The six whole mitochondrial genomes for the Diaporthe species: features, evolution and phylogeny

Gao W., Cao J., Xie Y., Sun X., Ma Q., Geng Y., Xu C., Guo Y., Zhang M.

Maple is an important ornamental plant in China. With the increasing use of maple trees in landscaping, a symptom of shoot dieback has been observed in Henan province, China. In this study, 28 Diaporthe isolates were obtained from symptomatic shoots of maple trees between 2020 and 2023. Phylogenetic analyses based on five loci (ITS, TEF, CAL, HIS and TUB) coupled with morphology of 12 representative isolates identified three known species (D. eres, D. pescicola and D. spinosa) and one new species, namely D. pseudoacerina sp. nov. Koch’s postulates confirmed that all these species were pathogenic. Additionally, D. pseudoacerina was able to infect China wingnut (Pterocarya stenoptera), pear (Pyrus sp.), and black locust (Robinia pseudoacacia). This study marks the first report of Diaporthe spinosa and D. pescicola pathogens infecting maple trees. These findings enhance the existing knowledge of the taxonomy and host diversity of Diaporthe species as, while also providing valuable information for managing of maple shoot dieback in Henan Province, China.

Li S., Hu X., Song Q.

Diaporthe longicolla (syn. Phomopsis longicolla) is an important seed-borne fungal pathogen and the primary cause of Phomopsis seed decay (PSD) in soybean. PSD is one of the most devastating seed diseases, reducing soybean seed quality and yield worldwide. As part of a genome sequencing project on the fungal Diaporthe–Phomopsis complex, draft genomes of eight D. longicolla isolates were sequenced and assembled. Sequences of mitochondrial genomes were extracted and analyzed. The circular mitochondrial genomes ranged from 52,534 bp to 58,280 bp long, with a mean GC content of 34%. A total of 14 core protein-coding genes, 23 tRNA, and 2 rRNA genes were identified. Introns were detected in the genes of atp6, cob, cox1, cox2, cox3, nad1, nad2, nad5, and rnl. Three isolates (PL7, PL10, and PL185E) had more introns than other isolates. Approximately 6.4% of the mitochondrial genomes consist of repetitive elements. Moreover, 48 single-nucleotide polymorphisms (SNPs) and were identified. The mitochondrial genome sequences of D. longicolla will be useful to further study the molecular basis of seed-borne pathogens causing seed diseases, investigate genetic variation among isolates, and develop improved control strategies for Phomopsis seed decay of soybean.

Dissanayake A.J., Zhu J., Chen Y., Maharachchikumbura S.S., Hyde K.D., Liu J.

AbstractDiaporthe is an important plant pathogenic genus, which also occurs as endophytes and saprobes. Many Diaporthe species that are morphologically similar proved to be genetically distinct. The current understanding of Diaporthe taxonomy by applying morphological characters, host associations and multi-gene phylogeny are problematic leading to overestimation/underestimation of species numbers of this significant fungal pathogenic genus. Currently, there are no definite boundaries for the accepted species. Hence, the present study aims to re-structure the genus Diaporthe, based on single gene phylogenies (ITS, tef, tub, cal and his), multi-gene phylogeny justified by applying GCPSR (Genealogical Concordance Phylogenetic Species Recognition) methodology as well as the coalescence-based models (PTP—Poisson Tree Processes and mPTP—multi-rate Poisson Tree Processes). Considering all available type isolates of Diaporthe, the genus is divided into seven sections while boundaries for 13 species and 15 species-complexes are proposed. To support this re-assessment of the genus, 82 Diaporthe isolates obtained from woody hosts in Guizhou Province in China were investigated and revealed the presence of two novel species and 17 previously known species. Synonymies are specified for 31 species based on molecular data and morphological studies. Dividing Diaporthe into several specific sections based on phylogenetic analyses can avoid the construction of lengthy phylogenetic trees of the entire genus in future taxonomic studies. In other words, when one conducts research related to the genus, only species from the appropriate section need to be selected for phylogenetic analysis.

Song X., Geng Y., Xu C., Li J., Guo Y., Shi Y., Ma Q., Li Q., Zhang M.

AbstractIn the present study, three mitogenomes from the Bipolaris genus (Bipolaris maydis, B. zeicola, and B. oryzae) were assembled and compared with the other two reported Bipolaris mitogenomes (B. oryzae and B. sorokiniana). The five mitogenomes were all circular DNA molecules, with lengths ranging from 106,403 bp to 135,790 bp. The mitogenomes of the five Bipolaris species mainly comprised the same set of 13 core protein-coding genes (PCGs), two rRNAs, and a certain number of tRNAs and unidentified open reading frames (ORFs). The PCG length, AT skew and GC skew showed large variability among the 13 PCGs in the five mitogenomes. Across the 13 core PCGs tested, nad6 had the least genetic distance among the 16 Pleosporales species we investigated, indicating that this gene was highly conserved. In addition, the Ka/Ks values for all 12 core PCGs (excluding rps3) were < 1, suggesting that these genes were subject to purifying selection. Comparative mitogenomic analyses indicate that introns were the main factor contributing to the size variation of Bipolaris mitogenomes. The introns of the cox1 gene experienced frequent gain/loss events in Pleosporales species. The gene arrangement and collinearity in the mitogenomes of the five Bipolaris species were almost highly conserved within the genus. Phylogenetic analysis based on combined mitochondrial gene datasets showed that the five Bipolaris species formed well-supported topologies. This study is the first report on the mitogenomes of B. maydis and B. zeicola, as well as the first comparison of mitogenomes among Bipolaris species. The findings of this study will further advance investigations into the population genetics, evolution, and genomics of Bipolaris species.

Anwar G., Mamut R., Wang J.

In the present study, the complete mitochondrial genomes of five Peltigera species (Peltigera elisabethae, Peltigera neocanina, Peltigera canina, Peltigera ponojensis, Peltigera neckeri) were sequenced, assembled and compared with relative species. The five mitogenomes were all composed of circular DNA molecules, and their ranged from 58,132 bp to 69,325 bp. The mitochondrial genomes of the five Peltigera species contain 15 protein-coding genes (PCGs), 2 rRNAs, 26–27 tRNAs and an unidentified open reading frame (ORF). The PCG length, AT skew and GC skew varied among the 15 PCGs in the five mitogenomes. Among the 15 PCGs, cox2 had the least K2P genetic distance, indicating that the gene was highly conserved. The synteny analysis revealed that the coding regions were highly conserved in the Peltigera mitochondrial genomes, but gene rearrangement occurred in the intergenic regions. The phylogenetic analysis based on the 14 PCGs showed that the 11 Peltigera species formed well-supported topologies, indicating that the protein-coding genes in the mitochondrial genome may be used as a reliable molecular tool in the study of the phylogenetic relationship of Peltigera.

Monkai J., Hongsanan S., Bhat D.J., Dawoud T.M., Lumyong S.

During our investigations of the microfungi on medicinal plants in Thailand, five isolates of Diaporthe were obtained. These isolates were identified and described using a multiproxy approach, viz. morphology, cultural characteristics, host association, the multiloci phylogeny of ITS, tef1-α, tub2, cal, and his3, and DNA comparisons. Five new species, Diaporthe afzeliae, D. bombacis, D. careyae, D. globoostiolata, and D. samaneae, are introduced as saprobes from the plant hosts, viz. Afzelia xylocarpa, Bombax ceiba, Careya sphaerica, a member of Fagaceae, and Samanea saman. Interestingly, this is the first report of Diaporthe species on these plants, except on the Fagaceae member. The morphological comparison, updated molecular phylogeny, and pairwise homoplasy index (PHI) analysis strongly support the establishment of novel species. Our phylogeny also revealed the close relationship between D. zhaoqingensis and D. chiangmaiensis; however, the evidence from the PHI test and DNA comparison indicated that they are distinct species. These findings improve the existing knowledge of taxonomy and host diversity of Diaporthe species as well as highlight the untapped potential of these medicinal plants for searching for new fungi.

Li Q., Xiao W., Wu P., Zhang T., Xiang P., Wu Q., Zou L., Gui M.

AbstractApiotrichum is a diverse anamorphic basidiomycetous yeast genus, and its mitogenome characterization has not been revealed. In this study, we assembled two Apiotrichum mitogenomes and compared them with mitogenomes from Agaricomycotina, Pucciniomycotina and Ustilaginomycotina. The mitogenomes of Apiotrichum gracile and A. gamsii comprised circular DNA molecules, with sizes of 34,648 bp and 38,096 bp, respectively. Intronic regions were found contributed the most to the size expansion of A. gamsii mitogenome. Comparative mitogenomic analysis revealed that 6.85–38.89% of nucleotides varied between tRNAs shared by the two Apiotrichum mitogenomes. The GC content of all core PCGs in A. gamsii was lower than that of A. gracile, with an average low value of 4.97%. The rps3 gene differentiated the most among Agaricomycotina, Pucciniomycotina and Ustilaginomycotina species, while nad4L gene was the most conserved in evolution. The Ka/Ks values for cob and rps3 genes were > 1, indicating the two genes may be subjected to positive selection in Agaricomycotina, Pucciniomycotina and Ustilaginomycotina. Frequent intron loss/gain events and potential intron transfer events have been detected in evolution of Agaricomycotina, Pucciniomycotina and Ustilaginomycotina. We further detected large-scale gene rearrangements between the 19 mitogenomes from Agaricomycotina, Pucciniomycotina and Ustilaginomycotina, and fifteen of the 17 mitochondrial genes shared by Apiotrichum varied in gene arrangements. Phylogenetic analyses based on maximum likelihood and Bayesian inference methods using a combined mitochondrial gene dataset revealed different taxonomic assignment of two Apiotrichum species, wherein A. gamsii had a more closely relationship with Trichosporon asahii. This study served as the first report on mitogenomes from the genus Apiotrichum, which promotes the understanding of evolution, genomics, and phylogeny of Apiotrichum.

Zhao X., Li K., Zheng S., Yang J., Chen C., Zheng X., Wang Y., Ye W.

Many species in the fungal Diaporthe (anamorph Phomopsis) genus have become a group of the most important pathogens that cause seed decay, stem and pot blight, and stem canker in soybean production worldwide, resulting in significant yield loss. Due to increased disease prevalence but a lack of research, we performed an extensive field survey to isolate and identify the Diaporthe species associated with soybean stem blight in six provinces of China between 2019 and 2020. A total of 92 Diaporthe isolates were identified based on morphological and multilocus phylogenetic analysis and classified into six species: D. longicolla, D. unshiuensis, D. sojae, D. caulivora, D. tectonigena, and an unknown Diaporthe sp. The most frequently identified species was D. longicolla with 57 isolates. High genetic diversity was observed for the D. longicolla isolates, and haplotype network analysis revealed a mixed structure among the population in the six provinces. In comparative pathogenicity assays, different virulence levels were observed among the 92 Diaporthe isolates. The results of this study provide new insights into the Diaporthe spp. associated with soybean stem blight in China and can help in the development of effective management strategies.

Zhang Z.

KaKs_Calculator 3.0 is an updated toolkit that is capable of calculating selective pressure on both coding and non-coding sequences. Similar to the nonsynonymous/synonymous substitution rate ratio for coding sequences, selection on non-coding sequences can be quantified as the ratio of non-coding nucleotide substitution rate to synonymous substitution rate of adjacent coding sequences. As testified on empirical data, KaKs_Calculator 3.0 shows effectiveness to detect the strength and mode of selection operated on molecular sequences, accordingly demonstrating its great potential to achieve genome-wide scan of natural selection on diverse sequences and identification of potentially functional elements at a whole-genome scale. The package of KaKs_Calculator 3.0 is freely available for academic use only at https://ngdc.cncb.ac.cn/biocode/tools/BT000001.

Zou H., Chen F., Li W., Li M., Lei H., Zhang D., Jakovlić I., Wang G.

Within the class Enoplea, the earliest-branching lineages in the phylum Nematoda, the relatively highly conserved ancestral mitochondrial architecture of Trichinellida is in stark contrast to the rapidly evolving architecture of Dorylaimida and Mermithida. To better understand the evolution of mitogenomic architecture in this lineage, we sequenced the mitogenome of a fish parasite Pseudocapillaria tomentosa (Trichinellida: Capillariidae) and compared it to all available enoplean mitogenomes. P. tomentosa exhibited highly reduced noncoding regions (the largest was 98 bp), and a unique base composition among the Enoplea. We attributed the latter to the inverted GC skew (0.08) in comparison to the ancestral skew in Trichinellidae (-0.43 to -0.37). Capillariidae, Trichuridae and Longidoridae (Dorylaimida) generally exhibited low negative or low positive skews (-0.1 to 0.1), whereas Mermithidae exhibited fully inverted low skews (0 to 0.05). This is indicative of inversions in the strand replication order or otherwise disrupted replication mechanism in the lineages with reduced/inverted skews. Among the Trichinellida, Trichinellidae and Trichuridae have almost perfectly conserved architecture, whereas Capillariidae exhibit multiple rearrangements of tRNA genes. In contrast, Mermithidae (Mermithida) and Longidoridae (Dorylaimida) exhibit almost no similarity to the ancestral architecture. Longidoridae exhibited more rearranged mitogenomic architecture than the hypervariable Mermithidae. Similar to the Chromadorea, the evolution of mitochondrial architecture in enoplean nematodes exhibits a strong discontinuity: lineages possessing a mostly conserved architecture over tens of millions of years are interspersed with lineages exhibiting architectural hypervariability. As Longidoridae also have some of the smallest metazoan mitochondrial genomes, they contradict the prediction that compact mitogenomes should be structurally stable. Lineages exhibiting inverted skews appear to represent the intermediate phase between the Trichinellidae (ancestral) and fully derived skews in Chromadorean mitogenomes (GC skews = 0.18 to 0.64). Multiple lines of evidence (CAT-GTR analysis in our study, a majority of previous mitogenomic results, and skew disruption scenarios) support the Dorylaimia split into two sister-clades: Dorylaimida + Mermithida and Trichinellida. However, skew inversions produce strong base composition biases, which can hamper phylogenetic and other evolutionary studies, so enoplean mitogenomes have to be used with utmost care in evolutionary studies.

Ma Q., Geng Y., Li Q., Cheng C., Zang R., Guo Y., Wu H., Xu C., Zhang M.

Exserohilum turcicum and E. rostratum, two closely related fungal species, are both economically important pathogens but have quite different target hosts (specific to plants and cross-kingdom infection, respectively). In the present study, complete circular mitochondrial genomes of the two Exserohilum species were sequenced and de novo assembled, which mainly comprised the same set of 13 core protein-coding genes (PCGs), two rRNAs, and a certain number of tRNAs and unidentified open reading frames (ORFs). Comparative analyses indicated that these two fungi had significant mitogenomic collinearity and consistent mitochondrial gene arrangement, yet with vastly different mitogenome sizes, 264,948 bp and 64,620 bp, respectively. By contrast with the 17 introns containing 17 intronic ORFs (one-to-one) in the E. rostratum mitogenome, E. turcicum involved far more introns (70) and intronic ORFs (126), which was considered as the main contributing factors of their mitogenome expansion/contraction. Within the generally intron-rich gene cox1, a total of 18 and 10 intron position classes (Pcls) were identified separately in the two mitogenomes. Moreover, 16.16% and 10.85% ratios of intra-mitogenomic repetitive regions were detected in E. turcicum and E. rostratum, respectively. Based on the combined mitochondrial gene dataset, we established a well-supported topology of phylogeny tree of 98 ascomycetes, implying that mitogenomes may act as an effective molecular marker for fungal phylogenetic reconstruction. Our results served as the first report on mitogenomes in the genus Exserohilum, and would have significant implications in understanding the origin, evolution and pathogenic mechanisms of this fungal lineage.

Norphanphoun C., Gentekaki E., Hongsanan S., Jayawardena R., Senanayake I., Manawasinghe I., Abeywickrama P., Bhunjun C., Hyde K.

Abuduaini A., Wang Y., Zhou H., Kang R., Ding M., Jiang Y., Suo F., Huang L.

In this study, the complete mitochondrial genome of O. gracilis was sequenced and assembled before being compared with related species. As the second largest mitogenome reported in the family Ophiocordycipitaceae, the mitogenome of O. gracilis (voucher OG201301) is a circular DNA molecule of 134,288 bp that contains numerous introns and longer intergenomic regions. UCA was detected as anticodon in tRNA-Sec of O. gracilis, while comparative mitogenome analysis of nine Ophiocordycipitaceae fungi indicated that the order and contents of PCGs and rRNA genes were considerably conserved and could descend from a common ancestor in Ophiocordycipitaceae. In addition, the expansion of mitochondrial organization, introns, gene length, and order of O. gracilis were determined to be similar to those of O. sinensis, which indicated common mechanisms underlying adaptive evolution in O. gracilis and O. sinensis. Based on the mitochondrial gene dataset (15 PCGs and 2 RNA genes), a close genetic relationship between O. gracilis and O. sinensis was revealed through phylogenetic analysis. This study is the first to investigate the molecular evolution, phylogenetic pattern, and genetic structure characteristics of mitogenome in O. gracilis. Based on the obtained results, the mitogenome of O. gracilis can increase understanding of the genetic diversity and evolution of cordycipitoid fungi.

Rizali A., Laili Aziz N., Sari N.

Li Q., Li L., Feng H., Tu W., Bao Z., Xiong C., Wang X., Qing Y., Huang W.

In this study, the mitogenome of Hannaella oryzae was sequenced by next-generation sequencing (NGS) and successfully assembled. The H. oryzae mitogenome comprised circular DNA molecules with a total size of 26,444 bp. We found that the mitogenome of H. oryzae partially deleted the tRNA gene transferring cysteine. Comparative mitogenomic analyses showed that intronic regions were the main factors contributing to the size variations of mitogenomes in Tremellales. Introns of the cox1 gene in Tremellales species were found to have undergone intron loss/gain events, and introns of the H. oryzae cox1 gene may have different origins. Gene arrangement analysis revealed that H. oryzae contained a unique gene order different from other Tremellales species. Phylogenetic analysis based on a combined mitochondrial gene set resulted in identical and well-supported topologies, wherein H. oryzae was closely related to Tremella fuciformis. This study represents the first report of mitogenome for the Hannaella genus, which will allow further study of the population genetics, taxonomy, and evolutionary biology of this important phylloplane yeast and other related species.