogeny of the myxomycete Physarum globuliferum was observed on corn meal agar and hanging drop cultures without adding sterile oat flakes, bacteria or other microorganisms. Its complete life cycle including spore germination, myxamoebae, swarm cells, plasmodial development, and maturity of fructifications was demonstrated. Details of spore-to-spore development are described and illustrated.     

STUDIES ON MARINE PLANKTON DIATOMS. II. RESTING SPORE MORPHOLOGY1

A morphological study of resting spores in five marine planktonic diatom species using electron microscopy indicates that Bacteriastrum delicatulum Cleve and Leptocylindrus danicus Cleve spores bear little resemblance, to their vegetative cells. Detonula confervacea (Cleve) Gran and Stephanopyxis turris (Grev. & Arn.) Ralfs spores have several features in common with their vegetative cells, and Rhizosolenia setigera Brightwell lies between the two extremes. The function of resting spores in relation to diatom life cycles is briefly discussed. Spore formation may be a primitive characteristic in the life cycle and may no longer have significant survival value for the species.     

Genomic divergence and lack of introgressive hybridization between two 13‐year periodical cicadas support life cycle switching in the face of climate change

Life history evolution spurred by post‐Pleistocene climatic change is hypothesized to be responsible for the present diversity in periodical cicadas (Magicicada), but the mechanism of life cycle change has been controversial. To understand the divergence process of 13‐year and 17‐year cicada life cycles, we studied genetic relationships between two synchronously emerging, parapatric 13‐year periodical cicada species in the Decim group, Magicicada tredecim and M. neotredecim. The latter was hypothesized to be of hybrid origin or to have switched from a 17‐year cycle via developmental plasticity. Phylogenetic analysis using restriction‐site‐associated DNA sequences for all Decim species and broods revealed that the 13‐year M. tredecim lineage is genomically distinct from 17‐year Magicicada septendecim but that 13‐year M. neotredecim is not. We detected no significant introgression between M. tredecim and M. neotredecim/M. septendecim thus refuting the hypothesis that M. neotredecim are products of hybridization between M. tredecim and M. septendecim. Further, we found that introgressive hybridization is very rare or absent in the contact zone between the two 13‐year species evidenced by segregation patterns in single nucleotide polymorphisms, mitochondrial lineage identity and head width and abdominal sternite colour phenotypes. Our study demonstrates that the two 13‐year Decim species are of independent origin and nearly completely reproductively isolated. Combining our data with increasing observations of occasional life cycle change in part of a cohort (e.g. 4‐year acceleration of emergence in 17‐year species), we suggest a pivotal role for developmental plasticity in Magicicada life cycle evolution.     

Optimization of AFLP for extremely large genomes over 70 Gb

Here, we present an improved amplified fragment length polymorphism (AFLP) protocol using restriction enzymes (AscI and SbfI) that recognize 8‐base pair sequences to provide alternative optimization suitable for species with a genome size over 70 Gb. This cost‐effective optimization massively reduces the number of amplified fragments using only +3 selective bases per primer during selective amplification. We demonstrate the effects of the number of fragments and genome size on the appearance of nonidentical comigrating fragments (size homoplasy), which has a negative impact on the informative value of AFLP genotypes. We also present various reaction conditions and their effects on reproducibility and the band intensity of the extremely large genome of Viscum album. The reproducibility of this octo‐cutter protocol was calculated using several species with genome sizes ranging from 1 Gb (Carex panicea) to 76 Gb (V. album). The improved protocol also succeeded in detecting high intraspecific variability in species with large genomes (V. album, Galanthus nivalis and Pinus pumila).     

MORPHOLOGY AND THERMAL DEATH POINT OF OLPIDIUM BRASSICAE

The morphology of single-sporangial isolates of lettuce, tomato, mustard, and oat Olpidium brassicae (Wor.) Dang. growing in their respective hosts as well as in cowpea were compared in situ and after extraction from the roots. The sporangia, zoospores, and resting spores of all isolates were within the established limits of the species. Single exit tubes or pores predominated which means that these isolates should not be transferred to the genus Pleotrachelus. A satisfactory assay for the presence of resting spores was developed by air-drying of the roots for a week or longer. This treatment killed zoospores and vegetative sporangia, but not resting spores. Factors affecting resting spore formation were investigated unsuccessfully. The thermal death point of zoospores of mustard isolates that did not form resting spores was between 40 and 45 C for 10 min.     

Differential Influence of Life Cycle on Growth and Toxin Production of three Pseudo‐nitzschia Species (Bacillariophyceae)

We used a multistrain approach to study the intra‐ and interspecific variability of the growth rates of three Pseudo‐nitzschia species – P. australis, P. fraudulenta, and P. pungens – and of their domoic acid (DA) production. We carried out mating and batch experiments to investigate the respective effects of strain age and cell size, and thus the influence of their life cycle on the physiology of these species. The cell size – life cycle relationship was characteristic of each species. The influence of age and cell size on the intraspecific variability of growth rates suggests that these characteristics should be considered cautiously for the strains used in physiological studies on Pseudo‐nitzschia species. The results from all three species do not support the hypothesis of a decrease in DA production with time since isolation from natural populations. In P. australis, the cellular DA content was rather a function of cell size. More particularly, cells at the gametangia stage of their life cycle contained up to six times more DA than smaller or larger cells incapable of sexual reproduction. These findings reveal a link between P. australis life cycle and cell toxicity. This suggest that life cycle dynamics in Pseudo‐nitzschia natural populations may influence bloom toxicity.     

Inoculum Density Relationships for Infection of Some Eastern US Forest Species by Phytophthora ramorum

Our objectives were to establish inoculum density relationships between P. ramorum and selected hosts using detached leaf and whole‐plant inoculations. Young plants and detached leaves of Quercus prinus (Chestnut oak), Q. rubra (Northern red oak), Acer rubrum (red maple), Kalmia latifolia (mountain laurel) and Rhododendron ‘Cunningham's White’ were dip‐inoculated with varying numbers of P. ramorum sporangia, and the total number of diseased and healthy leaves recorded following incubation at 20°C and 100% relative humidity. Calibration threshold estimates for obtaining 50% infected leaves based on linear analysis ranged from 36 to 750 sporangia/ml for the five hosts. Half‐life (LD50) estimates (the number of spores for which the per cent of diseased leaves reaches 50% of its total) from asymptotic regression analysis ranged from 94 to 319 sporangia/ml. Statistically significant differences (P = 0.0076) were observed among hosts in per cent infection in response to increased inoculum density. Inoculum threshold estimates based on studies with detached leaves were comparable to those obtained using whole plants. The results provide estimates of inoculum levels necessary to cause disease on these five P. ramorum hosts and will be useful in disease prediction and for development of pest risk assessments.     

小绒泡菌生活史及其形态建成期显微结构的观察

在完成小绒泡菌Physaum pusillum生活史的同时,观察了小绒泡菌P. pusillum生活史不同阶段显微结构的形成。生活史研究结果表明：小绒泡菌P. pusillum的孢子萌发方式为裂式,黏变形体具两根鞭毛,较短一根不易观察到,原质团类型为显型原质团,子实体形态建成过程约需12-13h。显微结构观察结果表明：石灰结和孢丝结构在原质团隆起约6h后开始形成,孢子在原质团隆起约9h后开始形成,石灰结和孢丝先于孢子形成。     

The Effect of Exposure to Decreasing Relative Humidity on the Viability of Phytophthora ramorum sporangia

Sporangia of three isolates of Phytophthora ramorum representing three different clonal lineages were subjected to relative humidity (RH) levels between 80 and 100% for exposure periods ranging from 1 to 24 h at 20°C in darkness. Plastic containers (21.5 × 14.5 × 5 cm) were used as humidity chambers with 130 ml of glycerine solution added to each container. Glycerine concentrations corresponded to 100, 95, 90, 85 and 80% RH based on refractive index measurements. Sporangia suspensions were pipeted onto nitrile mesh squares (1.5 × 1.5 cm, 15 micron pore size) which were placed in the humidity chambers and incubated at 20°C in darkness. Following exposure periods of 1, 2, 4, 8, 12 and 24 h, mesh squares were inverted onto Petri dishes of selective medium and sporangia germination assessed after 24 and 48 h. At 100% RH, we observed a mean value of 88% germination after 1 h exposure declining to 18% germination following 24 h incubation. At 95% RH, a steeper decline in germination was noted, with means ranging from 79% at 1 h to less than 1% at 24 h exposure. At 90% RH, no germination was noted after 8 or more h exposure, and values were 57%, 22% and 3% germination for the 1, 2 and 4 h exposures, respectively. Germination was only observed at 1 h exposure for both the 85% RH treatment (52% germination) and the 80% RH treatment (38% germination). The three isolates responded similarly over the range of RH values tested. The germination response of P. ramorum sporangia to RH values between 80% and 100% was comparable to that reported for other Phytophthora species. Knowledge of conditions that affect P. ramorum sporangia germination can shed light on pathogenesis and epidemic potential and lead to improved control recommendations.     

Corema album: unbiased dioecy in a competitive environment

• Corema album is a dioecious coastal shrub. Dioecious plants growing in these resource‐limited habitats may present spatial segregation of the sexes (SSS) or demographic biases because of the different reproductive effort between sexes. In these environments facilitation is a more common interaction between plants than competition. To assess factors determining the distribution of C. album male and female plants, we investigated the influence of habitat type (sand dunes and coastal woodlands), assessed the occurrence of SSS or demographic biases and also a possible role of these shrubs as nurse plants.
• We selected three C. album populations with the two habitat types. All C. album individuals were sexed, mapped and measured in three plots (20 m × 20 m) per population/habitat type. Presence and abundance of all plant species were recorded under five female and five male C. album plants as well as in equivalent open ground area in each of 15 plots.
• According to Ripley's K function result, C. album did not display SSS. Generalised linear mixed models (GLMM) show that differences in plant size were not related to plant sex. Plant inventory correspondence analysis showed that species composition and abundance were influenced by habitat type, population and the presence of a C. album individual, but not by its sex. GLMM indicated a detrimental effect of C. album on the co‐occurring plants.
• Our results show that sexual dimorphism has allowed C. album to adapt to the environment avoiding SSS or significant demographic bias, suggesting a positive outlook for its conservation.

     

The preparation,germination properties and stability of superdormant spores of Bacillus cereus

Aims: To determine yields, germination and stability of superdormant Bacillus cereus spores. Methods and Results: Superdormant B. cereus spores were isolated by germination with high concentrations of inosine or l ‐alanine in 2–5% yield and did not germinate with high concentrations of either of these germinants, but germinated like starting spores with Ca‐DPA, dodecylamine, l ‐alanine plus inosine or concentrated complete medium. Yields of superdormant spores from germinations with low inosine concentrations were higher, and these spores germinated poorly with low inosine, but relatively normally with high inosine. Yields of superdormant spores were also higher when nonheat‐activated spores were germinated. Superdormant spores stored at 4°C slowly recovered some germination capacity, but recovery was slowed significantly at ?20°C and ?80°C. Conclusions: Factors that influence levels of superdormant B. cereus spores and the properties of such spores are similar to those in B. megaterium and B. subtilis, suggesting there are common mechanisms involved in superdormancy of Bacillus spores. Significance: Superdormant spores are a major concern in the food industry, because the presence of such spores precludes decontamination strategies based on triggering spore germination followed by mild killing treatments. Studies of the properties of superdormant spores may suggest ways to eliminate them.     

Life cycle and morphology of Physarum pusillum (Myxomycetes) on agar culture

Myxomycetes (slime molds) are unique eukaryotic microorganisms with both characteristics of fungi and amoebae. Artificial cultures grown under controlled conditions were used to study the life cycle and morphogenesis. Physarum pusillum was collected from the field. Spores were inoculated and cultured with the hanging drop method. The complete life cycle was observed from spore to spore on agar without adding any solid nutrients or bacteria as food. Life cycle morphological characteristics were described for spore germination, myxamoebae, zygote, plasmodium and sporangia formation.     

Genome size stability across Eurasian Chenopodium species (Amaranthaceae)

Tremendous interspecific genome size variation is a well known phenomenon, whereas genome size within a species is supposed to be exceptionally stable and thus useful as a taxonomic trait. Using DAPI flow cytometry, we tested the stability of genome size in various representatives of Chenopodium s.s. (Amaranthaceae) across a broad geographical range (from Portugal to eastern Russia) in Eurasia. We sampled 1977 Chenopodium individuals of four different ploidies (di‐, tetra‐, hexa‐ and decaploids) from 347 populations. Intraspecific relative genome size variation was low, ranging from 2.0% in C. probstii to 7.7% in C. album, even in the species with broad distributions. We distinguished 12 homogeneous relative genome size groups among the 17 Chenopodium spp. tested. Genome size is useful for distinguishing certain morphologically similar groups of species such as C. suecicum/C. album, C. vulvaria/C. pamiricum–C. iljinii/C. sosnowskyi/C. karoi. Due to its genome size stability, the cosmopolitan species C. album can be used as an alternative internal standard in flow‐cytometric analyses with the additional advantages of annual life cycle, self‐compatibility and common occurrence all over the world. Finally, we did not detect any sign of hybridization between Chenopodium spp. of different ploidies.     

Temperature Effects on the Onset of Sporulation by Phytophthora ramorum on Rhododendron ‘Cunningham's White’

The effect of temperature and moist period on the onset of sporangia production by Phytophthora ramorum on Rhododendron ‘Cunningham's White’ was examined with misted detached leaves held in humid chambers. Following wound inoculation with sporangia, leaves were pre‐incubated at 20°C for either 24 or 72 h prior to placement at six different temperatures (4, 10, 15, 20, 25 and 30°C). The overall mean moist period required for first occurrence of sporulation over all six temperatures was 3.24 days with the 24‐h pre‐incubation time, compared with 1.49 days for the 72‐h pre‐incubation time. Following 24 h pre‐incubation at 20°C and at an incubation temperature of 15°C, sporangia were first collected from leaves following a 24 h incubation. At 10 and 20°C, sporangia were first collected after 48 h, whereas at 4, 25 and 30°C, sporangia were first collected after 3 days. Following 72 h pre‐incubation at 20°C, sporulation generally occurred within 1 day, even at temperatures such at 4 and 30°C that are suboptimal for sporulation. The highest levels of P. ramorum sporulation were observed at 20°C. P. ramorum formed sporangia on host tissue under moist conditions within the same time frame reported for P. phaseoli, P. palmivora and P. nicotianae, but substantially more slowly than certain other species such as P. infestans. Quantifying moisture and temperature conditions for initiation of sporangia production provides knowledge which leads to a greater understanding of the epidemic potential of P. ramorum.     

Exceptional preservation in Lower Devonian coalified fossils from the Welsh Borderland: a new genus based on reniform sporangia lacking thickened borders

Reniform sporangia, comprising two equal valves and containing retusoid spores, recovered from Lower Old Red Sandstone strata of Devonian age (micrornatus-newportensis Spore Biozone: lower Gedinnian lower Lochkovian) on North Brown Clee Hill in the Welsh Borderland are placed in Resilitheca salopensis gen. et sp. nov. Conventional compression fossils from Targrove, Ludlow of fertile axes showing isotomous branching with limited overtopping are considered conspecific because the terminal reniform sporangia contain the same spores. Spore ultrastructure is described using scanning and transmission electron microscopy. Sections show faint traces of lamellae. Particles associated with spores and sporangium wall are compared with the globules of pteridophytes and Ubisch bodies of angiosperms, and related to the development of the sporangium. The new plants are compared with Cooksonia caledonica Edwards known only from impressions, and with Renalia Gensel showing far more pronounced pseudomonopodial branching.     

Do differences in life‐history traits and the timing of peak mating activity between host‐associated populations of Chilo suppressalis have a genetic basis?

The development of host races, genetically distinct populations of the same species with different hosts, is considered to be the initial stage of ecological speciation. Ecological and biological differences consistent with host race formation have been reported between water‐oat and rice‐associated populations of Chilo suppressalis. In order to confirm whether these differences have a genetic basis, we conducted experiments to determine the extent to which various life‐history traits and the time of peak mating activity of these populations were influenced by the species of host plant larvae were raised on. Individuals from each population were reared for three consecutive generations on either water‐oat fruit pulp or rice seedlings. Descendants of both populations had higher larval survival rates, shorter larval developmental periods, higher pupal weight, and longer adult forewings, when reared on water‐oats than when reared on rice. The time of peak of mating activity differed between the descendants of each population, irrespective of whether they were raised on water‐oats or rice. These results indicate that although some life‐history traits of host‐associated populations of C. suppressalis are influenced by the host plant larvae are raised on, time of peak mating activity is not. Because it is a stable, objective, phenotypic trait, further research on difference in the time of peak mating activity between host‐associated populations of C. suppressalis should be conducted to clarify the mechanism responsible for host race formation in this species.     

Phytophthora austrocedrae sp. nov., a new species associated with Austrocedrus chilensis mortality in Patagonia (Argentina)

Phytophthora austrocedrae is a new species isolated from necrotic lesions of stem and roots of Austrocedrus chilensis. It is a homothallic species characterized by semipapillate sporangia, oogonia with amphigynous antheridia, and very slow growth (1–2 mm d⁻¹ on V-8 agar at 17.5 °C optimum temperature). Phylogenetic analysis of ITS rDNA sequence indicates that its closest relative is Phytophthora syringae, another species frequently isolated from soil and streams in A. chilensis forests.