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Abstract The pace at which Next Generation Sequence data is being produced continues to accelerate as technology improves. As a result, such data are increasingly becoming accessible to biologists outside of the field of bioinformatics. In contrast, access to training in the methods of genome assembly and annotation are not growing at a similar rate. In this issue, we report on a Genome Assembly Workshop for Mycologists that was held at the Forestry and Agricultural Biotechnology Institute (FABI) at the University of Pretoria, South Africa and make available the 12 draft genome sequences emanating from the event. With the aim of making the process of genome assembly and annotation more accessible to biologists, we provide a step-by-step guide to both genome assembly and annotation, intended to encourage and empower mycologists to use genome data in their research.

Abstract Calonectria leaf blight (CLB) is one of the most important diseases of Eucalyptus trees grown in plantations. This disease poses a serious threat to the sustainability of Eucalyptus plantations in southern China. To better understand the causal agents of CLB, we collected samples of diseased leaves and soil from Eucalyptus plantations from nine regions in Guangdong Province where the disease has become a serious problem. A total of 606 Calonectria isolates were purified from the samples, with 399 and 207 originating from diseased leaves and soils, respectively. Phylogenetic analyses utilizing six gene regions resolved 303 isolates in the C. kyotensis species complex and an equal number of isolates in the C. reteaudii species complex. These two complexes were represented by ten known Calonectria species, including C. aconidialis (12.0%), C. curvispora (0.3%), C. hongkongensis (24.8%), C. ilicicola (0.9%), and C. kyotensis (12.0%) in the C. kyotensis species complex, and C. crousiana (1.0%), C. Guangdongensis (0.3%), C. pseudoreteaudii (40.7%), C. queenslandica (7.3%), and C. reteaudii (0.7%) in the C. reteaudii species complex. Pathogenicity tests showed that all species were capable of causing disease on two tested Eucalyptus genotypes, albeit at varying degrees of aggressiveness. Most isolates (98.3%) in the C. reteaudii species complex were from the diseased leaves, indicating that species in this complex are the main causal agents of CLB outbreak. In addition, a significant number of the C. kyotensis species complex isolates (66.7%) from the soil samples could also cause the disease on Eucalyptus leaves.

Abstract Background Plant-parasitic nematodes (PPNs) that cause most damage include root-knot nematodes (RKNs) which are a major impediment to crop production. Root-knot nematodes, like other parasites, secrete proteins which are required for parasite proliferation and survival within the host during the infection process. Results Here, we used various computational tools to predict and identify classically and non-classically secreted proteins encoded in the Meloidogyne javanica genome. Furthermore, functional annotation analysis was performed using various integrated bioinformatic tools to determine the biological significance of the predicted secretome. In total, 7,458 proteins were identified as secreted ones. A large percentage of this secretome is comprised of small proteins of ≤ 300 aa sequence length. Functional analyses showed that M. javanica secretome comprises cell wall degrading enzymes for facilitating nematode invasion, and migration by disintegrating the complex plant cell wall components. In addition, peptidases and peptidase inhibitors are an important category of M. javanica secretome involved in compatible host-nematode interactions. Conclusion This study identifies the putative secretome encoded in the M. javanica genome. Future experimental validation analyses can greatly benefit from this global analysis of M. javanica secretome. Equally, our analyses will advance knowledge of the interaction between plants and nematodes.

Cyanobacteria are phototrophic organisms widely found in most types of natural habitats in the tropical regions of the world. In this study, we isolated and identified cyanobacterial strains from paddy soil in Hanoi (Vietnam) and investigated their cytotoxic activities. Five isolated cyanobacterial strains showed distinctive profiles of gene sequences (rRNA 16S and rbcL), phylogenetic placements, and morphological characteristics. Based on the polyphasic evaluation, they were classified as Scytonema bilaspurense NK13, Hapalosiphon welwitschii MD2411, Aulosira sp. XN1103, Desikacharya sp. NS2000, and Desmonostoc sp. NK1813. The cytotoxic screening revealed that the extract of strain Scytonema bilaspurense NK13 exhibited potent cytotoxic activities against four human cell lines of HeLa cells, OVCAR-8 cells, HaCaT cells, and HEK-293T cells, with IC50 values of 3.8, 34.2, 21.6, and 0.6 μg/mL, respectively. This is the first time a well-classified Scytonema strain from tropical habitat in Southeast Asia has been recognized as a potential producer of cytotoxic compounds.

Abstract Background Phytophthora cinnamomi is an oomycete pathogen of global relevance. It is considered as one of the most invasive species, which has caused irreversible damage to natural ecosystems and horticultural crops. There is currently a lack of a high-quality reference genome for this species despite several attempts that have been made towards sequencing its genome. The lack of a good quality genome sequence has been a setback for various genetic and genomic research to be done on this species. As a consequence, little is known regarding its genome characteristics and how these contribute to its pathogenicity and invasiveness. Results In this work we generated a high-quality genome sequence and annotation for P. cinnamomi using a combination of Oxford Nanopore and Illumina sequencing technologies. The annotation was done using RNA-Seq data as supporting gene evidence. The final assembly consisted of 133 scaffolds, with an estimated genome size of 109.7 Mb, N50 of 1.18 Mb, and BUSCO completeness score of 97.5%. Genome partitioning analysis revealed that P. cinnamomi has a two-speed genome characteristic, similar to that of other oomycetes and fungal plant pathogens. In planta gene expression analysis revealed up-regulation of pathogenicity-related genes, suggesting their important roles during infection and host degradation. Conclusion This study has provided a high-quality reference genome and annotation for P. cinnamomi. This is among the best assembled genomes for any Phytophthora species assembled to date and thus resulted in improved identification and characterization of pathogenicity-related genes, some of which were undetected in previous versions of genome assemblies. Phytophthora cinnamomi harbours a large number of effector genes which are located in the gene-poor regions of the genome. This unique genomic partitioning provides P. cinnamomi with a high level of adaptability and could contribute to its success as a highly invasive species. Finally, the genome sequence, its annotation and the pathogenicity effectors identified in this study will serve as an important resource that will enable future studies to better understand and mitigate the impact of this important pathogen.

Abstract Species of Armillaria are distributed globally and include some of the most important pathogens of forest and ornamental trees. Some of them form large long-living clones that are considered as one of the largest organisms on earth and are capable of long-range spore-mediated transfer as well as vegetative spread by drought-resistant hyphal cords called rhizomorphs. However, the virus community infecting these species has remained unknown. In this study we used dsRNA screening and high-throughput sequencing to search for possible virus infections in a collection of Armillaria isolates representing three different species: Armillaria mellea from South Africa, A. borealis from Finland and Russia (Siberia) and A. cepistipes from Finland. Our analysis revealed the presence of both negative-sense RNA viruses and positive-sense RNA viruses, while no dsRNA viruses were detected. The viruses included putative new members of virus families Mymonaviridae, Botourmiaviridae and Virgaviridae and members of a recently discovered virus group tentatively named “ambiviruses” with ambisense bicistronic genomic organization. We demonstrated that Armillaria isolates can be cured of viruses by thermal treatment, which enables the examination of virus effects on host growth and phenotype using isogenic virus-infected and virus-free strains.

Abstract Draft genomes of Penicillium roqueforti, Fusarium sororula, Chalaropsis populi, and Chrysoporthe puriensis are presented. Penicillium roqueforti is a model fungus for genetics, physiological and metabolic studies, as well as for biotechnological applications. Fusarium sororula and Chrysoporthe puriensis are important tree pathogens, and Chalaropsis populi is a soil-borne root-pathogen. The genome sequences presented here thus contribute towards a better understanding of both the pathogenicity and biotechnological potential of these species.

ABSTRACT Draft genomes of the fungal species Ambrosiella cleistominuta, Cercospora brassicicola, C. citrullina, Physcia stellaris, and Teratosphaeria pseudoeucalypti are presented. Physcia stellaris is an important lichen forming fungus and Ambrosiella cleistominuta is an ambrosia beetle symbiont. Cercospora brassicicola and C. citrullina are agriculturally relevant plant pathogens that cause leaf-spots in brassicaceous vegetables and cucurbits respectively. Teratosphaeria pseudoeucalypti causes severe leaf blight and defoliation of Eucalyptus trees. These genomes provide a valuable resource for understanding the molecular processes in these economically important fungi.

Abstract Bark beetle galleries are complex ecosystems where many microbes and other arthropods co-exist with the beetles. Fungi isolated from these galleries are often referred to as ‘beetle associates’, but the nature of these associations are poorly understood. The possibility that many of these fungi might in fact be mite associates is often overlooked. Several recent studies explored the diversity of fungi from conifer-infesting bark beetles and their galleries in China, but only one study considered phoretic mites and their fungi from conifer-infesting bark beetles in Yunnan, southwestern China. We studied the mites and fungi from galleries of four spruce-infesting bark beetle species in the high altitude forests of Qinghai province, western China. Mites were identified based on morphological characteristics, and fungi based on DNA sequences of four gene regions. In total, 173 mite individuals were collected belonging to 18 species in 11 genera. A total of 135 fungal isolates were obtained from the mites, representing 14 taxa from the Ophiostomatales. The most frequently isolated fungus was Ophiostoma nitidum, which represented 23.5% of the total isolates. More fungal species were found from fewer mites and bark beetle species than from the study in Yunnan. Although we could not elucidate the exact nature of interactions between mites and their fungi, our results re-enforce that these organisms should not be ignored in pest risk assessments of bark beetles, that often focus only on the beetles and their fungi. Three new species are described: Grosmannia zekuensis, O. manchongi, and O. kunlunense spp. nov., and our data revealed that O. typographi, recently described from China, is a synonym of O. ainoae.

Abstract Draft genome sequences of five Calonectria species [including Calonectria aciculata, C. crousiana, C. fujianensis, C. honghensis and C. pseudoturangicola], Celoporthe dispersa, Sporothrix phasma and Alectoria sarmentosa are presented. Species of Calonectria are the causal agents of Eucalyptus leaf blight disease, threatening the growth and sustainability of Eucalyptus plantations in China. Celoporthe dispersa is the causal agent of stem canker in native Syzygium cordatum and exotic Tibouchina granulosa in South Africa. Sporothrix phasma was first discovered in the infructescences of Protea laurifolia and Protea neriifolia in South Africa. Alectoria sarmentosa is fruticose lichen belongs to the alectorioid clade of the family Parmeliaceae. The availability of these genome sequences will facilitate future studies on the systematics, population genetics, and genomics of these fungi.

Abstract Draft genomes of the fungal species Fusarium xylarioides, Teratosphaeria gauchensis and T. zuluensis are presented. In addition an annotation of the genome of Ceratocystis fimbriata is presented. Overall these genomes provide a valuable resource for understanding the molecular processes underlying pathogenicity and potential management strategies of these economically important fungi.

To meet the growing demand for wood and pulp products, Eucalyptus plantations have expanded rapidly during the past two decades, becoming an integral part of the southern China landscape. Leaf blight caused by various Calonectria spp., is a serious threat to these plantations. In order to explore the diversity and distribution of Calonectria spp. in Fujian Province soils, samples were collected in Eucalyptus plantations and adjacent plantings of Cunninghamia lanceolata, Phyllostachys heterocycle and Pinus massoniana as well as in natural forests. Three hundred and fifty-three Calonectria isolates were recovered from soil samples and they were identified based on a comparison of multilocus DNA sequence data for the act (actin), cmdA (calmodulin), his3 (histone H3), rpb2 (the second largest subunit of RNA polymerase), tef1 (translation elongation factor 1-alpha) and tub2 (β-tubulin) gene regions, as well as morphological characteristics. Six known taxa including Calonectria aconidialis, Ca. hongkongensis, Ca. ilicicola, Ca. kyotensis, Ca. pacifica, Ca. pseudoreteaudii and one novel species described here as Ca. minensis sp. nov. were identified. Of these, Ca. aconidialis and Ca. kyotensis were the most prevalent species, and found in eight and seven sites, and four and five forest types, respectively. Calonectria spp. were most abundant in soils from Eucalyptus stands, followed by P. heterocycle and natural forests. Relatively few species were found in the soils associated with Cunninghamia lanceolata and Pinus massoniana. The abundance of known Calonectria spp. suggests that these fungi have been relatively well sampled in Fujian. The results are also consistent with the fact that most Calonectria diseases are found on Angiosperm as opposed to Gymnosperm plants.

Fusarium circinatum is an important global pathogen of pine trees. Genome plasticity has been observed in different isolates of the fungus, but no genome comparisons are available. To address this gap, we sequenced and assembled to chromosome level five isolates of F. circinatum. These genomes were analysed together with previously published genomes of F. circinatum isolates, FSP34 and KS17. Multi-sample variant calling identified a total of 461,683 micro variants (SNPs and small indels) and a total of 1828 macro structural variants of which 1717 were copy number variants and 111 were inversions. The variant density was higher on the sub-telomeric regions of chromosomes. Variant annotation revealed that genes involved in transcription, transport, metabolism and transmembrane proteins were overrepresented in gene sets that were affected by high impact variants. A core genome representing genomic elements that were conserved in all the isolates and a non-redundant pangenome representing all genomic elements is presented. Whole genome alignments showed that an average of 93% of the genomic elements were present in all isolates. The results of this study reveal that some genomic elements are not conserved within the isolates and some variants are high impact. The described genome-scale variations will help to inform novel disease management strategies against the pathogen.

Symbiosis between beetles and fungi arose multiple times during the evolution of both organisms. Some of the most biologically diverse and economically important are mutualisms in which the beetles cultivate and feed on fungi. Among these are bark beetles and Harringtonia, a fungal genus that produces Raffaelea-like asexual morph and hosts the causal agent of laurel wilt, H. lauricola (formerly Raffaelea lauricola). In this study, we propose four new species of Harringtonia associated with beetles from Belize and Florida (USA). We hope to contribute towards a more robust and inclusive phylogenetic framework for future studies on these beetle-fungi relationships and their potential impact in crops and forests worldwide.

Abstract This genome announcement includes draft genomes from Claviceps purpurea s.lat., including C. arundinis, C. humidiphila and C. cf. spartinae. The draft genomes of Davidsoniella eucalypti, Quambalaria eucalypti and Teratosphaeria destructans, all three important eucalyptus pathogens, are presented. The insect associate Grosmannia galeiformis is also described. The pine pathogen genome of Fusarium circinatum has been assembled into pseudomolecules, based on additional sequence data and by harnessing the known synteny within the Fusarium fujikuroi species complex. This new assembly of the F. circinatum genome provides 12 pseudomolecules that correspond to the haploid chromosome number of F. circinatum. These are comparable to other chromosomal assemblies within the FFSC and will enable more robust genomic comparisons within this species complex.

The Ophiostomatales is an Ascomycete order of fungi that accommodates several tree pathogens and many species that degrade wood. These fungi are commonly vectored by Scolytine bark and ambrosia beetles. In recent years it has also been shown that hyperphoretic mites on these beetles can vector some Ophiostomatales. Little is known regarding the Ophiostomatales in China and we have consequently explored the diversity of these fungi associated with conifer-infesting beetles and mites in Yunnan province. Galleries and beetles were collected for 17 beetle species, while 13 mite species were obtained from six of these beetle species. Collectively, 340 fungal isolates were obtained, 45 from beetles, 184 from mites, 56 from galleries and 55 isolates where the specific niche was not clear. DNA sequences for five gene regions (ITS, LSU, BT, EF, and CAL) were determined for fungal isolates representing different morphological groups. Phylogenetic analyses confirmed the presence of 19 fungal taxa, including five novel species described here as Ophiostoma acarorum sp. nov., Ophiostoma brevipilosi sp. nov., Graphilbum kesiyae sp. nov., Graphilbum puerense sp. nov., and Leptographium ningerense sp. nov. Ophiostoma ips was the most frequently isolated species, representing approximately 31% of all isolates. Six of 19 taxa were present on mites, beetles and in the galleries of the beetles, while three species were found on mites and galleries. Two species were found only on mites and one species only on a beetle. Although the numbers of beetles and mites were insufficient to provide statistical inferences, this study confirmed that mites are important vectors of the Ophiostomatales in China. We hypothesize that these mites are most likely responsible for horizontal transfer of fungal species between galleries of different beetle species. The fact that half of the fungal species found were new to science, suggests that the forests of east Asia include many undescribed Ophiostomatales yet to be discovered.

Species in the Ceratocystis manginecans complex are important fungal pathogens of plantation trees globally. The most important hosts include species of Eucalyptus, Acacia, Mangifera, and Punica. Despite their relevance and widespread occurrence, little is known regarding their population genetics and how this might relate to their host associations or geographic regions in which they occur. A global collection of 491 isolates representing the C. manginecans complex, from four different plant hosts and nine countries, were genotyped using microsatellite markers. Population genetic analyses using numerous tools were conducted to interrogate how their genetic diversity and structure might be affected by host or areas of occurrence. Results of genetic diversity studies showed that when grouping isolates into populations based on their host associations, the population on Eucalyptus was most diverse, and it also has a broad global distribution. When considering countries of origin as a basis for defining populations, the gene and genotypic diversity were highest in populations from China, Indonesia, and Brazil. In contrast, populations from Oman and Pakistan collected from Mangifera had the lowest genetic diversity and were clonal. Molecular variance, population differentiation, and network and structure analyses showed that the genetic structure of isolates in the C. manginecans complex is influenced by both host association as well as geographical isolation. Furthermore, the results reflected the movement of genotypes between plant hosts and geographic regions that have implications regarding the broad global distribution of this pathogen.

The European spruce bark beetle (Ips typographus) has become a major forest pest in Finland in recent years. The beetle is a well-known vector of mainly ophiostomatoid fungi causing blue-stain of timber and pathogens that have the ability to amplify the insect damage. It also vectors other associated organisms, such as phoretic mites. The ecology of these mites remains poorly understood, including their associations with fungi. In this study, we considered filamentous fungi and yeasts associated with mites phoretic on I. typographus. Fungal identifications were based on DNA sequences and phylogenetic analyses of the ITS and/or partial β-tubulin gene regions. Fifteen fungal species were detected, including eight yeasts and seven filamentous fungi. Eleven percent of the beetles carried mites and of these 74% carried at least one fungal species. An average of two fungal species were carried per mite. The most commonly found filamentous fungi were Grosmannia penicillata (25%), Ophiostoma bicolor (19%), O. ainoae (12%) and O. brunneolum (12%). Of the yeast species, the most commonly found was Wickerhamomyces bisporus (47%). This study is the first to report yeasts associated with I. typographus and its phoretic mites in Finland. Majority of the filamentous fungal species found are those previously reported in association with I. typographus. The results also confirmed that many of the fungal species commonly found on I. typographus are also associated with its phoretic mites. However, the nature of the symbiosis between the mites, beetles and fungal associates remains to be understood.

Recent reclassification of the Ceratocystidaceae (Microascales) based on multi-gene phylogenetic inference has shown that the oak wilt fungus Ceratocystis fagacearum does not reside in any of the four genera in which it has previously been treated. In this study, we resolve typification problems for the fungus, confirm the synonymy of Chalara quercina (the first name applied to the fungus) and Endoconidiophora fagacearum (the name applied when the sexual state was discovered). Furthermore, the generic placement of the species was determined based on DNA sequences from authenticated isolates. The original specimens studied in both protologues and living isolates from the same host trees and geographical area were examined and shown to represent the same species. A lectotype was designated for Chalara quercina and Endoconidiophora fagacearum and an epitype linked to a living ex-epitype isolate was designated. Phylogenetic analyses confirmed that the species resides in a well-supported monophyletic lineage in the Ceratocystidaceae, distinct from all other genera in the family. The new genus Bretziella is described to accommodate the oak wilt fungus.

Subsequence join in time series is to search for couples of similar subsequences from multiple time series. The task is useful in data mining on time series; nevertheless, it is extremely difficult because of its enormous computational cost. The task should use normalized time series during the search execution and be performed under an efficient distance measure to obtain accurate resulting couples. The task is more challenging when it works in a streaming environment in which time-series data might be collected very quickly. To address this problem, we propose an efficient method of subsequence join in streaming time series under Dynamic Time Warping (DTW), supporting z-score normalization. The proposed method utilizes a technique of subsequence extraction based on major extrema of streaming time series to search for couples of similar subsequences from coevolving time series. This method can identify couples of similar subsequences of the same length or different lengths. The experimental results show that the proposed method has high performance and can bring out interesting couples of similar subsequences. In addition, this method acts as an approximation algorithm suitable for a streaming scenario where users often expect fast responses from the task of subsequence join over time-series streams of high rates. [ABSTRACT FROM AUTHOR]