Small-spored Species Research Articles

Pathogenicity of Single and Combined Inoculations of Alternaria spp. on Potato

AbstractIn Europe, early blight has become an increasingly severe disease in potato cultivation. We investigated the lesion sizes caused by infection with different Alternaria spp. predominantly from the UK in detached-leaf assays and whole-plant assays of different potato cultivars. The analysis reveals that whilst there is no significant interaction between the potato cultivars used and the Alternaria spp. tested, statistically significant differences were observed between the group means of small-spored vs. large-spored Alternaria spp. The small-spored species grouped together with no significant differences amongst them, whilst large-spored species were able to cause large expanding lesions in all tests, with Alternaria solani exhibiting the most extensive lesion growth. We also explored the effect of co-inoculation treatments of Alternaria spp., revealing no significant difference between the inoculation with Alternaria solani alone and co-inoculation with Alternaria solani and Alternaria alternata or Alternaria infectoria. The findings contribute to our understanding of the behaviour of Alternaria spp. and their impact on potato cultivars.

Small-spored Alternaria spp. (section Alternaria) are common pathogens on wild tomato species.

The wild relatives of modern tomato crops are native to South America. These plants occur in habitats as different as the Andes and the Atacama Desert and are, to some degree, all susceptible to fungal pathogens of the genus Alternaria. Alternaria is a large genus. On tomatoes, several species cause early blight, leaf spots and other diseases. We collected Alternaria-like infection lesions from the leaves of eight wild tomato species from Chile and Peru. Using molecular barcoding markers, we characterized the pathogens. The infection lesions were caused predominantly by small-spored species of Alternaria of the section Alternaria, like A. alternata, but also by Stemphylium spp., Alternaria spp. from the section Ulocladioides and other related species. Morphological observations and an infection assay confirmed this. Comparative genetic diversity analyses show a larger diversity in this wild system than in studies of cultivated Solanum species. As A. alternata has been reported to be an increasing problem in cultivated tomatoes, investigating the evolutionary potential of this pathogen is not only interesting to scientists studying wild plant pathosystems. It could also inform crop protection and breeding programs to be aware of potential epidemics caused by species still confined to South America.

Taxonomic Revision of the pulcherrima Clade of Metschnikowia (Fungi): Merger of Species

The type strains of 10 small-spored species of the ascomycetous yeast genus Metschnikowia usually form a compact group on the phylogenetic trees inferred from barcode sequences. Based on the name of the species, which was described first (Metschnikowia pulcherrima), the group is frequently referred to as the pulcherrima clade. All strains produce the iron-chelate pigment pulcherrimin and have antagonistic effects on many microorganisms. Recent results of molecular phylogenetic, genetic, and genomic research raised doubts about the taxonomic division of the clade. Those data—combined with results obtained in this study by comprehensive analysis of primary and secondary barcode sequences, physiological tests, and hybridisation experiments—demonstrate that the species cannot be distinguished from each other by the criteria of any of the phenotypic, phylogenetic, and biological species concepts. Therefore, I propose that the species of the pulcherrima clade be merged into one species under the oldest species name, M. pulcherrima.

Draft Genome Sequence of Cumin Blight Pathogen Alternaria burnsii.

The fungal genus Alternaria consists of highly diverse species. They can be isolated readily from soil, water, and many plants, and even from animals and humans. Alternaria burnsii is a small-spored species of section Alternaria. It has been reported as a pathogen, an endophyte, and a saprophyte, and can also be found in indoor air. It causes cumin blight, a destructive disease on cumin (Cuminum cyminum), and also causes other serious diseases, such as pumpkin seed rot, date palm leaf spot, wheat leaf spot, and gray spot of Notopterygium incisum. In this study, we sequenced and assembled the first genome of A. burnsii isolate CBS 107.38. The draft genome can be used as a reference for the further study of related pathogens and comparative genomics of Alternaria species.

Nanoglomus plukenetiae, a new fungus from Peru, and a key to small-spored Glomeraceae species, including three new genera in the “Dominikia complex/clades”

A new arbuscular mycorrhizal fungus was found in an agricultural plantation of Plukenetia volubilis, the inka nut (also called “sacha inchi” or “inka peanut”) in the Amazonia region of San Martin State in Peru. In this site, the inka nut was grown in mixed cultures together with Zea mays and Phaseolus vulgaris. The fungus was propagated in bait and single-species cultures on Sorghum vulgare, Brachiaria brizantha, Medicago sativa, and Plukenetia volubilis as host plants. The fungus differentiates hyaline spores terminally or intercalary on cylindrical to slightly funnel-shaped hyphae, singly or in spore clusters with up to ca. 90 spores per cluster. The spores are bi-layered, (20–)25–36(–45) μm in diameter and show regularly a visible septum at the spore base, despite the small spore and tiny hyphae sizes. Phylogenetically, the new fungus represents a new genus in a separated clade, near to the already known Dominikia clades. It can be distinguished from other species by the small spore size, the characteristics of the spore wall layers, and the clearly visible septum at the spore base, which in Kamienskia and Microkamienskia species has rarely to never been reported, while Dominikia species usually have a higher variability of spore sizes and spore wall characteristics. The fungus is here described under the epithet Nanoglomus plukenetiae, serving as type species of the new genus Nanoglomus. The revision of the species and environmental sequences in the Dominikia clades, based on both morphological and molecular phylogenetic analyses, revealed at least two other new genera: Microdominikia gen. nov., based on D. litorea, and Orientoglomus gen. nov., based on D. emiratia. Finally, in the present study, a key for all small-spored species in the Glomeraceae is included comprising all known Dominikia, Kamienskia, Microdominikia, Microkamienskia, Nanoglomus, Orientoglomus spp., and all small-spored Rhizoglomus spp.

Bridging reproductive and microbial ecology: a case study in arbuscular mycorrhizal fungi

Offspring size is a key trait for understanding the reproductive ecology of species, yet studies addressing the ecological meaning of offspring size have so far been limited to macro-organisms. We consider this a missed opportunity in microbial ecology and provide what we believe is the first formal study of offspring-size variation in microbes using reproductive models developed for macro-organisms. We mapped the entire distribution of fungal spore size in the arbuscular mycorrhizal (AM) fungi (subphylum Glomeromycotina) and tested allometric expectations of this trait to offspring (spore) output and body size. Our results reveal a potential paradox in the reproductive ecology of AM fungi: while large spore-size variation is maintained through evolutionary time (independent of body size), increases in spore size trade off with spore output. That is, parental mycelia of large-spored species produce fewer spores and thus may have a fitness disadvantage compared to small-spored species. The persistence of the large-spore strategy, despite this apparent fitness disadvantage, suggests the existence of advantages to large-spored species that could manifest later in fungal life history. Thus, we consider that solving this paradox opens the door to fruitful future research establishing the relationship between offspring size and other AM life history traits.

Polyozellus multiplex (Thelephorales) is a species complex containing four new species

ABSTRACTGeographic, morphological, and internal transcribed spacer (ITS)-based molecular review of collections identified as Polyozellus multiplex revealed that it is a complex of five phylogenetic species. Average spore size—either less or more than 7 × 6 µm—splits the complex into a small-spored group of two (P. multiplex and P. atrolazulinus) and a large-spored group of three (P. mariae, P. marymargaretae, and P. purpureoniger). Basidiocarps of the small-spored species are somewhat smaller than the large-spored ones, are various shades of blue, dark all the way to black, with brownish tomentum only in early growth, have dark context, and have pilei that tend to flare out at the edge. The large-spored species produce somewhat larger sporocarps, have light or lighter context than the pileipelis, and usually retain some brown on the mature pileipellis, the edge of which tends to curl like a cabbage leaf. All will darken or blacken with age. The species of the P. multiplex complex are distributed in the northern coniferous region, with the exception of Europe. One species (P. atrolazulinus) is known from three regions, eastern Asia, western North America, and northeastern North America. Two species are known from two regions: P. purpureoniger in eastern Asia and northwestern North America and P. multiplex in eastern Asia and eastern North America. Two species have been documented in one region only: P. mariae in northeastern North America and P. marymargaretae in western North America. A combination of location, macromorphology, and spore size will usually differentiate the species of the complex.

Sporormiella longicolla sp. nov. and new Sporormiella records on herbivore dung from Brazil

Seven species of Sporormiella are reported from fresh dung collected in a survey including different vegetation areas throughout Pernambuco, Northeastern Brazil. An undescribed, long-necked, small-spored species is proposed as Sporormiella longicolla. Sporormiella isomera is a new record for the Neotropics; S. australis, S. leporina, and S. pentamera are new records for Brazil; and S. herculea (a rare species) and S. minima are also recorded. Descriptions, photographic plates and digital line drawings are provided, along with an identification key to all Sporormiella species recorded in Brazil. Substrate relationships and distribution data of these fungi are also discussed.

Natural occurrence of mycotoxins and toxigenic capacity of Alternaria strains from mouldy peppers

Sweet pepper (Capsicum annuum) is an important crop cultivated worldwide, with Argentina being one of the major producers in South America. The fruit is susceptible to several fungal diseases, leading to severe economic losses for producers. In this study, Alternaria was found as the prevalent genus in mouldy peppers (50% fruits infected). Morphological identification revealed that all 64 Alternaria isolates belonged to small-spored species, most of them corresponding to A. tenuissima, A. arborescens and A. alternata species-groups. Their secondary metabolite profile was evaluated in vitro; alternariols were synthesized by most of the isolates (91% for alternariol and 92% for alternariol monomethyl ether). A high number of Alternaria spp. also produced tenuazonic acid (64%), altenuene (84%) and tentoxin (72%). In addition, damaged pepper fruits were analysed for the presence of tenuazonic acid and alternariols. A total 32 out of 48 spoiled pepper fruits were contaminated with at least one of these metabolites. Half of the samples were positive for tenuazonic acid (range 8–11,422μg/kg), while alternariol and its monomethyl ether were less frequently detected (21 and 29%, respectively) and at lower concentrations. This is the first report on the natural occurrence of Alternaria mycotoxins in Argentinean sweet pepper, and highlights a consumer risk when mouldy fruits are used in industrialized products because these compounds are not destroyed by conventional heat treatments.

Chloroplast DNA sequences indicate the grammitid ferns (Polypodiaceae) in New Zealand belong to a single clade, Notogrammitis gen. nov.

Eleven species of grammitid ferns have been recognized from New Zealand, in Ctenopteris (one species) and Grammitis (10 species). On account of their morphology, it has previously been hypothesized that these species belong to a single clade. We found this to be supported by phylogenetic analysis of chloroplast DNA sequences, with the species misclassified in their current genera. Consequently, a new genus is described, Notogrammitis, containing a global total of 12 species. The only grammitid species south of 35°30′S worldwide belong to Notogrammitis. Within Notogrammitis, it appears that small-spored species of more benign habitats have originated from within a grade of larger-spored species, generally of colder environments. The chloroplast DNA sequences indicate several cases where the sampled populations may not actually be conspecific: New Zealand and Tasmanian N. billardierei; the circum-Antarctic N. crassior; and the morphologically variable, New Zealand endemic N. ciliata.

Genus assignment of small-spored aquatic and terrestrial species of the Metschnikowia yeasts

Genus assignment of small-spored aquatic and terrestrial species of the Metschnikowia yeasts

Differential gene expression in Alternaria gaisen exposed to dark and light

Character states during sporulation have been used to segregate and describe many small-spored species of Alternaria, but some are not supported by published phylogenetic analyses. The conidiation response of Alternaria gaisen was characterized by selective subtractive hybridization of cDNA produced from cultures of A. gaisen grown either in total darkness or in total darkness followed by scarification and 24 h exposure to light. Transcripts or their translation products were identified using BLAST. Multiple transcripts with similarity to ORF-1 of the AM-toxin gene were obtained from the light library. L152 is a full reading frame EST in the light library whose ORF translation has similarity to the conserved domain aegerolysin (pfam06355). A set of 11 ex-type or representative isolates including A. alternata, A. gaisen, A. yaliinficiens, A. arborescens, A. tenuissima and A. brassicicola were resolved by UPGMA analysis of a partial genomic sequence (415–425 base pairs) of L152, but were not resolved by a similar analysis of ITS sequences. Furthermore, the resolved lineages of the L152 dataset were reflective of the diversity previously hypothesized by morphological evaluations of sporulation patterns. Although the ITS rDNA sequence region is generally accepted as the most likely candidate for fungal barcoding, the analysis of L152 sequences presented here resolved closely related species or species groups where other loci, including ITS, have not. Based on these results, the sequences of putative aegerolysin homologs were variable, parsimony-informative and warrant additional analyses with a broader isolate set including related genera and species.

Molecular and genetic differentiation of small-spored species of the yeast genus Metschnikowia Kamienski

Information concerning molecular reidentification of the species of the rejected genus Chlamydozyma is generalized. The yeasts Chl. pulcherrima Wickerham (1964) and Chl. reukaufii Wickerham (1964) are shown to be sibling species of Metschnikowia pulcherrima Pitt et Miller (1968) and M. reukaufii Pitt et Miller (1968), respectively. Restoration of the species M. zygota (Wickerham) Fell at Hunter (1968) is proposed. The parasexual cycle and the prospects of its application for investigation of the Metschnikowia yeasts are discussed.

Mushroom's spore size and time of fruiting are strongly related: is moisture important?

Most basidiomycete fungi produce annual short-lived sexual fruit bodies from which billions of microscopic spores are spread into the air during a short time period. However, little is known about the selective forces that have resulted in some species fruiting early and others later in the fruiting season. This study of relationships between morphological and ecological characteristics, climate factors and time of fruiting are based upon thorough statistical analyses of 66 520 mapped records from Norway, representing 271 species of autumnal fruiting mushroom species. We found a strong relationship between spore size and time of fruiting; on average, a doubling of spore size (volume) corresponded to 3 days earlier fruiting. Small-spored species dominate in the oceanic parts of Norway, whereas large-spored species are typical of more continental parts. In separate analyses, significant relationships were observed between spore size and climate factors. We hypothesize that these relationships are owing to water balance optimization, driven by water storage in spores as a critical factor for successful germination of primary mycelia in the drier micro-environments found earlier in the fruiting season and/or in continental climates.

First Report of Leaf Spot on Japanese Plum Caused by an Alternaria sp. in Korea.

Japanese plum (Prunus salicina Lindley) is a deciduous tree in the family Rosaceae. In Korea, this plant is widely distributed in orchards as an important stone fruit as well as in gardens as an ornamental tree because of their abundant white blossoms. Every September to November since 2001, leaf spots were observed on Japanese plum in a garden in Cheongyang, Chungnam District, Korea. Early symptoms consisted of small, brown spots that were 2 to 5 mm in diameter. Later, the leaf lesions became circular or irregular, dark brown, expanded to 15 mm in diameter, and resulted in discoloration with necrosis on twisted leaves that was followed by defoliation. In November, older lesions sometimes appeared blackish brown as sporulation occurred on the lesions. The causal fungus was isolated from diseased leaves and cultured on potato dextrose agar. A culture has been placed in the CABI Herbarium (IMI Accession No. 387139). Conidial dimension averaged 34 × 12 μm. On the basis of morphological characteristics of conidia and conidiophores, the causal fungus was identified as a small-spored species of Alternaria as described by E. G. Simmons (1). Pathogenicity tests were conducted by inoculating slightly wounded and nonwounded leaves with a conidial suspension adjusted to 1 × 106 conidia/ml. Four leaves per each experiment were either wounded or not and inoculated with a spore suspension. The eight leaves were placed in a moist chamber at 25°C. After 6 to 10 days, small brown spots appeared on 87% of the wounded and nonwounded leaves. Control leaves sprayed with distilled water did not develop any symptoms. The causal fungus was consistently reisolated from the leaf spots. Results from pathogenicity tests were similar in a repeated test. It is possible that small-spored Alternaria spp. isolates are host specific (2). Eight Alternaria spp., including A. alternata, A. tenuis, A. tenuissima, and A. citri, have been found to cause black spot on fifteen Prunus spp. in China, Japan, Hong Kong, Libya, Mexico, Australia, and the United States (2). Further studies on the host-specific toxin production, geographical distribution, and host ranges for the species of Alternaria isolated from Japanese plum are in progress. To our knowledge, this is the first report of leaf spot on Japanese plum (P. salicina) caused by a small-spored Alternaria sp. in Korea.

A remarkable new freshwater Verrucaria from Europe

Verrucaria madida is described as a new species with a green, subgelatinous thallus, and small ascospores; it is unique in the genus in having 4-spored asci. Three other small-spored freshwater species, V. aquatilis, V. rheitrophila and V. scabra, are compared. The type of pigment in the thallus cortex is a useful taxonomic character in the genus.

Trichoderma brevicompactum sp. nov.

Trichoderma brevicompactum, a new species, was isolated from soil or tree bark in North, Central and South America, including the Caribbean Islands, and southwestern and southeastern Asia. Morphological and physiological characters, the internal transcribed spacer regions of the rDNA cluster (ITS1-5.8SrDNA-ITS2) and partial sequences of translation elongation factor 1-alpha (tef1) are described. Trichoderma brevicompactum is characterized by a pachybasium-type morphology, morphologically resembling other small-spored species referable to Trichoderma section Pachybasium but with essentially subglobose conidia. It is most closely related phylogenetically to Hypocrea lutea, from which it differs in morphological and physiological characters.

Molecular, ecological and evolutionary approaches to understanding Alternaria diseases of citrus

SUMMARY Alternaria fungi cause four different diseases of citrus: Alternaria brown spot of tangerines, Alternaria leaf spot of rough lemon, Alternaria black rot of several citrus fruits and Mancha foliar of Mexican lime. The first three diseases are caused by the small-spored species, Alternaria alternata and the causal agents can only be differentiated using pathogenicity tests, toxin assays or genetic markers. Mancha foliar is caused by the morphologically distinct, large-spored species A. limicola. Substantial progress has been made in understanding the biology, ecology, population biology, systematics, molecular biology and biochemistry of the interactions between these pathogens and citrus. Epidemiological studies have focused on brown spot of tangerines and their hybrids and have contributed to the development of a model of disease development which has improved control and reduced fungicide use. Studies of the population genetics, host specificity and ecology of A. alternata from different ecological niches on citrus have revealed host specific forms of the pathogen which cause disease on different citrus species, the existence of three phylogenetic lineages of the fungus which cause brown spot world-wide, and closely related non-pathogenic isolates which colonize healthy citrus tissue. The role of host-specific toxins in Alternaria diseases of citrus has been extensively studied for over 20 years, and these pathosystems have become model systems for host-pathogen interactions. Recent molecular research has started to unravel the genetic basis of toxin production and the host susceptibility to toxin, and the role of extracellular, degradative enzymes in disease.

Morphology, taxonomy, and nomenclature of theChrysomyxa ledicomplex and related rust fungi on spruce and Ericaceae in North America and Europe

Rust fungi in the genus Chrysomyxa Unger occur in boreal forests of the northern hemisphere on Pinaceae (mostly Picea A. Dietr.), and most species alternate to angiosperm hosts in the Ericaceae. About 30 species are known worldwide. Although several species are economically important pathogens of spruce and rhododendrons, knowledge about species delineations, relationships among species on different continents, and life cycles is lacking. A group of species with similar spore size, including the Chrysomyxa ledi de Bary complex, was re-examined using field observations, inoculation experiments, and light and scanning electron microscopy. In addition to host specificity, characters found useful in species delineation were urediniospore and aeciospore size and shape, and ornamentation of spores and the aecial peridium. Detailed descriptions are given for eight Chrysomyxa species, including synonyms, types, distribution, relationship to other species, and disease impact. The members of the C. ledi complex are considered separate species: Chrysomyxa ledi on Ledum palustre, Chrysomyxa nagodhii sp.nov. on Ledum groenlandicum and Ledum decumbens, Chrysomyxa neoglandulosi sp.nov. on Ledum glandulosum, Chrysomyxa cassandrae on Chamaedaphne calyculata, Chrysomyxa rhododendri on Rhododendron spp., and Chrysomyxa vaccinii comb.nov. on Vaccinium parvifolium. Chrysomyxa chiogenis, with similar spore size, is included for comparison. A previously unrecognized small-spored species, Chrysomyxa reticulata sp.nov., is described on Ledum spp. and Rhododendron spp. Evidence is presented that C. reticulata spreads from native Ledum spp. in North America to cultivated rhododendrons. A new anamorphic species, Peridermium zilleri, likely belonging in Chrysomyxa, is described on Picea sitchensis from coastal British Columbia.Key words: Uredinales, Rhododendron, needle rust, Ledum, systematics.

Recombination in Calonectria morganii and phylogeny with other heterothallic small-spored Calonectria species

The Calonectria morganii cultures studied were isolated from several hosts in the USA. Isolates were mated in all combinations, and one successful mating was selected to establish whether recombination occurred. RAPD and mating type data of parental isolates and progeny confirmed C. morganii to have a heterothallic mating system. Furthermore, to infer the phylogenetic relationships of C. morganii with morphologically similar Calonectria spp., DNA sequences of the ribosomal 5.8S RNA gene and the flanking internal transcribed (ITS) spacers, as well as part of the high mobility group (HMG) box (forming part of the MAT2 mating type gene) and the β-tu-bulin gene, were analyzed. Maximum parsimony yielded concordant trees for all three data sets. These data supported the morphological and biological species concepts proposed for C. morganii and other Calonectria species with, similar, small-spored Cylindrocladium anamorphs.