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Acacia mangium plantations account for more than 50 % of the exotic plantations in Vietnam. A new black butt symptom was discovered in 2012, followed by the wilting sign in Acacia seedlings in Tuyen Quang Province. Isolations recovered two Phytophthora species, the well-known Acacia pathogen P. cinnamomi, and an unknown species. The new species is described here as Phytophthora acaciivora sp. nov. Phylogenetically this species resides in clade 2d and is most closely related to P. frigida. Phytophthora acaciivora is a heterothallic species, oospores are aplerotic and antheridia are amphigynous. It produces predominantly elongated ovoid, semi papillate, persistent sporangia, no hyphal swellings and no chlamydospores. Optimum temperature for the growth is 25–30 °C and the maximum temperature is over 37.5 °C. Studies are underway to determine the impact of this new species on Acacia plantations in Vietnam.

A stem rot disease appeared on hydrangea plants in Tochigi Prefecture, Japan. Four Phytophthora-like isolates were obtained, and a representative isolate was selected for analysis. Hydrangea plants inoculated with the isolate developed similar symptoms. Molecular phylogenetic analyses using the rDNA ITS region and cox1, EF1-α, and β-tubulin genes indicated that the isolate was Phytophthora hedraiandra. The sporangia were papillate, laterally sessile, and caducous. Antheridia were paragynous or amphygynous, and laterally sessile, and the oospores were aplerotic. Based on these characteristics, the isolate was identified as P. hedraiandra. We suggest naming this hydrangea disease “Ajisai kukieki byo”.

In this study, Aphanomyces brasiliensis is proposed as a new species. It was collected from soil and water samples of a trout farming system located in Atlantic Rainforest areas, São Paulo State, Brazil. This species is mainly characterized by the presence of commonly short and unbranched oogonial stalks, antheridia usually androgynous and monoclinous with clavate, bell-shaped or cylindrical antheridial cells. We also collected Aphanomyces helicoides, A. raphani and A. stellatus from samples of submerged leaves, surface sediment and water in freshwater bodies. All species were isolated growing on cellulosic and/or keratinous substrates used as bait. Phylogenetic analysis of the ITS (rDNA) region showed that the new species and the other Brazilian isolates clustered within the Aphanomyces clade.

To test the premise that fertilization distances in bryophytes are limited by spermatozoid motility, we measured the distances between over 80 000 male and female thalli of Marchantia polymorpha on heathland over 2 successive years after a major fire and recorded the incidence of sporophytes. The first comparative data on spermatozoid numbers per antheridium across bryophytes and pteridophytes were calculated from antheridial sizes and spermatid diameters and cross‐checked with the likely numbers produced by successive antheridial mitoses. Individual antheridia of Marchantia produce over 200 000 spermatozoids. Extrapolating from individual antheridia to the numbers of mature antheridia per antheridiophore and then to individual male plants, we calculated that a single flooding event on a male thallus with 10–12 antheridiophores will result in the release of over 50 million spermatozoids. Assuming radial dispersal in surface water films by lipids released from the dehiscing antheridia, spermatozoids can reach distances exceeding 20 m from the parent plants, in line with our finding of 100% fertilization in female plants of Marchantia up to 19 m from the nearest males, far beyond published fertilization distances in dioicous bryophytes. We attribute this to (i) much greater spermatozoid production and numbers in Marchantia than in other bryophytes and (ii) highly effective sperm transport both within the antheridiophores and archegoniophores via their overlapping scales and grooves in the stalks and between male and female thalli via surface water films. These features, coupled with the massive production of small spores, explain the success of Marchantia as the primary colonist of open habitats.

Several populations of a partially submerged, robust, morphologically distinct Fossombronia were personally collected in central Mexico and several high elevation subpáramo communities in Venezuela. Comparative morphological studies of these populations with the type collections of F. peruviana, F. herzogii, F. ptychophylla, and F. australis, and a phylogenetic analysis of three chloroplast loci in 18 morphologically defined species of the genus were conducted to determine the taxonomic status of these populations. Both studies support the recognition of these populations as a new species of Fossombronia, which is named and described herein as Fossombronia delgadilloana. This new species resolves in a clade that is sister to F. peruviana, and is characterized by horizontal, planate, sub-longitudinally inserted leaves, widely dispersed antheridia, with basally expanded stalks, stipitate caulocalyces that lack lamellae, obovoid to ellipsoidal capsules, I-type inner capsule wall thickenings, and spores that are > 40µm, with reticulate distal wall ornamentation. Fossombronia herzogii is recognized as a new synonym of F. peruviana, and a new lectotype is designated for F. ptychophylla.

The impact of exogenous cytokinins (kinetin, zeatin, 6-benzylaminopurine, and N6-2-isopentenyladenine) on the pattern of Dryopteris filix-mas (L.) Schott spore germination, gametophyte growth and morphology in vitro have been studied. It was found that all studied cytokinins significantly retarded spore germination, inhibited gametophyte growth, caused deformations and decrease in the thallus size, and suppressed the development of reproductive structures and sporophyte growth at the concentration of 10–5 М. The reduction of the hormone concentration to 10–8 М stimulated the gametophyte development, induced cell divisions, particularly in the apical zone, due to which some of thalli were deformed, promoted the production of rhizoids, affected the formation of antheridia and archegonia, and slowed the sporophyte development.

During routine surveys of Pinus radiata plantations in the Nelson region, New Zealand, a Phytophthora species was isolated in association with bleeding stem cankers and rhizosphere soil. This isolate grew more slowly than other Phytophthora species associated with P. radiata in New Zealand, and was morphologically similar to isolates of Phytophthora cactorum previously associated with horticulture production, and isolates that were morphologically identified as P. cactorum from P. radiata in Nelson since the 1970s. Phylogenetic analyses of the ITS, cox1, and s-tubulin _F1A and s-tubulin_F2A regions confirmed this to be a new species closely related to P. hedraiandra in Clade 1. The new species is described here as Phytophthora aleatoria sp. nov. It produces partially caducous, papillate, ovoid to rarely globose or limoniform sporangia, markedly aplerotic oogonia forming thin-walled oospores, and paragynous (mainly) or amphigynous antheridia on some cultures. To date, this species has been confirmed on P. radiata in New Zealand across a range of sites within the North Canterbury, Nelson, Hawkes Bay, Taupo and the Bay of Plenty regions.

Cytogenetic studies in bryophytes have been limited by the difficulty of obtaining sufficient dividing nuclei and by the absence of modern protocols. The technical difficulties stem from the plants’ small size and lack of roots and pollen mother cells, the main sources of cells in division in vascular plants. In bryophytes instead, tiny sporophytes, antheridia, or phyllid meristems must be used to obtain meiotic or mitotic chromosome spreads. We here describe the preparation of such spreads from phyllids, antheridia, and sporophytes in several species of liverworts and compare available protocols with or without prefixation treatments. We also provide illustrated step-by-step instructions. The three prefixation agents (including colchicine) that we tested failed to improve synchronization of cell divisions. Young sporophytes were the best source of diploid synchronized cells, while antheridia were the best source of haploid cells. For meiotic nuclei, a short fixation of capsule tissue at the right developmental stage with 45% acetic acid sufficed to conserve the DNA for cytological investigations, while for mitotic nuclei, fixation in 3:1 ethanol/glacial acetic acid for a longer period (4–24 h) worked well.

espanolSe estudio la fase gametofitica de las especies Amblovenatum immersum y Christella subpubescens (Thelypteridaceae) incluyendo la germinacion, el desarrollo morfologico, caracteristicas vegetativas y estrategias reproductivas. El modelo de germinacion para ambas especies se ajusta al tipo Vittaria, mientras que el modelo de desarrollo del gametofito corresponde con el tipo Drynaria. Los gametofitos adultos eran cordados, con multitud de pelos unicelulares y secretores tanto por el margen, como por ambas caras del protalo. Ademas, en C. subpubescens se detectaron otro tipo de pelos aciculares, unicamente en el margen. Los gametangios presentaban el aspecto tipico de los helechos leptosporangiados. En A. immersum todos los gametofitos se desarrollaron como femeninos. En C. subpubescens los gametofitos primeramente son femeninos, con el paso del tiempo se vuelven bisexuales. Se detecto actividad de anteridiogeno en ambas especies, ya que despues del desarrollo de los gametofitos femeninos aparecieron en los alrededores protalos masculinos en fases muy tempranas de desarrollo (con ausencia de meristemo), pero sexualmente maduros. EnglishThe gametophytic generation of Amblovenatum immersum and Christella subpubescens (Thelypteridaceae), including spore germination, morphological development of the gametophytes, major vegetative features and reproduction strategies, was studied. For both species, the spore germination was of the Vittaria type and the developmental pattern was of the Drynaria type. Adult gametophytes were cordate and hairy, with unicellular and secretory hairs located in the margins and both the ventral and dorsal surfaces of the prothalli. C. subpubescens has another type of acicular hairs only in the margin of the prothallus. Gametangia were of the normal type described for leptosporangiate ferns. In A. immersum all the gametophytes were female. In C. subpubescens the gametophytes produced at first instance female gametangia and then became bisexual with time. Antheridiogen activity was observed in both species, suggested by the presence of small young ameristic gametophytes with antheridia surrounding well-developed female ones.

The gametophytes of the Athyriaceae are not very well known, such as Diplazium caudatum. The aim of this work is to analyze the gametophytic phase of this species, which includes germination study, morphological development and reproductive phase. Spores belonging to two different sporophytes from La Gomera, Spain, were mixed and sown in multisporic cultures. Plates have been cultured in chambers with nutritive agar at 25ºC and 12 hours photoperiod. The germination was registered every three days, and the main vegetative and reproductive development was checked throughout the observational period. The germination rate reached a maximum of 58%. The spore germination followed a Vittaria pattern meanwhile prothallial development followed an Adiantum type. Regarding sexual expression, all gametophytes developed archegonia and later some of them developed antheridia becoming bisexual. Los gametofitos de las Athyriaceae no son muy conocidos, tal es el caso de Diplazium caudatum. El objetivo de este trabajo es analizar la fase gametofítica de esta especie, que incluye el estudio de la germinación, desarrollo morfológico y fase reproductiva. Las esporas pertenecientes a dos esporofitos diferentes de La Gomera (España) han sido mezcladas y sembradas en placas de cultivo. Éstas se han cultivado en cámaras con agar nutritivo a 25ºC y 12 horas de fotoperíodo. La germinación se registró cada tres días y se comprobó el principal desarrollo vegetativo y reproductivo durante todo el período de observación. La tasa de germinación alcanzó un máximo del 58%. La germinación de las esporas siguió un patrón de tipo Vittaria, mientras que el desarrollo del prótalo siguió un patrón de tipo Adiantum. En cuanto a la expresión sexual, todos ellos desarrollaron arquegonios, y posteriormente una pequeña parte desarrolló anteridios convirtiéndose en bisexuales.

Two new species of Pythium Pringsheim isolated from soils of northwestern regions in Iran, are described: P. monoclinum Abrinbana, Abdollahz. & Badali, sp. nov., and P. iranense, sp. nov. Both species are morphologically distinct from all known species. Pythium monoclinum Abrinbana, Abdollahz. & Badali, sp. nov., is morphologically characterized by the presence of a swollen element in oogonial stalks below terminal oogonia, mostly monoclinous antheridia originating at various distances from the oogonia that occasionally arising from inflated part of oogonial stalk and, rarely production of peanut-shaped and double oospores. Pythium iranense, sp. nov., is distinguished from other species of the genus by the production of smooth-walled oogonia that occasionally have 1-3 papillae, globose to rarely elongated oogonia, 1-2(-5) antheridia per oogonium or crowd of antheridia around oogonia, 1-2 rarely 3-4 oospores per oogonium, rarely peanut-shaped oospores and immature oospores provided with 1-6 projections. Phylogenetic relationships of the isolates belonging to these new species with other related species were investigated using internal transcribed spacers of rRNA genes and partial cytochrome c oxidase subunit I sequence data. The phylogenetic analyses separated the two species from closely related species and placed P. iranense, sp. nov., in clade J1 but the clade of P. monoclinum Abrinbana, Abdollahz. & Badali, sp. nov., was unknown.

Main conclusionThis study shows differences in gene expression between male and female gametophytes of the simple thalloid liverwort with a distinction between the vegetative and reproductive phases of growth.AbstractPellia endiviifoliais a simple thalloid liverwort that, together with hornworts and mosses, represents the oldest living land plants. The limited taxon sampling for genomic and functional studies hampers our understanding of processes governing evolution of these plants. RNA sequencing represents an attractive way to elucidate the molecular mechanisms of non-model species development. In the present study, RNA-seq was used to profile the differences in gene expression betweenP. endiviifoliamale and female gametophytes, with a distinction between the vegetative and reproductive phases of growth. By comparison of the gene expression profiles from individuals producing sex organs with the remaining thalli types, we have determined a set of genes whose expression might be important for the development ofP. endiviifoliareproductive organs. The selected differentially expressed genes (DEGs) were categorized into five main pathways: metabolism, genetic information processing, environmental information processing, cellular processes, and organismal systems. A comparison of the obtained data with theMarchantia polymorphatranscriptome resulted in the identification of genes exhibiting a similar expression pattern during the reproductive phase of growth between members of the two distinct liverwort classes. The common expression profile of 87 selected genes suggests a common mechanism governing sex organ development in both liverwort species. The obtained RNA-seq results were confirmed by RT-qPCR for the DEGs with the highest differences in expression level. FivePellia-female-specific and twoPellia-male-specific DEGs showed enriched expression in archegonia and antheridia, respectively. The identified genes are promising candidates for functional studies of their involvement in liverwort sexual reproduction.

Pityrogramma calomelanos (L.) Link, popularly known as “Silver fern” has significant importance as a medicinal plant used traditionally for its astringent, analgesic, anti-haemorrhagic, anti-hypertensive, anti-pyretic and anthelminthic properties. This fern demonstrates an increased morphogenetic potential towards sporophyte formation, upon exposure to low doses of gamma radiation. Young sporophytic leaf crosier cultures were established in vitro on agar based Knop’s media with and without 20 g/l sucrose. The cultures were subjected to 60Co radiations in the range of 2.5–100 Gy. Apospory (production of gametophytes on sporophytic tissue without spores) was observed on leaf tissue cultured on Knops media with and without sucrose in P. calomelanos, at the end of 60 days. 5 Gy treated explants showed high number of aposporous gametophytes and was comparable to the control. Other tested doses reduced the aposporous gametophyte production significantly. In the second phase of the experimentation, the cultures were retained on the gametophyte induction media for a period of 4 weeks. Aposporous gametophytes were observed to proliferate with occasional development of antheridia. At the end of 4 weeks, morphogenetic development on the gametophytic tissue resulted in a significantly higher number of apogamous sporophytes (production of sporophytes without fusion of gametes) were obtained on 5 Gy treated tissue as compared to control and all the other treated explants. Apogamous sporophytes thus produced were successfully grown in the greenhouse and transferred to the field. Thus the use of gamma radiation in vitro not only reduced the need for sucrose for induction of apospory in P.calomelanos, it also exhibited hormesis at 5 Gy for improved sporophyte production.

A morphological comparison between Targionia hypophylla L. and T. stellaris Hassel is carried out based on fertile material from northern Argentina. The two species differed in a large number of gametophytic traits, including: epidermal pores, antheridia location and ventral scales shape. The spores of T. stellaris are also described in detail and compared to those of T. hypophylla; their potential taxonomic value is discussed in depth. The first photomicrographs of T. stellaris are provided.

During routine vegetation health surveys in the southwest of Western Australia (SWWA), several Phytophthora isolates with affinity to Clade 6a have been recovered. In this study, all known taxa from Clade 6a, P. inundata, P. humicola, P. gemini, P. ‘walnut’ and P. ‘personii’, and the new isolates were compared based on morphology and DNAsequence data from three nuclear genes and two mitochondrial genes resulting in the description of five new species, P. balyanboodja, P. condilina, P. cooljarloo, P. kwongonina and P. pseudorosacearum. With the exception of P. gemini and P. humicola, all species from Clade 6a have been recovered from natural ecosystems in SWWA. These species are morphologically similar, with predominantly ovoid sporangia and nested and extended internal proliferation. If oospores are present, they tend to be aplerotic with paragynous antheridia mostly attached adjacent to the oogonial stalk. They can all grow at 35 °C and have a fast growth rate on most agar media. These species have all been recovered from the rhizosphere soil and dead and dying plants within dry kwongon heathlands, often from water gaining sites and frequently from very isolated areas. The radiation, origin and potential ecological role of these species are discussed.

Root rot is a major constraint on field pea ( Pisum sativum L.) production. Species of Phytophthora can cause seed rot, pre- and post-emergence damping-off, and stem and root rot in many legume crops, but have not been reported in association with pea root rot in Canada. Following a survey of pea fields in Alberta, Canada, in 2015, root tissue with typical symptoms of root rot was plated onto a growth medium, and one isolate was identified as a putative Phytophthora spp. based on its colony characteristics. The isolate was homothallic, producing smooth, thick-walled oospores and both paragynous and amphigynous antheridia on lima bean agar. The sporangia were ovoid or elliptical, non-papillate, and had internal proliferations. Based on these criteria, the isolate was tentatively identified as Phytophthora sansomeana . Sequencing of the internal transcribed spacer (ITS) and Cox II regions revealed 100% identity with the ITS and Cox II sequences of P. sansomeana available in GenBank, confirming this identification. A pathogenicity test indicated that the isolate was pathogenic and caused pre-emergence damping-off, root discoloration, and root rot with brown to dark brown lesions on pea cultivars. This is the first report of P. sansomeana causing root rot in pea in North America.

During routine surveys of Corymbia calophylla, a widespread tree currently experiencing decline over a large area of its native range of south west Western Australia, a slow growing Phytophthora species was often recovered from roots and rhizosphere soil. This species grew more slowly, produced a wide array of morphological features and was visibly different to all other Phytophthora species present in Western Australia. Phylogenetic analyses of the ITS, cox1, HSP90, β-tubulin and NADH gene regions confirmed this to be a new species closely related to P. quercina and the provisional species P. ‘ohioensis’. The new species is described here as Phytophthora versiformis sp. nov. It produces persistent, papillate sporangia of variable shape, oogonia with thick-walled oospores, and paragynous antheridia. To date this species has only been recovered from C. calophylla. Pathogenicity trials indicate that while P. versiformis can infect the roots of C. calophylla it does not lead to seedling death.

Ślipiko, M., Sawicki, J. & Chmielewski, J. 2017. Ramet-based expressed sex ratio in Polish populations of Nyholmiella obtusifolia (Orthotrichaceae). — Herzogia 30: 51–57. We assessed ramet-based expressed sex ratio in eight Polish populations of Nyholmiella obtusifolia based on our own sampling (n = 4) and herbarium collections (n = 4). In each specimen of ca. 5 cm2 in size, we visually scored all individual ramets for the presence of sexual organs, i.e. antheridia and archegonia. We also counted gametophytes with sporophytes. Generally, most bisexual specimens (n = 5) were female-skewed with female ramets outnumbering males 7–10 fold. Only one population was almost balanced. Unisexual specimens (n = 3) were always female. Thirtyfive to 91 % of the ramets per specimen were non-expressing.

Reproductive performance is known to differ between co-sexual and non-co-sexual species. Thus, our aim was to determine whether: (i) the distance between sex structures is negatively associated with sex expression; (ii) male gametangia take longer to mature in rhizautoicous species than in gonioautoicous species; and (iii) the gonioautoicous sexual system has greater reproductive success (i.e. percentage of ramets with sporophyte) than the rhizautoicous sexual system. One population each of the mosses Fissidens scariosus and F. submarginatus, rhizautoicous and gonioautoicous, respectively, in a remnant of Atlantic Forest in Brazil were sampled monthly from September 2016 until August 2017. The number and phenophases of antheridia, archegonia and sporophytes were analysed for each species. Sexual expression and reproductive success were calculated, and reproductive phenology compared across environmental variables. As expected, sexual expression was significantly higher for the gonioautoicous species, which produced antheridia throughout the year and archegonia over many months, while gametangia production by the rhizautoicous species occurred only during the rainy season. Mean number of gametangia per perigonium and perichaetium were slightly higher for the rhizautoicous species (6.84 antheridia; 11.38 archegonia) than for the gonioautoicous species (4.39; 7.62). Gametangia and sporophyte production in the rhizautoicous species were markedly seasonal compared to that of the gonioautoicous species, although reproductive success did not differ. Therefore, we infer that the rhizautoicy (i.e. a functional dioicy) promotes lower expression of gametangia compared to gonioautoicy but is more efficient and so obtains the same reproductive success.

Introducción: Pteridium aquilinum es un helecho heliófilo ampliamente distribuido en México. Es una especie pionera que usualmente se encuentra en hábitats alterados y tiene gran relevancia ecológica por sus propiedades alelopáticas, su resistencia al fuego y a condiciones de sequía. Objetivo: realizar una descripción integral de la forma y ornamentación de las esporas de P. aquilinum, así como de la morfogénesis del gametofito, incluyendo el tipo de germinación y el desarrollo protálico (resaltando su morfología). Además, describir la anatomía del esporofito joven desarrollado en condiciones in vitro. Metodología: se utilizó microscopía electrónica de barrido y técnica histológica de parafina para la descripción de las fases del gametofito y esporofito. Resultados: Las esporas son globulares y el trilete tiene ornamentación granulada, mientras que su germinación fue de tipo Vittaria y el desarrollo protálico correspondió al tipo Adiantum. Los anteridios se observaron 13 días después de la siembra, mientras que los arquegonios surgieron al día 17. La primera hoja del esporofito apareció entre los días 60 y 70. Al cuarto mes, la fase esporofítica desarrollada in vitro mostró la anatomía típica de la especie a esta edad, dado que no presentó vitrificación. El análisis histológico del rizoma, la base del peciolo, el raquis de primer orden y la lámina mostraron los sistemas de tres tejidos bien diferenciados. Las modificaciones anatómicas observadas in vitro, como una dictiostela monocíclica y sólo una banda de esclerénquima en el rizoma, podrían atribuirse a la edad de los individuos. Por otra parte, los estomas están presentes en la superficie adaxial de la lámina, que corresponde al tipo anomocítico. Aunque estos estomas se formaron en condiciones in vitro, es importante resaltar que son completamente funcionales. Conclusiones: Nuestro trabajo describe por primera vez la morfo-anatomía de las fases gametofítica y esporofítica del ciclo de vida de P. aquilinum en condiciones in vitro. Nuestros resultados posiblemente permitan la exploración con mayor profundidad de las propiedades biológicas, fisiológicas y ecológicas de la especie.

BackgroundPost-embryonic growth of land plants originates from meristems. Genetic networks in meristems maintain the stem cells and direct acquisition of cell fates. WUSCHEL-RELATED HOMEOBOX (WOX) transcription factors involved in meristem networks have only been functionally characterized in two evolutionarily distant taxa, mosses and seed plants. This report characterizes aWOXgene in a fern, which is located phylogenetically between the two taxa.ResultsCrWOXBtranscripts were detected in proliferating tissues, including gametophyte and sporophyte meristems ofCeratopteris richardii. In addition,CrWOXBis expressed in archegonia but not the antheridia of gametophytes. Suppression ofCrWOXBexpression in wild-type RN3 plants by RNAi produced abnormal morphologies of gametophytes and sporophytes. The gametophytes of RNAi lines produced fewer cells, and fewer female gametes compared to wild-type. In the sporophyte generation, RNAi lines produced fewer leaves, pinnae, roots and lateral roots compared to wild-type sporophytes.ConclusionsOur results suggest thatCrWOXBfunctions to promote cell divisions and organ development in the gametophyte and sporophyte generations, respectively. CrWOXBis the first intermediate-clade WOX gene shown to function in both generations in land plants.

In 1995 loquat trees (Eriobotrya japonica [Thunb.] Lindl.) in several orchards in central Taiwan suffered from a severe stemrot and rootrot (2). Two populations of Phytophthora spp. isolates were recovered from diseased trees by placing pieces of plant tissue in a petri dish containing selective medium (2). One group of isolates possessed papillate sporangia, appeared to be heterothallic with amphigynous antheridia, and had a maximum growth temperature of 36°C. Upon mating, isolates of this group produced oogonia and oospores measuring 26.1 and 21.4 μm in diameter, respectively. These isolates were identified as typical strains of P. parasitica (=P. nicotianae) (2). The second group of isolates showed papillate sporangia, was homothallic with predominantly amphigynous antheridia, and had a maximum growth temperature of 34.5°C. However, the sizes of oogonia and oospores were considerably larger (average 34.7 and 30.3 μm in diameter, respectively) than those of P. nicotianae. Amplification of genomic DNA of one isolate (95023) of the second group by polymerase chain reaction (PCR) using P. nicotianae species specific primers PAR1 and PAR2, produced a special 1,000 bp sequence, indicating that the atypical isolates of second group were closely related to P. nicotianae. Isozyme analysis of two isolates (95023 and 95034, provided by L. L. Chern, Taiwan) of the second group, using the dimeric enzymes malic enzyme (MDHP; EC 1.1.1.40) and malate dehydrogenase (MDH; EC 1.1.1.37), was applied as described before (1,3). These two isolates generated the same three banded patterns using either enzyme (genotype: MDHP: 92/100; MDH-2: 93/100); the Mdhp92 allele and the Mdh-293 allele are diagnostic for P. cactorum, whereas the Mdhp100 allele and the Mdh-2100 allele are diagnostic for P. nicotianae. These well-defined heterozygous band patterns were exactly identical to those generated in atypical isolates which were identified earlier as hybrids of P. nicotianae and P. cactorum (1,3). Based on their atypical morphology and isozyme genotyping, it was concluded that these two isolates of the second group represent hybrids of P. nicotianae and P. cactorum. One of the parents, P. nicotianae, was also found closely associated with diseased loquat trees; the other parental species, P. cactorum, was not isolated from loquat in this study. It is the first time that natural hybrids of P. nicotianae and P. cactorum were found on loquat in Asia. References: (1) P. J. M. Bonants et al. Phytopathology 90:867–874, 2000. (2) L. L. Chern et al. Plant Dis. 82:651–656, 1998. (3) W. A. Man in 't Veld et al. Phytopathology 88:922–929, 1998.

Gametophytes of the fern Ceratopteris richardii develop into either hermaphrodites or males. As hermaphrodites develop, they secrete antheridiogen, or ACE, into the environment, inducing male development in undifferentiated gametophytes. Hermaphrodites are composed of archegonia, antheridia, rhizoids and a notch meristem, while males consist of antheridia and rhizoids. Much of the research on sexual and morphological development concerns gametophytes grown in sterile environments. Using biochemical and molecular techniques we identify a soil bacterium and explore its effects on sexual and rhizoid development. Hermaphrodite and male gametophytes were exposed to this bacterium and the effects on sexual development, rhizoid length and rhizoid number were explored. The bacterium was identified as a pseudomonad, Pseudomonas nitroreducens. Gametophytes grown in the presence of the pseudomonad were more likely to develop into hermaphrodites across all gametophyte densities. Across all gametophyte sizes, hermaphrodites had rhizoids that were 2.95× longer in the presence of the pseudomonad while males had rhizoids that were 2.72× longer in the presence of the pseudomonad. Both hermaphrodite and male gametophytes developed fewer rhizoids in the presence of the pseudomonad. Control hermaphrodites produced 1.23× more rhizoids across all gametophyte sizes. For male gametophytes grown in the absence of the pseudomonad, the rate of increase in the number of rhizoids was greater with increasing size in the control than the rate of increase in males grown in the presence of the pseudomonad. The pseudomonad may be acting on gametophyte sexual development via several potential mechanisms: degradation of ACE, changes in nutrient availability or phytohormone production. The pseudomonad may also increase rhizoid number through production of phytohormones or changes in nutrient availability.

Root rot of cocoyam (Xanthosoma sagittifolium L. Schott) caused by Pythium myriotylum Drechsler is a major disease of this crop in Africa (1,2) but is unreported from other regions of the world. During September 1999, commercially grown cocoyam (cv. Ratu-kiri-ala) in Gampaha (7°05′N, 80°00′E), Sri Lanka suffered from severe root rot. Initial symptoms were water-soaked lesions at the root tips that gradually enlarged to rot the entire root system and tuber. Wilting and yellowing of leaves were observed in advanced stages of disease. A Pythium sp. was regularly isolated from the affected roots and an isolate, SC5, was identified as P. myriotylum on the basis of morphology and the internal transcribed spacer (ITS) rDNA sequence. Characteristics of isolate SC5, grown on a grass-leaf water culture (3) were main hyphae up to 8.5 μm wide, oogonia terminal or intercalary (22.5 to 33.8 μm in diameter), antheridia diclinous occasionally monoclinous, one to eight per oogonium, stalks branched, often more or less loosely enveloping the oogonium, antheridium clavate or crook-necked, making apical contact with the oogonium, breadth of antheridium 2.5 to 7.0 μm, oospores aplerotic (17.0 to 22.5 μm in diameter), oospore wall 0.8 to 2.0 μm in thickness, sporangia terminal or intercalary, filamentous, inflated lobulate, and digitate, of variable length, breadth of sporangia 7.0 to 17.5 μm, formed in water; zoospores formed at 25°C, and diameter of encysted zoospores 10.0 to 12.5 μm. Cardinal temperatures on potato carrot agar 8°C minimum, 34°C optimum, and 37°C maximum with daily radial growth rate for 34°C at 32.8 mm. The ITS rDNA sequence of the isolate matched the sequences of P. myriotylum in GenBank (Accession Nos. AB095051 and AF452156) and isolate CBS254.70 used for the species description by van der Plaats-Niterink (3). The sequence of SC5 has been deposited in GenBank, Accession No. DQ102701. Pathogenicity tests used potted cocoyam plants (20 cm high), planted in an autoclaved potting mix. Four agar disks (8 mm in diameter) of isolate SC5 grown at 25°C for 48 h on potato dextrose agar was mashed and injected at a depth of 2 to 3 cm in the soil around the roots. Inoculated plants were placed in transparent plastic bags and kept for 7 days in a growth chamber maintained at 24 to 26°C with continuous light (52 to 98 μmol m-2·s-1). The experiment was carried out twice with three replications for each test. Dark brown rotting on roots and wilting of leaves were observed in 7 days after the inoculation. P. myriotylum was reisolated from diseased tissues and found to be morphologically identical to the original isolate SC5. Noninoculated control plants remained healthy. On the basis of the symptoms, morphological and molecular characteristics and confirmation of pathogenicity, P. myriotylum is the causal agent of root rot of cocoyam. To our knowledge, this is the first report of P. myriotylum causing root rot of cocoyam in Sri Lanka. References: (1) S. Nzietchueng. L'agronomie Tropicale 38:321, 1983. (2) R. P. Pacumbaba et al. J. Phytopathol. 135:265, 1992. (3) A. J. Van Der Plaats-Niterink. Stud. Mycol. 21:1, 1981.

During August 2005, wilted cucumber (Cucumis sativus cv. Tasty Green) plants were observed in a commercial greenhouse with a closed hydroponic system in the state of Mexico. Disease incidence was 50%. Diseased plants were detected 15 days after transplanting, when plants were overwatered. Yield was severely reduced when disease affected mature plants. Wilted plants showed basal stem lesions and root rot. Phytophthora capsici was consistently isolated from diseased tissue on corn meal agar (CMA) with tartaric acid. Oomycete identification was based on sporangial and gametangial characteristics (2). Sporangia produced on blocks of CMA at 25°C were spherical, broadly ellipsoid or obovoid with one papillae, and deciduous with a long pedicel (1). The isolates were heterothallic, and oogonia with amphigynous antheridia were observed in pairings with an A1 isolate of P. capsici, therefore, the isolates were determined to be an A2. Pathogenicity tests were conducted on 2-month-old cucumber seedlings under controlled conditions (25°C). Inoculation was performed by placing small pieces of agar with mycelium of 5- to 7-day-old cultures on the stem base and wrapping with Parafilm. Control plants were inoculated with CMA agar. No symptoms were observed on the control. Plants inoculated with the P. capsici isolated from the diseased cucumbers showed a basal stem lesion, followed by wilting and death 7 to 14 days after inoculation. The isolate was also pathogenic on tomato and eggplant that were grown at the same time in the commercial greenhouse sharing the nutrient solution. P. capsici sporangia were observed on the roots of both hosts. To our knowledge, this is the first report of P. capsici affecting cucumber in a hydroponics system in Mexico. References: (1) M. Aragaki and J. Y. Uchida. Mycologia 93:137, 2001. (2) D. C. Erwin and O. K. Ribeiro. Phytophthora Diseases Worldwide. The American Phytopathological Society. St. Paul MN, 1996.

Water survey for Phytophthora spp. by baiting with rhododendron leaves in April 2006 at a perennial ornamental plant nursery in Virginia detected five isolates showing a unique, previously unknown single-strand conformation polymorphism (SSCP) fingerprint (1). These cultures were isolated from two reservoirs at different depths of water column from surface to 2 m. They were homothallic and produced smooth-surfaced spherical oogonia with an average diameter of 27 μm on 10% V8 agar. Oospores were aplerotic. The paragynous antheridia were averaging 12 μm in diameter. Sporangia were papillate, spherical to ovoid, averaging 39 by 28 μm (length by width). They were caducous with short (2 mycelial plug from a 7-day-old culture and securing with Parafilm. V8 agar was used instead of mycelial plugs on control plants. After inoculation, each plant was enclosed in a plastic bag for 1 day and then incubated at 22°C with a 12-h photoperiod. Distilled water was sprayed daily for 5 days postinoculation (dpi) until disease symptoms were observed. At 15 dpi, 3 of the 10 inoculated rhododendron leaves and 6 of the 10 stems showed leaf lesions, wilting, dieback, and cankers, eventually leading to rhododendron death. Two of the 10 viburnum leaves and 4 of the 10 stems showed similar symptoms. Leaf lesions were approximately 3 to 5 cm in diameter. P. hedraiandra was recovered from diseased tissues and all resulting cultures showed an identical SSCP fingerprint to tested isolates as well as a P. hedraiandra isolate from Minnesota (3). No symptom developed on control plants. To our knowledge, this is the first report of P. hedraiandra in Virginia. Considering neither host plant has been grown or bought for resale by this nursery, this study indicates that P. hedraiandra may have a wider host range than is currently known. This possibility and the importance of water dispersal for P. hedraiandra in disease epidemiology warrant further investigation. References: (1) P. Kong et al. Fungal Genet. Biol. 39:238, 2003. (2) W. A. Man in't Veld et al. Eur. J. Plant Pathol. 117:25, 2007. (3) B. W. Schwingle et al. Plant Dis. 90:109, 2006.

High mortality was noticed in a 10-year-old stand of Pinus pinea in the Alberese area (Grosseto, central Italy, elev. 40 m, 42° 39′ 46″ N, 11° 06′ 25″ E) in July 2010. Aerial symptoms of trees included chlorosis, crown thinning, stunted growth, bark lesions at the stem base with resinous exudations, and extensive necroses of the underlying xylem tissue. Woody roots of two uprooted trees exhibited bark necroses and a high proportion of fine roots was destroyed. Soil around necrotic roots was baited using apple fruits (cv. Gala). After 1 week of incubation at 24°C, typical firm fruit rot developed and small tissue samples were transferred to clarified V8 agar (V8A) amended with 5 ml/l PARPNH and incubated at 24°C. After 7 days, stellate to rosaceous, finely lobed cottony colonies arose that were transferred to FPM medium and incubated at 24°C. Within 7 days, spherical oogonia with a smooth surface and predominantly paragynous antheridia formed; sporadic amphyginous antheridia could be observed. Colony squares (1 cm2) were then placed in filtered and sterilized pond water. After 48 h, ovoid, obpyriform, or clavate, nonpapillate, persistent sporangia with internal nested and extended proliferation were formed. Fifty oogonia and 30 sporangia were measured. The diameter of the 50 spherical oogonia varied from 33.6 to 44.9 μm (avg. 39 μm); dimensions of the 30 sporangia were 42.6 to 59.8 × 28.9 to 47.8 μm (avg. 52.95 × 38.98 μm; 1:b ratio 1.37). The isolate was identified as Phytophthora humicola W. H. Ko & Ann on the basis of colony type, size and morphology of oogonia and sporangia, average length/width ratio of sporangia, the homothallic formation of oogonia (4), and ITS rDNA sequence information (GenBank Accession No. JQ757060). A BLAST search of the ITS sequence of P. humicola isolate B33 revealed a 99% identity with the Phytophthora ITS Clade 6 species P. humicola and P. inundata (2). This latter species could be ruled out, however, since it is self-sterile, whereas our isolate B33 was self-fertile (3). A strain of P. humicola was deposited in the CBS-KNAW Fungal Biodiversity Centre, strain no. CBS129249. Pathogenicity tests were conducted on 10 one-year-old twigs of Pinus pinea. A bark portion was removed aseptically and a V8A disc (0.5 cm diam.) of P. humicola mycelium was placed on the wound. Control twigs (3) received sterile V8A discs. Inoculated and control twigs were incubated at 20°C in the dark. Clearly noticeable necrotic lesions (avg. length 2.2 × 0.68 cm) were observed after 15 days on inoculated twigs. Control twigs showed no symptoms. Reisolations on selective V8-PARPNH-agar confirmed P. humicola as the causal agent. P. humicola is mainly associated with woody horticultural crops (1, 3), while the other taxa grouped with this species in Clade 6 are mainly found in forest and riparian ecosystems (1). These aquatic Phytophthora species normally have a saprophytic lifestyle, but under favourable environmental circumstances can act as opportunistic pathogens, attacking susceptible trees and causing scattered mortality in forest stands and natural ecosystems (3). To our knowledge, this is the first report of P. humicola from a pine stand. It is supposed that the pathogen reached the stand through infected plant material or infested soil introduced into the stand. References: (1) C. M. Brasier et al. Mycol. Res. 107:277, 2003. (2) D. E. L. Cooke et al. Fungal Genet. Biol. 30:17, 2000. (3) T. Jung et al. Persoonia 26:13, 2011. (4) W. H. Ko and P. J. Ann. Mycologia 77:631, 1985.

We investigated the seasonality of the development of the gametangia and sporophytes of Racomitrium lanuginosum growing at a seasonally snow-covered site (ca. 2200 m altitude) on Mt. Fuji, Central Honshu, Japan. Shoots of R. lanuginosum were collected every 2 weeks during the snow-free period (June–November) in 2014. The number of inflorescences and the numbers, sizes, and developmental stages of the male and female gametangia and sporophytes were recorded. Archegonia developed quickly in early spring, but antheridia took longer to develop from the previous summer. Fertilization occurred in June and July and spore dispersal occurred in June of the following year. The archegonia took 1 month to mature, the antheridia took 7–10 months, and the sporophytes took 10 months. The development of the antheridia and sporophytes stopped during the winter when the study site was covered by snow.

International audience; Combresomyces cornifer is a conspicuous spheroidal microfossil (

Members of the P. citricola complex (Phytophthora clade 2c), such as P. plurivora, are destructive pathogens of trees and shrubs in nursery, landscape and forest setting worldwide. During surveys of Phytophthora species from streams and rivers in Massachusetts and North Carolina, a novel species in the P. citricola complex was recovered. Based on sequences from three nuclear (ITS, β-tub and tef1) and two mitochondrial (cox1 and nadh1) loci, morphological characters, temperature–growth relationships and host plant inoculations, this novel species is described as Phytophthora caryae sp. nov.. Phytophthora caryae resembles several other species in the P. citricola complex, demonstrating homothallism and producing paragynous antheridia and semipapillate and noncaducous sporangia. However, P. caryae exhibits smaller sexual structures, higher rates of oogonia with a tapered base and sporangia with an off-set attachment of the sporangiophores. Phylogenetic analyses using maximum likelihood and Bayesian inference placed isolates of P. caryae into a unique clade with significant statistical support. Based on the mitochondrial dataset, P. caryae is most closely related to P. pini and P. citricola III, which are believed to be native in eastern North America. Inoculations of P. caryae on one-year-old twigs of 12 tree species representing nine genera resulted in under-bark lesions on species of Carya and Juglans. Sapling inoculations under greenhouse conditions suggest that P. caryae may be pathogenic to shagbark hickory (C. ovata) but not to black walnut (J. nigra). This article is protected by copyright. All rights reserved.

In land plants, there are two types of male gametes: one is a non-motile sperm cell which is delivered to the egg cell by a pollen tube, and the other is a motile sperm cell with flagella. The molecular mechanism underlying the sexual reproduction with the egg and pollen-delivered sperm cell is well understood from studies using model plants such as Arabidopsis and rice. On the other hand, the sexual reproduction with motile sperm has remained poorly characterized, due to the lack of suitable models. Marchantia polymorpha L. is a model basal land plant with sexual reproduction involving an egg cell and bi-flagellated motile sperm. To understand the differentiation process of plant motile sperm, we analyzed the gene expression profile of developing antheridia of M. polymorpha. We performed RNA-sequencing experiments and compared transcript profiles of the male sexual organ (antheridiophore and antheridium contained therein), female sexual organ (archegoniophore) and a vegetative organ (thallus). Transcriptome analysis showed that the antheridium expresses nearly half of the protein-coding genes predicted in the genome, but it also has unique features. The antheridium transcriptome shares some common features with male gamete transcriptomes of angiosperms and animals, and homologs of genes involved in male gamete formation and function in angiosperms and animals were identified. In addition, we showed that some of them had distinct expression patterns in the spermatogenous tissue of developing antheridia. This study provides a transcriptional framework on which to study the molecular mechanism of plant motile sperm development in M. polymorpha as a model.

Dobbeler, P. 2017. Antheridia of Bryum capillare release moving spermatozoids after two months in the air-dry state. – Herzogia 30: 501–503. A specimen of the moss Bryum capillare collected in August 2004 in Tuscany (Italy) was preserved for two months at room temperature. The specimen was rehydrated and screened for the presence of fungi. It was observed that mature antheridia released moving spermatozoids. Antheridia and spermatozoids survived for 60 days in the air dry state.