Multilocus Phylogeny Research Articles

Haplotype diversity and phylogeny within Alternaria alternata and A. arborescens species complexes from tomatoes

Tomato (Solanum lycopersicum L.) is an economically important vegetable susceptible to various fungal diseases, including leaf spot caused by Alternaria spp. from the section Alternaria. In our study, a total of 72 tomato-associated Alternaria spp. strains from Latvia, Lithuania, Estonia, and Algeria were analysed by integrating morphological data, pathogenicity assay, multi-locus phylogeny, and haplotype assignment. Recovered Alternaria spp. strains were characterized by considerable variation in phenotypic diversity, non-pathogenicity to their host of origin and absence of the AAL-toxin biosynthesis gene (ALT1). Multi-locus phylogeny of the RNA polymerase II second largest subunit (rpb2), putative F-box-domain-containing protein (ASA-10), and putative histone-like transcription factor (ASA-19) confirmed the occurrence of both A. alternata and A. arborescens species complexes along with A. longipes and A. postmessia on symptomatic tomatoes. The discordant tree topology among single-gene phylogenies suggested the occurrence of potential recombination between phylogenetic lineages in the section Alternaria, resulting in putative alternata-arborescens and alternata-longipes hybrids. DNA polymorphism analysis of the rpb2, ASA-10, and ASA-19 loci revealed a high level of genetic diversity in the section Alternaria, and the number of single nucleotide polymorphisms (SNPs) and haplotypes varied among loci and lineages studied. A total of 16 and 6 multi-locus haplotypes were assigned in alternata and arborescens lineages, respectively. Global genetic diversity analysis of A. alternata and A. arborescens strains at the rpb2 locus confirmed that major haplotypes described from tomatoes were shared among other hosts of origin.

Nigrospora solani (Amphisphaeriales, Apiosporaceae) a new endophytic species from Brazil

Nigrospora species are found in various substrates as saprobes and are associated with plants as pathogens and endophytes. In this study, we describe N. solani from healthy Solanum lycopersicum var. cerasiforme leaves in an organic farming system in Brazil. Based on morphological data and multi-locus phylogeny using internal transcribed spacer, beta-tubulin, and translation elongation factor 1, N. solani can be recognized as a new species for further research. This anamorphic species is characterized by conidiogenous cells discrete, globose, subglobose, and or ampulliform, solitary, initially hyaline becoming pale brown with age and by conidia solitary, globose to subglobose, black, smooth, and aseptate.

Identification and Characterization of Bipolaris secalis and B. setariae Associated with Brown Stripe Disease on Saccharum officinarum in Yunnan, China.

Sugarcane (Saccharum officinarum L.) is the most important sugar crop that belongs to the Poaceae family and is mainly cultivated in tropical and subtropical regions worldwide. Brown stripe disease is a common and serious foliar fungal disease of sugarcane and has become a serious threat to sugarcane production in China. Yunnan Province is the second-largest sugar base in China. From 2022 to 2023, brown stripe-like symptoms from different sugarcane variety were observed in Yunnan. Thirty-nine Bipolaris-like isolates were obtained from symptomatic leaves of different sugarcane varieties and characterized by means of morphological identification combined with multilocus phylogeny comprising internal transcribed spacer rDNA (ITS), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and the translation elongation factor-1α (TEF-1α) gene sequence date. The analysis provided strong support for the delineation and identification of two Bipolaris species associated with sugarcane brown stripe, including Bipolaris secalis and B. setariae. Pathogenicity text in potted sugarcane plants confirmed that these two Bipolaris species can cause brown stripe of sugarcane, thus fulfilling Koch's postulates. This study confirmed that B. secalis and B. setariae were the pathogens of brown stripe in Yunnan Province. To the best of our knowledge, this is the first to report B. secalis as causal agents of brown stripe of sugarcane. Collectively, this finding provides a basis for sugarcane brown stripe disease accurate diagnosis and would be helpful to the development of effective management strategies.

Molecular phylogeny reveals a new genus and species of freshwater mussel (Bivalvia, Unionidae, Gonideinae) from Jiangxi, China

Freshwater mussels of the tribe Gonideini (Bivalvia: Unionidae: Gonideinae) constitute one of the most taxonomically diverse groups and serve as keystone species in riverine and lacustrine ecosystems across East Asia. A new genus and species of Gonideini (Bivalvia: Unionidae) is described from Jiangxi, China, as Pseudopostolata angulagen. et sp. nov. based on an integrative analysis of shell morphology and molecular data. The validity of the new genus and species is supported by distinct conchological features: a short, rounded anterior; a long, wide posterior; a slightly downward-curved dorsal margin; and a distinctly obtuse angle at the center of the posterior margin. The multi-locus (COI + 16S rRNA + 28S rRNA) phylogeny showed that the species formed a monophyletic group in the tribe Gonideini of the subfamily Gonideinae. Pseudopostolata angulagen. et sp. nov. is identified as the sister group to a clade comprising the genera Postolata, Cosmopseudodon, Obovalis, Ptychorhynchus, Parvasolenaia, and Koreosolenaia. We emphasize the significant morphological convergence in freshwater mussels, particularly within Gonideini, highlighting the necessity of an integrated taxonomic approach for accurate generic classification of this group.

Assessment of nine markers for phylogeny, species and haplotype identification of Kappaphycus species and Eucheuma denticulatum (Solieriaceae, Rhodophyta)

ABSTRACT Molecular studies have contributed to the taxonomy of carrageenan-producing Kappaphycus spp. and Eucheuma denticulatum. However, unresolved species complexes and the lack of standardization in the use of genetic markers impede the identification of specimens and the delineation of a robust taxonomic framework. Here, nine molecular markers (cox1, cox2–3 spacer, cox2, cox3, COB, ITS, psbA, UPA and rbcL) were used to generate a multilocus phylogeny for 113 fresh eucheumatoid samples and four herbarium specimens. Analyses of species delineation and genetic distances confirmed the monophyly of currently accepted taxa. These analyses suggest that clades previously reported as K. striatus KS1 and KS2 are conspecific, and that E. denticulatum EDA ‘spinosum’ and EDB ‘endong/cacing’ are also conspecific. The results also unveiled possible new taxa from Hawaii and Indonesia. Each molecular marker and combinations thereof were assessed with regard to species identification, ease of amplification and sequencing, and haplotype characterization. All genetic markers recorded at least 94% success in the amplification and sequencing of fresh specimens, with cox1 being the most phylogenetically informative. Automatic partitioning, phylogenetic and tree-based assessments showed cox1, cox2–3 spacer, cox2 and rbcL were able to correctly identify species while cox1+ rbcL, COB+rbcL, cox2+ rbcL or cox1+ COB+rbcL trees best represented the phylogeny with consistently high nodal support. Among individual markers, cox1 identified the greatest number of haplotypes, while UPA, partial rbcL (750 bp), ITS, cox3 and cox2–3 spacer were able to retrieve information from herbarium specimens of 12–16 years of age. These molecular results provide a basis for a database essential for the taxonomic framework, cultivar development and germplasm conservation of eucheumatoids.

Multi-gene phylogeny and phenotypic analyses revealed an association of different Colletotrichum species with inflorescence dieback and leaf spot of arecanut in India

Arecanut is one of the economically important plantation crops. Among the various diseases, inflorescence dieback and leaf spot/blight diseases caused by Colletotrichum spp. have re-emerged as major factors impeding arecanut production in India in the last few years. To understand the Colletotrichum species complex in the arecanut system, we have characterized Colletotrichum spp. associated with inflorescence dieback and leaf spot/blight of arecanut based on a combination of phenotypic, multi-locus (ITS, ACT, CHS-1, GAPDH, HIS3 and TUB2) phylogeny and pathogenicity studies. A total of 26 Colletotrichum isolates were obtained from arecanut inflorescence dieback (13 isolates) and leaf spot/blight (13 isolates) infected tissues collected from disease-endemic regions of Karnataka, Kerala and Tripura states of India. The combined multi-locus sequence analyses and a critical examination of phenotypic characters revealed the association of Colletotrichum aoteaora as a dominant species with arecanut inflorescence dieback disease. On the other hand, two Colletotrichum species, viz., C. siamense and C. fructicola, were distinctly associated with leaf spot/blight disease. All the Colletotrichum isolates studied were pathogenic to arecanut. Significant variations in lesion size were observed among the different species with cross-infection ability on inflorescence and leaves. This study represents the first comprehensive study of Colletotrichum spp. associated with arecanut inflorescence dieback and leaf spot/blight diseases. In addition, the study is the first record of C. aoteaora, C. siamense, and C. fructicola associated with the arecanut crop in this country. It provides valuable information for implementing effective IDM strategies against these diseases in India.

Integrative taxonomy reveals a new species of deep-sea squat lobster (Galatheoidea, Munidopsidae) from cold seeps in the Gulf of Mexico

The western Atlantic Ocean harbors a diverse fauna of squat lobsters, particularly in the family Munidopsidae. This study introduces Munidopsis sednasp. nov., a species only found in the Gulf of Mexico and the first species reported to be endemic to cold seeps in the western Atlantic. Our investigation incorporates morphological analyses including micro-CT scanning evidence, multilocus molecular phylogeny, and mtDNA phylogeography, as well as ecological data derived from in situ observations and geographic distribution patterns to substantiate the recognition of the new species. Shallow molecular divergences and multiple morphological differences differentiate the new species from its closest relative, M. longimanus (A. Milne-Edwards, 1880). Additionally, we explore the potential scenario for ecological speciation within this newly identified taxon and discuss its significance in the context of conservation efforts in the Gulf of Mexico.

Three New Truffle Species (Tuber, Tuberaceae, Pezizales, and Ascomycota) from Yunnan, China, and Multigen Phylogenetic Arrangement within the Melanosporum Group.

Based on a multi-locus phylogeny of a combined dataset of ITS, LSU, tef1-α, and rpb2 and comprehensive morphological analyses, we describe three new species from the Melanosporum group of genus Tuber and synonymize T. pseudobrumale and T. melanoexcavatum. Phylogenetically, the three newly described species, T. yunnanense, T. melanoumbilicatum and T. microexcavatum, differ significantly in genetic distance from any previously known species. Morphologically, T. yunnanense is distinctly different from its closest phylogenetically related species, T. longispinosum, due to its long shuttle-shape spores (average the ratio of spore length to spore width for all spores (Qm) = 1.74). Tuber melanoumbilicatum differs from the other species in having a cavity and long shuttle-shaped spores (Qm = 1.65). Although T. microexcavatum sampled ascomata have relatively low maturity, they can be distinguished from its closely related species T. pseudobrumale by the ascomata size, surface warts, and spore number per asci; additionally, phylogenetic analysis supports it as a new species. In addition, molecular analysis from 22 newly collected specimens and Genebank data indicate that T. pseudobrumale and T. melanoexcavatum are clustered into a single well-supported clade (Bootstrap (BS) = 100, posterior probabilities (PP) = 1.0); and morphological characteristics do not differ. Therefore, based on the above evidence and publication dates, we conclude that T. melanoexcavatum is a synonym of T. pseudobrumale. By taking into account current knowledge and combining the molecular, multigene phylogenetic clade arrangement and morphological data, we propose that the Melanosporum group should be divided into four subgroups. Diagnostic morphological features and an identification key of all known species in the Melanosporum group are also included. Finally, we also provide some additions to the knowledge of the characterization of T. pseudobrumale, T. variabilisporum, and T. pseudohimalayense included in subgroup 1 of the Melanosporum group.

Arthrobotrys chinensis, a novel fungal species on marbled bamboo (Chimonobambusa marmorea) leaves in Sichuan Province, China

ABSTRACT During an ongoing survey on fungal diversity in Sichuan Province, China, a white filamentous fungus growing on decaying marbled bamboo (Chimonobambusa marmoreal) leaves was collected. Upon conducting the initial morphological diagnosis and BLASTn searches, the new collection was assigned to the genus Arthrobotrys. A multi-locus phylogeny, based on a concatenated dataset (SSU-ITS-LSU-TEF1-RPB2), revealed the new collection in close phylogenetic proximity to Arthrobotrys eryuanensis and A. hyrcanus. However, they differ in shape and size of conidia. The conidia of the new collection are comparatively shorter than those of A. hyrcanus and wider than those of A. eryuanensis. Based on the difference in morphology and molecular data, we proposed the new collection as a new species, Arthrobotrys chinensis sp. nov. Detailed morphological characteristics, illustrations, and an updated phylogeny is provided for the new species.

Six Species of Phyllachora with Three New Taxa on Grass from Sichuan Province, China.

Phyllachora (Phyllachoraceae, Phyllachorales) species are parasitic fungi with a wide global distribution, causing tar spots on plants. In this study, we describe three newly discovered species: Phyllachora chongzhouensis, Phyllachora neidongensis, and Phyllachora huiliensis from Poaceae in China. These species were characterized using morphological traits and multi-locus phylogeny based on the internal transcribed spacer region (ITS) with the intervening 5.8S rRNA gene, the large subunit of the rRNA gene (LSU), and the 18S ribosomal RNA gene (SSU). Three known species of P. chloridis, P. graminis, and P. miscanthi have also been redescribed, because, in reviewing the original references of P. chloridis, P. graminis, and P. miscanthi, these were found to be relatively old and in Chinese or abbreviated. In addition, the illustrations were simple. In molecular identification, the ITS sequence is short, while the ITS, LSU, and SSU are incomplete. Therefore, this study provides new important references for the redescription of three known species and provides further evidence for the identification of new taxa.

Diversity and pathogenicity of Fusarium spp. isolated from cultivated sorghum stems and roots in eastern Australia

AbstractStalk and root rots of cultivated sorghum (Sorghum bicolor) are caused by several Fusarium species worldwide. This study evaluated Fusarium diversity, pathogenicity and population structure amongst 212 isolates obtained from 169 sorghum plants from commercial field crops in eastern Australia. Sequences of translation elongation factor‐1α (tef‐1α), RNA polymerase II largest subunit (rpb1), RNA polymerase II second largest subunit (rpb2) and calmodulin (cmdA) were used to construct multilocus phylogenies that enabled the identification of 16 Fusarium species in Fusarium chlamydosporum species complex (FCSC), Fusarium fujikuroi species complex (FFSC), Fusarium incarnatum‐equiseti species complex and Fusarium oxysporum species complex (FOSC). The majority of isolates (n = 171) belonged to FFSC. The pathogenicity of 17 selected isolates was determined by artificial inoculation of sorghum seedlings and completing Koch's postulates. Isolates of species in FFSC were significantly (p < 0.05) more aggressive as root pathogens in sorghum seedlings than isolates of other species tested and widely distributed across all sampling sites. Amongst the 35 isolates of FOSC, 26 belonged to Fusarium cili, which is only known as an endophyte from healthy roots of Rosa roxburghii in China. Fusarium sporodochiale (in FCSC) and Fusarium contaminatum (in FOSC) are reported as sorghum seedling root rot pathogens for the first time.

Recent fieldwork and fungarium studies double known diversity of Chlorosplenium and improve understanding of species distributions

ABSTRACT Chlorosplenium is a small genus comprising five species of inoperculate discomycetes in the order Helotiales (Leotiomycetes) often recognizable by their bright yellowish-green colors and gregarious growth on wood. In this study, we describe five new species—C. aotearoa, C. australiense, C. cusucoense, C. epimorsicum, and C. hawaiiense—based on a combination of recent fieldwork and examination of previously collected fungarium specimens. We use an integrative taxonomic approach to support the distinction of new species, incorporating morphology and DNA sequence data with biogeography. Macro- and micromorphological features of apothecia for all species and culture characteristics for four of the five new species are documented. A multilocus phylogeny based on nuc rDNA internal transcribed spacer region ITS1-5.8S-ITS2, partial large subunit nuc ribosomal DNA (28S nuc rDNA), and A–B regions of the largest subunit of RNA polymerase II (RPB1) gene is presented. Additionally, we report Chlorosplenium chlora from Europe for the first time and expand our knowledge of the diversity and distributions of species in this genus in America, Australia, and New Zealand.

Taxonomic reintroduction of the holarctic saprotrophic fungus Crepidotus cinnamomeus

Crepidotus is a genus of common saprotrophic fungi well known especially in the Northern Hemisphere, but distribution patterns of individual species are not sufficiently understood. We redefined a taxonomic circumscription of Crepidotus cinnamomeus based on morphological and molecular congruencies between the type material and recent collections. The species is well delimited from other similar and currently accepted species of the genus. Crepidotus cinnamomeus was found to have a broad holarctic distribution with occurrences in North America, Europe and Asia where it grows on twigs and branches of deciduous trees and shrubs in preferably cold humid habitats. Here we present the first multilocus phylogeny of the genus, including portions of the RPB2 gene. Our study highlights the importance of sufficient sampling from broader areas supported by sequence data, which is essential for estimation of species delimitation, distribution and correct name assignment for Crepidotus species.

Diplodia seriata Isolated from Declining Olive Trees in Salento (Apulia, Italy): Pathogenicity Trials Give a Glimpse That It Is More Virulent to Drought-Stressed Olive Trees and in a Warmth-Conditioned Environment.

The fungi Botryosphaeriaceae are involved in olive declines in both the world hemispheres and in all continents where this species is cultivated. In Salento (Apulia, Italy), the Botryosphaeriaceae Neofusicoccum mediterraneum and N. stellenboschiana have been reported as the agents of a branch and twig dieback that overlaps with olive quick decline syndrome caused by Xylella fastidiosa subsp. pauca. In this study, we report the finding of Diplodia seriata, another Botryosphaeriaceae species, in Salento in Xylella fastidiosa-infected olive trees affected by symptoms of branch and twig dieback. Given that its presence was also reported in olive in the Americas and in Europe (Croatia) with different degrees of virulence, we were prompted to assess its role in the Apulian decline. We identified representative isolates based on morphological features and a multilocus phylogeny. In vitro tests showed that the optimum growth temperature of the isolates is around 25-30 °C, and that they are highly thermo-tolerant. In pathogenicity trials conducted over eleven months, D. seriata expressed a very low virulence. Nonetheless, when we imposed severe water stress before the inoculation, D. seriata significatively necrotized bark and wood in a time frame of 35 days. Moreover, the symptoms which resulted were much more severe in the trial performed in summer compared with that in autumn. In osmolyte-supplemented media with a water potential from -1 to -3 Mpa, the isolates increased or maintained their growth rate compared with non-supplemented media, and they also grew, albeit to a lesser extent, on media with a water potential as low as -7 Mpa. This suggests that olives with a low water potential, namely those subjected to drought, may offer a suitable environment for the fungus' development. The analysis of the meteorological parameters, temperatures and rainfall, in Salento in the timeframe 1989-2023, showed that this area is subjected to a progressive increase of temperature and drought during the summer. Thus, overall, D. seriata has to be considered a contributor to the manifestation of branch and twig dieback of olive in Salento. Coherently with the spiral decline concept of trees, our results suggest that heat and drought act as predisposing/inciting factors facilitating D. seriata as a contributor. The fact that several adverse factors, biotic and abiotic, are simultaneously burdening olive trees in Salento offers a cue to discuss the possible complex nature of the olive decline in Salento.

Single-fly genome assemblies fill major phylogenomic gaps across the Drosophilidae Tree of Life.

Long-read sequencing is driving rapid progress in genome assembly across all major groups of life, including species of the family Drosophilidae, a longtime model system for genetics, genomics, and evolution. We previously developed a cost-effective hybrid Oxford Nanopore (ONT) long-read and Illumina short-read sequencing approach and used it to assemble 101 drosophilid genomes from laboratory cultures, greatly increasing the number of genome assemblies for this taxonomic group. The next major challenge is to address the laboratory culture bias in taxon sampling by sequencing genomes of species that cannot easily be reared in the lab. Here, we build upon our previous methods to perform amplification-free ONT sequencing of single wild flies obtained either directly from the field or from ethanol-preserved specimens in museum collections, greatly improving the representation of lesser studied drosophilid taxa in whole-genome data. Using Illumina Novaseq X Plus and ONT P2 sequencers with R10.4.1 chemistry, we set a new benchmark for inexpensive hybrid genome assembly at US $150 per genome while assembling genomes from as little as 35 ng of genomic DNA from a single fly. We present 183 new genome assemblies for 179 species as a resource for drosophilid systematics, phylogenetics, and comparative genomics. Of these genomes, 62 are from pooled lab strains and 121 from single adult flies. Despite the sample limitations of working with small insects, most single-fly diploid assemblies are comparable in contiguity (>1 Mb contig N50), completeness (>98% complete dipteran BUSCOs), and accuracy (>QV40 genome-wide with ONT R10.4.1) to assemblies from inbred lines. We present a well-resolved multi-locus phylogeny for 360 drosophilid and 4 outgroup species encompassing all publicly available (as of August 2023) genomes for this group. Finally, we present a Progressive Cactus whole-genome, reference-free alignment built from a subset of 298 suitably high-quality drosophilid genomes. The new assemblies and alignment, along with updated laboratory protocols and computational pipelines, are released as an open resource and as a tool for studying evolution at the scale of an entire insect family.

Elucidation of the life cycle of a saprotrophic inoperculate discomycete that is associated with pinesap using a polyphasic taxonomic approach

This study deals with the elucidation of the life cycle of an inoperculate discomycete that was eventually collected from conifer wood in South Germany and has become famous for the extraordinary diversity of its secondary metabolites when it was studied extensively during the 1990s. It had then been identified as Lachnum papyraceum (Lachnaceae, Helotiales) based on morphological traits, and extracts from its mycelial cultures were found to possess extraordinary nematicidal and antibiotic activities. Over 60 different secondary metabolites were finally identified from this fungus after extensive variation of culture media and scale-up of production up to 100-L scale. Among the main active principles were mycorrhizin A and chloromycorrhizin A, which had first been reported in 1977 from an unnamed “mycorrhizal” fungus of the hemiparasitic plant, Monotropa hypopitys (pinesap), that was isolated in Sweden. We noted that both the Lachnum strain and the original mycorrhizin producer were still available in the public domain and decided to study them for comparison using a multilocus phylogeny and also generated secondary metabolite profiles of both strains using analytical high performance liquid chromatography coupled to diode array and mass spectrometric detection (HPLC–DAD/MS). Surprisingly, the sequence data as well as the secondary metabolite profiles of both strains were highly similar, and it was also confirmed by phylogenetic methods that the strains are indeed nested within the genus Lachnum by comparison of their ITS, LSU, and RPB2 sequences. The specimen called L. papyraceum in the old publications was tentatively re-identified by Hans-Otto Baral as L. cf. subvirgineum, but substantial further work on the taxonomy of the genus remains to be done, anyway. We conclude that some Lachnum species have a highly complex but all the more interesting life cycle, and the mycorrhizal symbiont partner may invade the host plant, where it may persist as an endophyte and finally turn saprotrophic on the wood of the senescent pine tree. The taxonomy of these fungi should also be further resolved in the future, using a polythetic concept that includes chemotaxonomic data and a multi-locus genealogy.

A multiscale analysis of coralline algae Lithophylloideae (Corallinophycidae, Rhodophyta) shedding new light on understanding cryptic diversity

Cryptic diversity abounds in many biological species, posing challenges to our understanding of biological diversity, conservation and management. Taking the common coralline algae, the subfamily Lithophylloideae as an illustration, this study delved into the implications of cryptic diversity through global-level phylogenetic and geographical analysis based upon Lithophylloideae molecular data worldwide, as well as a multi-locus time-calibrated phylogeny to elucidate their possible evolutionary process. The multiscale analysis revealed the polyphyly in current concept of the genus Lithophyllum. Geographic isolation resulting from the Tethys terminal event (TTE) has led to two distinct distribution regions for this so-called cosmopolitan genus: one regionally distributed along European coasts/Mediterranean that should include the taxonomical Lithophyllum; others widely distributed, particularly among pan-tropic waters, suggesting at least five groups to be rediscovered within the subfamily Lithophylloideae. Meanwhile, the cryptic genus Titanoderma, lacking morphological identification features with Lithophyllum, exhibited differences in distribution and evolutionary patterns consistent with their ecological habits, thus supporting their separation. This study provided useful hints for cryptic diversity, which advocated an integrative thinking to investigating global cryptic diversity and exploring the broad linkages between phylogenetic relationships and evolutionary origin, biogeography, morphological and ecological traits to achieve a more comprehensive understanding of biodiversity.

Phylogenomics, taxonomy and morphological characters of the Microdochiaceae (Xylariales, Sordariomycetes).

Species of the family Microdochiaceae (Xylariales, Sordariomycetes) have been reported from worldwide, and collected from different plant hosts. The proposed new genus and two new species, viz., Macroidriella gen. nov., M.bambusae sp. nov. and Microdochiumaustrale sp. nov., are based on multi-locus phylogenies from a combined dataset of ITS rDNA, LSU, RPB2 and TUB2 with morphological characteristics. Microdochiumsinense has been collected from diseased leaves of Phragmitesaustralis and this is the first report of the fungus on this host plant. Simultaneously, we annotated 10,372 to 11,863 genes, identified 4,909 single-copy orthologous genes, and conducted phylogenomic analysis based on genomic data. A gene family analysis was performed and it will expand the understanding of the evolutionary history and biodiversity of the Microdochiaceae. The detailed descriptions and illustrations of species are provided.

One new species and a new record of saprobic fungi from Ananas comosus in northern Thailand

Pineapple is one of the major crops in northern Thailand. Like all the other plants, it has associated with it, a wide range of fungi, which may be saprobic, pathogenic, or endophytic. This study investigated fungi associated with pineapple in Chiang Rai Province, northern Thailand. Based on morphology and multilocus phylogeny of the ITS and LSU sequence data, Anthracocystis ananatum is described as a new species with its detailed asexual morph. Compared to other Anthracocystis species, A. ananatum has a distinctive morphotype that includes septate hyphae, conidigenous cells, and conidiophores. In addition, Spegazzinia tessarthra is reported for the first time as a new host and geographic record. The complete descriptions, photo-micrographs, and phylogenies of the new species Anthracocystis ananatum and a new host and geographic record S. tessarthra are provided and discussed.

First report of leaf spot on Daphniphyllum macropodum caused by Neopestalotiopsis clavispora in China.

Daphniphyllum macropodum Miq., an evergreen arbor, is widely cultivated in southern China for its ornamental and medicinal value (Su et al. 2013). In October 2019, a severe leaf spot was observed on D. macropodum in Jinggangshan National Nature Reserve in Ji'an city, Jiangxi, China (114°06'23″E, 26°32'25″N). The plants were about 15 years old, and the disease incidence was estimated to be 15% (4/26 plants). The disease primarily appeared as small black spots on the leaves. At the late stage, the spots enlarged and coalesced into regular or irregular gray necrotic lesions with dark margins. We collected five samples per plant and total 20 samples were collected to isolated the pathogen strains. The margin of the diseased tissues was cut into 5 mm × 5 mm pieces; surface disinfected with 70% ethanol and 2% NaOCl for 30 s and 60 s, respectively; and rinsed thrice with sterile water. Tissues were placed on potato dextrose agar (PDA) and incubated at 25°C in the dark. Pure cultures were obtained by single-spore isolation method, and the representative isolates, JRM3, JRM6, and JRM8 were used for morphological studies and phylogenetic analyses. The colonies of three isolates grown on PDA were white, cottony, and flocculent, contained undulate edges with dense aerial mycelium on the surface at 25 °C. Conidiomata was black conidial masses on PDA. Conidia were 5-celled, clavate to fusiform, smooth, 19.3 to 24.4 long × 6.1 to 8.6 μm wide (n = 50). The 3 median cells were dark brown to olivaceous, the central cell was darker than the other 2 cells, and the basal and apical cells were hyaline. All conidia developed one basal appendage (3.4 to 8.3 μm long; n = 50), and 2 to 3 apical appendages (18 to 32 μm long; n = 50), filiform. The morphological characteristics of the isolates are comparable with those of the genus Neopestalotiopsis (Maharachchikumbura et al. 2014). The internal transcribed spacer (ITS) regions, β-tubulin 2 (TUB2) and translation elongation factor 1-alpha (TEF1-α) were amplified from genomic DNA for the three isolates using primers ITS1/ITS4, T1/Bt-2b, EF1-728F/EF-2 (Maharachchikumbura et al. 2014), respectively. The sequences of the isolates were submitted to GenBank (ITS, OQ372202 to OQ372204; TUB2, OQ390129 to OQ390131; TEF1-α, OQ390126 to OQ390128). A maximum likelihood and Bayesian posterior probability analyses using IQtree v. 1.6.8 and Mr. Bayes v. 3.2.6 with the concatenated sequences placed JRM3, JRM6, and JRM8 in the clade of N. clavispora. Based on the multi-locus phylogeny and morphology, three isolates were identified as N. clavispora. To confirm pathogenicity, eight healthy 10-year-old D. macropodum plants growing in the field were chosen, and 4 leaves per plant were wounded with a sterile needle and inoculated with 10 μL conidial suspension per leaf (106 conidia/ml). Eight plants inoculated with sterile water were used as control. All the inoculated leaves were covered with plastic bags to maintian a humidity environment for 2 days. The leaves inoculated with conidial suspension showed similar symptoms to those observed in the field, whereas control leaves were asymptomatic for 10 days. The same fungus were re-isolated from the lesions, whereas no fungus was isolated from control leaves. N. clavispora was determined as the pathogen of a variety of plant diseases, including Kadsura coccinea (Xie et al. 2018), Dendrobium officinale (Cao et al. 2022), Macadamia integrifolia (Santos et al. 2019). However, this is the first report of N. clavispora infecting D. macropodum in China. This work provided crucial information for epidemiologic studies and appropriate control strategies for this newly emerging disease.