Mature Ascospores Research Articles

Phylogeny of the Pyrenula ochraceoflava group (Pyrenulaceae) reveals near-cryptic diversification and the inclusion of the Mazaediothecium album aggregate

In this study we present an analysis of the Pyrenula ochraceoflava group (Pyrenulaceae), focusing on the Neotropics and based on morphological, chemical, and molecular data of the mtSSU, nuLSU and ITS markers. We described three new species from tropical dry forests of Mexico, confirm the monophyly of the P. ochraceoflava group and provide evidence for the inclusion of species currently placed in the genus Mazaediothecium within Pyrenula. Pyrenula aurantiacoretis sp. nov. is characterized by an orange pigment covering the thallus in net-like fashion, muriform ascospores with 4 rows of 1–4 cells each, 12–15.5 × 8–10.5 µm, and 7-chloroemodin and emodin as major compounds. Pyrenula connexa sp. nov. is closely related to Mazaediothecium album, being characterized by mazaedioid pyrenocarps, basal and lateral excipular carbonization, highly variable mature ascospores, 1-septate to submuriform, thallus with abundant white verrucae, and lichexanthone as major compound. Pyrenula moldenkeorum sp. nov. is characterized by an orange thallus, submuriform ascospores that frequently show pigmented septa forming a cross septation pattern, 7.5–11 × 5.5–8.5 µm in size, and 7-chloroemodin and emodin as major compounds. The taxonomy of the most common and widespread species of the group, P. ochraceoflava and P. ochraceoflavens, is briefly discussed, presenting evidence to support the consideration of P. ochraceoflava as a species complex. The two species Mazaedothecium album and M. mohamedii are transferred to Pyrenula as P. aptrootiana nom. nov. [non Pyrenula alba (Schrad.) A.Massal.] and P. mohamedii comb. nov.

A novel fossil-species of Meliolinites Selkirk (fossil Meliolaceae) and its life cycle stages associated with an angiosperm fossil leaf from the Siwalik (Mio-Pliocene) of Bhutan sub-Himalaya

Here, we report the in-situ occurrence of a new fossil-species of Meliolinites (fossil Meliolaceae), Meliolinites bhutanensis sp. nov. on the cuticle fragments of a compressed angiosperm dicot leaf recovered from the middle Siwalik (Formation II: latest Miocene to Pliocene) of Bhutan, eastern Himalaya. This unique foliicolous new fossil fungal species features well-preserved mycelia consisting of superficial, brown to dark brown, septate, thick-walled, branching hyphae with bi-cellular appressoria, unicellular phialides, and a characteristic long, slightly curved hyphal seta. The web-like, brown to dark brown fungal colonies also include globose to sub-globose, dark brown ascomata, and oblong to broadly cylindrical, 5-celled, 4-septate, brown to dark brown, mature ascospores. As almost all features of different stages in the life cycle (ascospores, mature germinating ascospores, superficial lateral hyphae, hyphal seta, hyphopodia, mycelial colony, and ascomata) of this new fossil-species are found, we have proposed the first time a possible life cycle of fossil-species of Meliolaceae. The in-situ evidence of M. bhutanensis on the host leaf cuticle indicates the possible existence of a host-ectoparasite relationship in Bhutan sub-Himalaya's ancient warm and humid tropical evergreen forest during the deposition. So, M. bhutanensis might have thrived generally under warm and humid climate conditions for its growth and development in the Mio-Pliocene time, which is in conformity with our recently published quantitative climatic data by CLAMP (Climate Leaf Analysis Multivariate Program) analysis.

First Report of Microdochium nivale and M. majus Causing Brown Foot Rot of Wheat in China.

Microdochium nivale and M. majus not only cause seedling blight of wheat (Triticum aestivum L.) in cold dry soils, but also cause foot rot and ear blight of wheat under favorable conditions (Haigh et al. 2009). In May 2017, 2019, and 2020, a serious foot rot of wheat with an incidence of 92%, 45%, and 51% was observed in the field in Xiangcheng County (33.43° N, 114.84° E), Tanghe County (32.43° N, 112.66° E), and Linzhou City (36.13° N, 113.75° E), Henan Province, respectively. The serious brown lesions of the lower leaf sheaths is visible. The pathogens were isolated from brown leaf sheaths on potato dextrose agar (PDA) after being surface-sterilized (70% EtOH for 30 s followed by 3% NaClO for 1.5 min) and rinsed three times in sterile distilled water. After 5 d, mycelia were transferred to fresh PDA, and nine representative isolates (G17ZK2-1, G17ZK2-2, G17ZK2-3, g19TH10-4, g19TH10-5, g19TH10-6, G20LZ1-6, G20LZ1-7, and G20LZ1-8) were further purified by hyphal tipping. Species were identified based on morphological characteristics, and sequence analysis of partial sequences of the translation elongation factor-1α (TEF), the RNA polymerase II subunit (RPB2) gene and β-tubulin gene (Abdelhalim et al. 2020). Among the nine isolates, six isolates belonged to M. majus, three isolates belonged to M. nivale. Sequences of six isolates M. majus and three isolates M. nivale were deposited in GenBank with accession numbers MW428296-MW428298, MZ734119-MZ734121and MZ734139-MZ734141(TEF), MW384889, MW428291, MW428292, MZ734203-MZ734205 and MZ734161-MZ734163(RPB2), MW428293-MW428295, MZ501004-MZ501006 and MZ501024-MZ501026 (β-tubulin). For all the genes, isolates revealed 98-100% similarity to M. majus and M. nivale accessions, respectively. Microscopy of the six M. majus isolates showed: the conidia were falcate, straight to curved, apex pointed or obtuse to subacute, lacking basal differentiation, with 1 to 6 septa, 3.6 to 5.0 × 15.0 to 30.5 μm (av.= 4.5 × 23.2; n = 60). The three M. nivale isolates showed: the conidia were hyaline, 1 to 3 septa, 2.4 to 4.4 × 11.9 to 26.0 μm (av.= 3.5 × 14.7; n = 60). Perithecia of M. majus are dark brown, globose, and 95.2 to 190.5 × 95.2 to 228.6 μm (av.= 144.4 ×152.5; n = 30). Asci are clavate, and 6.8 to 11.0 ×68.2 to 77.3 μm (av.= 8.6×72.0; n = 30), contain eight ascospores. Mature ascospores are ellipsoidal, and 3.8 to 4.9 ×11.5 to 19.2 μm (av.= 4.0 ×15.2; n = 30), with 1 to 3 septa. These morphological characteristics were consistent with previous descriptions of these two species (Glynn et al. 2005). For pathogenicity tests, mycelia of M. nivale and M. majus was prepared using the modified procedure of Zhang et al. (2015). Two-week-old healthy wheat seedlings (cv. Aikang 58) were inoculated using 1 mL of prepared mycelia to one seedling, which was sprayied on soil. Control seedlings were inoculated with 1 mL distilled water containing 0.2% gelatin. After 10 days under 15/10℃, 16h/8h, all the inoculated plants had developed brown spots; while control plants remained healthy. The pathogens were reisolated from inoculated plants and identified as M. nivale and M. majus based on morphological characteristics and molecular methods described above. Although there are reports of M. majus associated with brown foot rot of wheat in Anhui Province and M. nivale associated with seedling blight of oat in Gansu Province (Chen et al. 2021; Tai et al. 2019). To our knowledge, this is the first report of brown foot rot of wheat caused by M. nivale and M. majus in China.

First report of Chaetomium pseudocochliodes causing flower rot disease of Camellia reticulata in Yunnan, China.

Camellia reticulata is the world-famous ornamental flower (Wang et al. 2021). In February 2021, the infected flowers of C. reticulata 'Shizitou' were collected in Zixi Mountain, Chuxiong city, Yunnan province, China (24°9'95″ N, 101°42'53″ E). Flower rot disease incidence ranged from 40% to 75% in the garden. The infected flowers showed symptoms of varying degrees of yellow-browning, dry or wet rot to the whole flower wilted and even dropped (Figure 1A, B, C). Three symptomatic flowers were randomly collected in the garden. Tissues from the infected flowers (cut to 5×5 mm size) were surface-disinfected by 75% ethyl alcohol for 30s, rinsed in sterile water for 3 times to air dry, and cultured in Potato Dextrose Agar medium (PDA) at 25℃±2 in the normal light for 5-7 days (Fang, 1998). Similar fungal colonies were isolated from 50%-75% of the infected flowers. Three isolates from different flowers showed similar colony morphology. After sub-culturing of hyphal tips on PDA for 5-7 days, the initially yellow colored colonies showed abundant white aerial mycelium, with sporulation (Figure 1E, F). The asci (Figure 1G) sporulation site is 24(-37) ×7(-14) μm, and the stalk length is 17-42 μm, with 8 biseriate acuminate ascospores. The mature ascospores (Figure 1H) are olive-brown or brown, lemon-like, double-pointed, with slightly umbilical protrusions at both ends, flat on both sides, 9(-11.5) × 7(-9) × 5.5(-7) μm in size, with germ holes on the top (Wang et al. 2016). These morphological features are consistent with Chaetomium pseudocochliodes. The genomic DNA was extracted from the isolated strains. To identified this fungal pathogen genetically, sequence analyses were conducted using the ITS1/ITS4 (Henson et al. 1993), NL1/NL4 (Liu et al. 2011), EF1-938F /EF1-2218R (Tan et al. 2016) primers for the internal transcribed spacer (ITS), large ribosomal subunit (LSU), and elongation factor 1-α (EF1-α) genes. The obtained sequences were deposited in GenBank with accession numbers MZ817067 (ITS), MZ817072 (LSU), MZ820167 (EF1-α). The phylogenetic trees (Figure 2) were constructed to determine the phylogenetic relationships based on MEGA 6.0 maximum likelihood method. In order to confirm the pathogenicity, the tests were conducted with fungus plug (5 mm) from a 7-day-old colony placed onto the surface of healthy petals. The sterile water-absorbent cottons place onto healthy petal surface near fungus plug and plastic wraps cover in petri dish were used for moisturizing. A total of 3 replicates in each of 3 groups were included (3 healthy petals for a group, 1 for wounded inoculation, 1 for unwounded inoculation, and 1 for sterile PDA plug). A sterile PDA plug was placed onto the surface of healthy petals as a control. After incubation at room temperature for 5 days (Ren. 2019), 3 replicates in each of 2 groups of treated healthy petals for wounded inoculation showed obvious symptoms (Figure 1D), and no symptoms appeared in the control, and healthy petals unwounded showed symptoms for 7-10 days. The fungus was re-isolated from the lesions of inoculated tissues. The re-isolated fungal colonies showed identical morphology and high sequence similarity with ITS, LSU and EF-1α of the initial isolate. No fungus was isolated from the controls. The first extraction of C. pseudocochliodes was also obtained from the roots of the Panax notoginseng in Wenshan, Yunnan (Wang et al. 2016). To our knowledge, this is the first report of flower rot caused by C. pseudocochliodes on C. reticulata in China.

Methods for observing, culturing, and studying living ascospores

Ascospore morphologies provide important characters with which to diagnose and describe taxa in Ascomycota. Ascospore features such as size, shape, color, septation, wall thickness, and guttulation, among others, are provided in identification manuals and descriptions of new species. Yet, by tradition, ascospores are usually described from dead fungarium material, and unfortunately, occasionally from immature or overmature ones. However, living, mature ascospores display a wealth of taxonomically informative morphological features that are lost or obscured when they die. Examples of the severe morphological changes that ascospores undergo when they die are provided here. Data from living ascospores may not be observed and recorded by mycologists because field and laboratory practices do not prioritize the study of freshly collected specimens. In this review, we discuss how to assess ascospore maturity and describe methods to produce an ascospore deposit for the purpose of obtaining living, mature ascospores. Ascospores are ejected from living, mature asci onto a cover glass or growth medium. The ascospores collected on these surfaces can be used in microscopy and culture studies. Notes on a method for isolating conidia on growth medium are also provided. This guide is aimed at those who have a basic understanding of ascomycetes, including the various types of ascomata and mechanisms of ascospore liberation. Methods given in this paper are primarily applied to ascomycete fungi that have active ascospore discharge. Some methods may be adapted for use with other groups that have passive discharge. Our purpose is to promote standardized, accurate, and thorough morphological characterization of living ascospores, as well as to encourage the routine employment of culture-based methods.

Pathogenic Fitness of Pyrenophora tritici-repentis Isolates Causing Tan Spot of Winter Wheat in Oklahoma

Tan spot of wheat (Triticum aestivum L.) caused by the fungus Pyrenophora tritici-repentis can be a significant disease, and thus developing resistant varieties using fit and virulent isolates is critical to wheat breeding programs. Hence, variability in fitness characters and virulence was determined for P. tritici-repentis isolates collected in three different decades (1980s, 1990s, and 2000s) from Oklahoma wheat fields. Growth (hyphal extension on media), conidia production on media, and conidia production on wheat leaves of a susceptible (TAM 105), an intermediate (Deliver) and a resistant (Red Chief) variety were determined for 17 isolates. Isolates also were evaluated for pseudothecia production and ascospore maturity on wheat straw, and isolate virulence was determined on all three wheat varieties. Isolates differed significantly (p<0.01) for growth on media, and for conidia production on agar and on wheat leaves. Isolates varied significantly in pseudothecia production and ascospore maturity on wheat straw. Isolates differed significantly (p<0.01) in virulence (percent leaf area infection) on the three wheat cultivars. Isolates collected from the 2000s were more virulent than those collected in the 1980s and 1990s, and also were deemed to be more fit than isolates collected in the 1980s and 1990s in terms of traits such as growth, conidia production, pseudothecia formation, mature ascospore production, and virulence. This fitness could affect the competitiveness of these isolates in the field, as well as facilitate selection of isolates used to test wheat germplasm lines for reaction to tan spot.

Trichaleurina javanica from West Java

Trichaleurina is a fleshy mushroom with goblet-shaped within Pezizales. Many genera have a morphology similar to Trichaleurina, such as Bulgaria and Galiella. Some previous reports had been described fungi like Trichaleurina as Sarcosoma. Indonesia has been reported that has Trichaleurina specimen (the new name of Sarcosoma) by Boedijn. This research aimed to obtain, characterize, and determine the Trichaleurina around IPB University. Field exploration for fungal samples was used in the Landscape Arboretum of IPB University. Ascomata of Trichaleurina were collected, observed, and preserved using FAA. The specimen was deposited into Herbarium Bogoriense with collection code BO 24420. The molecular phylogenetic tree using RAxML was used to identify the species of the specimen.&#x0D; Morphological data were used to support the species name of the specimen. Specimen BO 24420 was identified as Tricahleurina javanica with 81% bootstrap value. Molecular identification was supported by the morphological data, such as the two oil globules and the size of mature ascospores.

&lt;p&gt;&lt;strong&gt;Morphological and molecular identification of &lt;em&gt;Alternaria hedjaroudei&lt;/em&gt; &lt;em&gt;sp. nov&lt;/em&gt;., a new species in section &lt;em&gt;Panax&lt;/em&gt; from Iran&lt;/strong&gt;&lt;/p&gt;

Alternaria section Panax currently includes seven species which are characterized by simple or branched and short to moderately long primary conidiophores as well as by solitary to short simple or branched chains of conidia with moderate to relatively long secondary conidiophores. Two species within the section have been reported to have sexual morphs. During the study of fungi on overwintered stems of Serratula coriacea Fisch. &amp; C.A.Mey. (Asteraceae) from different regions of Urmia, West Azarbaijan Province, Iran, 26 isolates (16 from conidia and 10 from single ascospores) representing Alternaria characteristics were isolated and studied taxonomically. All the studied isolates formed sexual morph on PDA, PCA, V8–A, and HA media after 2 months incubation in the dark at 4 ºC and fully matured after 3 months. Based on morphological characteristics of sexual and asexual morphs and multiple gene sequences analyses (ITS-rDNA, GAPDH, RPB2 and TEF), the newly studied isolates represented a new species in Alternaria section Panax, which is described and illustrated here as Alternaria hedjaroudei. Alternaria hedjaroudei is phylogenetically close to A. avenicola, A. calycipyricola and A. photistica but it can be distinguished morphologically based on the formation of short chains of conidia (2–3 in a chain), absence of chlamydospores, smaller conidia size, relatively short secondary conidiophores, and fewer transverse septa in mature ascospores. Detailed morphological comparisons with other species in the section Panax are provided.

Influence of moisture on maturation rate of the Venturia inaequalis (Cooke) Wint. ascospores in central Poland

Influence of moisture on maturation rate of the airborne Venturia inaequalis (Cooke) Wint. ascospores was numerically examined using field data collected from spore traps in Skierniewice, Poland, during 2005–2008 and 2010–2014. In order to avoid a bias resulting from different abundance of airborne ascospores in different seasons, the cumulative proportions of mature ascospores observed in the season were analyzed. The maturation rate was expressed with the probit of cumulative proportion which linearized its sigmoidal shape and made the effect of the weather during the entire season of the same importance. The significant, positive association between the probit slope and the moisture indicators related to precipitation, leaf wetness, and high relative humidity were observed. Additionally, the significant, negative relationships with the average and maximal air temperature were found. The results suggested that rain should be treated as the major factor driving the ascospore development and leaf wetness as the second factor. In conclusion, the main sources of moisture should be taken into account in statistical models of ascospore development, but it seems reasonable to take into account different strength of their association with the development rate.

Two new species of Amphisphaeria (Amphisphaeriaceae) from northern Thailand

Amphisphaeria flava and A. thailandica sp. nov. collected from dead woody branches in northern Thailand, are described and illustrated in this study. Both taxa share ascomata with a brown outer peridium and 1-septate brown ascospores in uniseriate asci. Amphisphaeria flava is characterized by a pale yellow host surface around the ostiole, a comparatively thin peridium and multi-guttulate mature ascospores. Amphisphaeria thailandica is characterized by subglobose to oval ascospores. Maximum parsimony, maximum likelihood and Bayesian inference analyses of a combined LSU and ITS DNA sequence matrix shows that A. flava and A. thailandica are distinct from other Amphisphaeria species (with molecular data available) in family Amphisphaeriaceae. A key to accepted Amphisphaeria species is provided.

Sexuality and compatibility of Bipolaris sorokiniana and segregation pattern in teleomorph (Cochliobolus sativus): geographic origin and segregation ratio

Bipolaris sorokiniana causes leaf blotch of wheat, inflicting substantial grain yield losses in worldwide. Knowledge of sexual recombination of the fungal pathogen can help to implement effective disease management strategies. Randomly selected 51 isolates of B. sorokiniana was studied to test compatibility with all possible combinations. Six most fertile crosses were found, among them two combinations (BS-81 x BS-83 and BS-60 x BS-134) produced abundant pseudothecia with mature and viable ascospores. This result disclosed that most compatible isolates belonging in the Lower and High Ganges river floodplain area of Bangladesh. One hundred and fifty ascospores were examined to evaluate morphological segregation pattern. Morphological characters (ascospores color, shape, size, Septation number and colony color) of ascospores reject the null hypothesis. A clear 3:1 and 9:7 segregation ratios for ascospores color, mycelial colony color, shape of ascospores; and size of ascospores, septation number of ascospores were found in F2 progeny. RAPD technique was tested to determine molecular segregation pattern in 75 F2 progeny following reproducible polymorphic prominent bands. After test of independence, each trait showed significant and fit to the 1:1 segregation ratio expected for Mendelian trait.

Morphological and molecular characterization, sexual reproduction, and pathogenicity of Setosphaeria rostrata isolates from rice leaf spot

Setosphaeria rostrata, a common plant pathogen causing leaf spot disease, affects a wide range of plant species, mainly grasses. Fungi were isolated from brown spots on rice leaves throughout Peninsular Malaysia, and 45 isolates were identified as Setosphaeria rostrata. The isolates were then characterized using morphological and molecular approaches. The mating type was determined using PCR amplification of the mating type alleles, and isolates of opposite mating types were crossed to examine sexual reproduction. Based on nuclear ribosomal DNA ITS1-5.8S-ITS2 region (ITS) and beta-tubulin (BT2) sequences, two phylogenetic trees were constructed using the maximum likelihood method; S. rostrata was clustered in one well-supported clade. Pathogenicity tests showed that S. rostrata isolates are pathogenic, suggesting that it is the cause of the symptoms. Mating-type analyses indicated that three isolates carried the MAT1-1 allele, and the other 42 isolates carried MAT1-2. After isolates with opposite mating types were crossed on Sach’s medium and incubated for 3 wk, six crosses produced pseudothecia that contained eight mature ascospores, and 12 other crosses produced numerous pseudothecia with no ascospores. To our knowledge, this is the first report on S. rostrata isolated from leaf spots on rice.

Effects of Weather Variables on Ascospore Discharge from Fusarium graminearum Perithecia.

Fusarium graminearum is a predominant component of the Fusarium head blight (FHB) complex of small grain cereals. Ascosporic infection plays a relevant role in the spread of the disease. A 3-year study was conducted on ascospore discharge. To separate the effect of weather on discharge from the effect of weather on the production and maturation of ascospores in perithecia, discharge was quantified with a volumetric spore sampler placed near maize stalk residues bearing perithecia with mature ascospores; the residues therefore served as a continuous source of ascospores. Ascospores were discharged from perithecia on 70% of 154 days. Rain (R) and vapor pressure deficit (VPD) were the variables that most affected ascospore discharge, with 84% of total discharges occurring on days with R≥0.2 mm or VPD≤11 hPa, and with 70% of total ascospore discharge peaks (≥ 30 ascospores/m3 air per day) occurring on days with R≥0.2 mm and VPD≤6.35 hPa. An ROC analysis using these criteria for R and VPD provided True Positive Proportion (TPP) = 0.84 and True Negative Proportion (TNP) = 0.63 for occurrence of ascospore discharge, and TPP = 0.70 and TNP = 0.89 for occurrence of peaks. Globally, 68 ascospores (2.5% of the total ascospores sampled) were trapped on the 17 days when no ascospores were erroneously predicted. When a discharge occurred, the numbers of F. graminearum ascospores sampled were predicted by a multiple regression model with R2 = 0.68. This model, which includes average and maximum temperature and VPD as predicting variables, slightly underestimated the real data and especially ascospore peaks. Numbers of ascospores in peaks were best predicted by wetness duration of the previous day, minimum temperature, and VPD, with R2 = 0.71. These results will help refine the epidemiological models used as decision aids in FHB management programs.

New Insights into the Life Cycle of the Wheat Powdery Mildew: Direct Observation of Ascosporic Infection in Blumeria graminis f. sp. tritici.

Although Blumeria graminis is an intensively studied pathogen, an important part of its life cycle (namely, the way ascospores initiate primary infections on cereal leaves) has not yet been explored in detail. This study reports, for the first time, the direct observation of this process in B. graminis f. sp. tritici using light and confocal laser-scanning microscopy. All the germinated ascospores produced a single germ tube type both in vitro and on host plant surfaces; therefore, the ascosporic and conidial germination patterns are markedly different in this fungus, in contrast to other powdery mildews. Germinated ascospores penetrated the epidermal cells of wheat leaves and produced haustoria as known in the case of conidial infections. This work confirmed earlier studies reporting that B. graminis chasmothecia collected from the field do not contain mature ascospores, only asci filled with protoplasm; ascospore development is induced by moist conditions and is a fast process compared with other powdery mildews. Although ascosporic infections are frequent in B. graminis f. sp. tritici in the field, as shown by this study and other works as well, a recent analysis of the genomes of four isolates revealed the signs of clonal or near-clonal reproduction. Therefore, chasmothecia and ascospores are probably more important as oversummering structures than genetic recombination factors in the life cycle of this pathogen.

The filamentous ascomycete Sordaria macrospora can survive in ambient air without carbonic anhydrases

The rapid interconversion of carbon dioxide and bicarbonate (hydrogen carbonate) is catalysed by metalloenzymes termed carbonic anhydrases (CAs). CAs have been identified in all three domains of life and can be divided into five evolutionarily unrelated classes (α, β, γ, δ and ζ) that do not share significant sequence similarities. The function of the mammalian, prokaryotic and plant α-CAs has been intensively studied but the function of CAs in filamentous ascomycetes is mostly unknown. The filamentous ascomycete Sordaria macrospora codes for four CAs, three of the β-class and one of the α-class. Here, we present a functional analysis of CAS4, the S. macrospora α-class CA. The CAS4 protein was post-translationally glycosylated and secreted. The knockout strain Δcas4 had a significantly reduced rate of ascospore germination. To determine the cas genes required for S. macrospora growth under ambient air conditions, we constructed double and triple mutations of the four cas genes in all possible combinations and a quadruple mutant. Vegetative growth rate of the quadruple mutant lacking all cas genes was drastically reduced compared to the wild type and invaded the agar under normal air conditions. Likewise the fruiting bodies were embedded in the agar and completely devoid of mature ascospores.

Herpothallon inopinatum(Arthoniaceae), a New Lichen Species from Mexico

Herpothallon inopinatum Frisch & G. Thor sp. nova (Arthoniaceae) is described from Chiapas, Mexico. It is the second species of Herpothallon known with mature ascospores besides H. fertile. It differs from H. fertile in smaller asci immersed in the thallus instead of strongly protruding barrel-shaped thallus warts, smaller bean-shaped ascospores and a thallus chemistry of confluentic and chiodectonic acids. Herpothallon inopinatum most closely resembles H. pustulatum, but differs in the absence of pustules. The closely related H. rubrocinctum has distinct bright-red pseudoisidia and a bright-red pigmentation of pro- and hypothallus, while H. inopinatum lacks pseudoisidia and has a pro- and hypothallus white mottled pale yellow to orange to purplish brown. A revised key to all 34 currently accepted species in Herpothallon is provided.

Molecular phylogeny resolves a taxonomic misunderstanding and placesGeisleriaclose toAbsconditellas. str. (Ostropales:Stictidaceae)

AbstractThe phylogenetic position of the genusGeisleriaand its type speciesG. sychnogonioideswas reconstructed using sequence data of the mitochondrial small subunit (mtSSU), the nuclear large subunit rDNA (nuLSU) and the first subunit of the RNA polymerase (RPB1). The species, previously classified inVerrucariaceae(Eurotiomycetes) andStrigulaceae(Dothideomycetes), is sister to the type of the genusAbsconditella,A. sphagnorum, and nested within the generaAbsconditellaandCryptodiscuscombined (which also includes the lichenizedBryophagus). At first glance it appears to be a further example of parallel evolution of perithecioid ascomata withinStictidaceae(Lecanoromycetes:Ostropales), besidesOstropaandRobergea, adding to the growing list of perithecioid forms nested within apothecioid lineages in Ostropomycetidae, and specificallyOstropales, with other examples known fromGraphidaceae(several genera),Gyalectaceae(Belonia), andPorinaceae. However, revision of type material collected by Nitschke revealed that the species actually develops typical apothecia with a narrowly exposed disc. We conclude thatGeisleria sychnogonoideswas erroneously considered a pyrenocarpous taxon, because in dry conditions the apothecia are closed and not recognizable as such. The species usually grows on unstable soil and therefore often only develops young, more or less closed ascomata (yet with mature ascospores), and has also been confused with the superficially similarBelonia incarnata, in which the ascomata remain closed even when mature.Geisleriasychnogonioideshas so far only been known as a rarely reported pioneer species from loamy soils in Europe and North America. Here it is reported to occur abundantly on lateritic soils in subtropical Brazil, suggesting that it is cosmopolitan and possibly common, but much overlooked.

Eisosome distribution and localization in the meiotic progeny of Aspergillus nidulans

In the model filamentous fungus Aspergillus nidulans, PilA and PilB, two homologues of the Saccharomyces cerevisiae eisosome proteins Pil1/Lsp1, and SurG, a strict orthologue of Sur7, are assembled and form tightly packed structures in conidiospores. As A. nidulans differs in its reproduction pattern from the Saccharomycotina in that it has the ability to reproduce through two different types of spores, conidiospores and ascospores, the products of the asexual and the sexual cycle respectively, we investigated the eisosome distribution and localization during the sexual cycle. Our results show that core eisosome proteins PilA, PilB and SurG are not expressed in hülle cells or early ascospores, but are expressed in mature ascospores. All eisosomal proteins form punctate structures at the membrane of late ascospores. In mature but quiescent ascospores, PilA forms static punctate structures at the plasma membrane. PilB also was observed at the ascospore membrane as well, with higher concentration at the areas where the two halves of ascospores are joined together. Finally, SurG was localized both at the membrane of ascospores and perinuclearly. In germlings originating from ascospores the punctate structures were shown to be composed only of PilA. PilB is diffused in the cytoplasm and SurG was located in vacuoles and endosomes. This altered localization is identical to that found in germlings originated from conidiospores. In germinated ascospores PilA foci did not colocalise with the highly mobile and transient peripheral punctate structures of AbpA, a marker for sites of clathrin-mediated endocytosis. Deletions of each one or all the three core eisosomal genes do not affect viability or germination of ascospores. In the presence of myriocin – a specific inhibitor of sphingolipid biosynthesis – PilA-GFP foci of ascospore germlings were less numerous and their distribution was significantly altered, suggesting a correlation between PilA foci and sphingolipid biosynthesis.

Functional analyses of the nitrogen regulatory gene areA in Gibberella zeae

Fusarium head blight caused by Gibberella zeae is a prominent disease of cereal crops that poses serious human health concerns due to the contamination of grains with mycotoxins. In this study, we deleted an orthologue of areA, which is a global nitrogen regulator in filamentous fungi, to characterize its functions in G. zeae. The areA deletion resulted in an inability to use nitrate as a sole nitrogen source, whereas urea utilization was partially available. The virulence of ΔareA strains on wheat heads was markedly reduced compared with the wild-type strain. The areA mutation triggered loss of trichothecene biosynthesis but did not affect zearalenone biosynthesis. The ΔareA strains showed immaturity of asci and did not produce mature ascospores. Chemical complementation by urea restored normal sexual development, whereas the virulence and trichothecene production were not affected by urea addition. GFP-AreA fusion protein was localized to nuclei, and its expression increased in response to nitrogen-limiting conditions. These results suggest that areA-dependent regulation of nitrogen metabolism is required for vegetative growth, sexual development, trichothecene biosynthesis, and virulence in G. zeae.

Three new Pronectria species in terricolous and saxicolous microlichen communities (Bionectriaceae, Ascomycota)

Abstract: Three new species of Bionectriaceae are described; (1) Pronectria algicola grows within Gloeocystis-like algal crusts with Steinia geophana and Sarcosagium campestre; (2) P. caloplacae is lichenicolous on areoles of Caloplaca austrocitrina; (3) P. diplococca grows in squamules of Collema spp. The latter species has early dissolving asci and 1-septate ascospores disintegrating into simple globose mature ascospore fragments. Fragmentation of ascospores appears in three additional species of Pronectria. A key to the known Pronectria species is provided.