The filamentous fungus Arcopilus aureus (Sordariale: Chaetomiaceae) is notable for its multi-domain significance across agriculture, medicine, and industry. In this study, we generated a chromosome-level nuclear genome and a complete circular mitogenome for A. aureus by integrating data from next-generation sequencing, PacBio HiFi, and Hi-C technologies. The final nuclear genome assembly spans 33.77 Mb (GC content: 57.67%), and was organized into seven chromosomal-sized scaffolds (only one gap) with an N50 size of 5.09 Mb and BUSCO completeness of 95.91%. A total of 10,282 protein-coding genes, 228 non-coding RNAs, and ~1.77 Mb of repetitive elements were predicted in the nuclear genome. By contrast, the mitogenome of A. aureus is 33,820 bp in length, with a GC content of 25.96%. It harbors 15 typical mitochondrial protein-coding genes, one unidentified ORF, two rRNAs (small subunit rns and large subunit rnl), and 28 tRNAs. This high-quality genome assembly provides a valuable resource for understanding the ecology, genetics, and evolution of A. aureus, which facilitates elucidating its mechanisms of biocontrol, infection, and metabolite synthesis.

Gpi7 gene, encoding the catalytic subunit of GPI ethanolamine-phosphate (Etn-P) transferase II, is primarily involved in the synthesis, maturation, and sorting of GPI-anchored proteins (GPI-APs), thereby playing a crucial part in cell wall functions and host-pathogen interactions. This study aimed to investigate the role of Gpi7 (here designated CcGpi7) in Corynespora cassiicola, a devastating fungal pathogen causing leaf spot in cucumber and many other cash crops. We systematically identified and inventoried 138 GPI-APs in C. cassiicola, followed by a detailed structural characterization of the CcGpi7 protein that is strongly induced during infection. Homologous recombination was employed to construct a CcGpi7-deleted mutant (ΔCcGpi7) and its corresponding complementary strain (cCcGpi7). Compared with the wild type and cCcGpi7, deletion of CcGpi7 led to markedly reduced vegetative growth and conidia formation. The ΔCcGpi7 mutant displayed obvious defects in cell wall architecture, manifested as enhanced susceptibility to cell wall-perturbing agents and degrading enzymes. Under stress conditions, ΔCcGpi7 exhibited increased sensitivity to KCl but reduced sensitivity to sorbitol and H2O2. Pathogenicity assays revealed a dramatic attenuation in the virulence of ΔCcGpi7 on cucumber leaves, directly correlating with its impaired ability to form invasive hyphae. Transcriptome profiling identified a total of 3,604 differentially expressed genes in ΔCcGpi7, which were enriched in multiple processes including cellular growth and development, cell wall organization, sporulation, and unexpectedly, transcription and translation. Together, our findings demonstrate that CcGpi7 exerts pleotropic effects on vegetative growth, reproduction, cell wall integrity, and pathogenesis of C. cassiicola. This work lays a theoretical foundation for developing CcGpi7-targeted control strategies against cucumber target spot disease.

Bipolaris maydis, the fungus responsible for southern corn leaf blight, seriously impacts maize yield and quality worldwide. In this study, a rapid diagnostic method that targets the EF-1 alpha gene of B. maydis, combining recombinase polymerase amplification (RPA) with CRISPR/Cas12a detection at a constant 37 degrees C. The entire process takes about 15 min, comprising 5 min for RPA amplification and 10 min for CRISPR/Cas12a-mediated detection. Based on the one-tube RPA-CRISPR/Cas12a platform, results can be visualized using lateral flow test strips (LFS) and fluorescent-based methods. The assay demonstrated high specificity for B. maydis and improved sensitivity, detecting as few as 8 copies using fluorescence, with a visual detection range of 8-80 copies on test strips. This represents a 10-fold increase in sensitivity compared to conventional real-time PCR. The method successfully identified the pathogen in maize leaves as early as two days after inoculation. In conclusion, this one-tube RPA-CRISPR/Cas12a platform provides an effective method for B. maydis detection with significant potential for on-site applications in field settings.

Background/Objectives: Wheat root and crown rot, caused by Fusarium pseudograminearum, Fusarium graminearum, and Bipolaris sorokiniana, are devastating soil-borne diseases that cause substantial yield losses worldwide. Endophytic fungi are promising sources of bioactive metabolites for agricultural applications. This study aimed to isolate and characterize an endophytic fungus with antifungal activity against major wheat pathogens, identify its active compound, and investigate the underlying transcriptional response. Methods: An endophytic strain Y2 was isolated from Hedyotis diffusa leaves and identified through morphological and phylogenetic analysis based on TEF-1α and RPB2 sequences. Pathogenicity of strain Y2 was evaluated on wheat stem bases. The bioactive compound was purified by HPLC and identified by HR-ESI-MS and NMR. Antifungal activity was assessed using dual-culture and microbroth dilution assays. Transcriptomic analysis (RNA-seq) was performed on F. pseudograminearum treated with equisetin, with qRT-PCR validation of seven representative differentially expressed genes. Results: Strain Y2 was identified as Fusarium incarnatum or a closely related member of the F. incarnatum-equiseti species complex (FIESC) and confirmed to be non-pathogenic to wheat. The purified bioactive compound was characterized as equisetin, which exhibited significant antifungal activity with MIC values of 16, 32, and 64 μg/mL against F. pseudograminearum, B. sorokiniana, and F. graminearum, respectively. Transcriptomic analysis revealed that equisetin treatment induced a polarized transcriptional response in F. pseudograminearum, characterized by strong upregulation of ribosome and translation-related genes and widespread downregulation of other metabolic pathways, particularly nitrogen metabolism. qRT-PCR validation of seven representative genes confirmed the reliability of the RNA-seq data. Conclusions: Our findings demonstrate that equisetin is the active antifungal metabolite produced by F. incarnatum Y2, with potent in vitro activity against major wheat root and crown rot pathogens. The transcriptomic data provide insights into the potential mechanism of action, while the non-pathogenic nature of strain Y2 supports its biosafety. Although these results highlight equisetin as a promising lead compound for antifungal development, further in planta efficacy and safety studies are required before it can be considered for practical biocontrol.

Bipolaris sorokiniana, a pathogenic fungus responsible for root rot, crown rot, leaf spot, and black point in wheat, poses a serious threat to both yield and grain quality. To investigate its pathogenic mechanisms, we identified the BsSYT1 gene with the Sec7 domain by amplifying the flanking sequence of a T-DNA insertion mutant, BSK-353—which exhibited markedly reduced mycelial growth and attenuated virulence—from a B. sorokiniana mutant library using hiTAIL-PCR. Expression of BsSYT1 was significantly upregulated during the infection stage compared to the mycelial stage. Deletion of BsSYT1 impaired hyphal growth but unexpectedly enhanced conidiation. Moreover, the ΔBsSYT1 mutant exhibited significantly enhanced tolerance to osmotic, oxidative, and cell wall stresses compared to the wild-type strain. Virulence assays demonstrated that while the loss of BsSYT1 did not affect appressorium formation, it drastically reduced lesion development on wheat and barley leaves. Furthermore, we identified BsSYT1 as a functional target of the mycotoxin brefeldin A. Collectively, our results establish that BsSYT1 is a critical regulator of multiple biological processes in B. sorokiniana, including growth, development, stress responses, and virulence.

Here, we describe a novel ourmia-like virus, Botryosphaeria dothidea ourmia-like virus 3 (BdOLV3), obtained from the phytopathogenic fungus Botryosphaeria dothidea strain ZM211299-2 infecting apple shoots in Shandong province of China. The genome of BdOLV3 is a positive-sense single-stranded RNA with a length of 2,650 nucleotides (nt), containing a large open reading frame (ORF) that encodes a putative RNA-dependent RNA polymerase (RdRp) consisting of 673 amino acids (aa) with a molecular mass of 75.53 kDa. This RdRp protein contains eight typical conserved motifs associated with ourmia-like viruses. BLASTp analysis revealed that the RdRp protein of BdOLV3 had the highest similarity (61.98%, 61.83%, and 51.47% identity, respectively) to viruses previously identified as Plasmopara viticola lesion associated ourmia-like virus 45, Erysiphe necator associated ourmia-like virus 130, and Magnaporthe oryzae botourmiavirus 7. Phylogenetic analysis based on RdRp sequences indicated that BdOLV3 is a new member of the genus Epsilonscleroulivirus in the family Botourmiaviridae. This is the first report of the complete genomic sequence of a member of the genus Epsilonscleroulivirus found in B. dothidea.

Natural essential oils (EOs) are recognized as promising bioactive agents for the development of biopesticides. Herein, five EOs were encapsulated into oil-in-water nanoemulsions using a one-pot ultrasonic cavitational emulsification approach. The physicochemical properties of the nanoemulsions were characterized and an optimal nanoscale size was achieved at ultrasonication of 280 W for 5 min using 1% natural tea saponin as emulsifier within ice-bath. All nanoemulsions exhibited both contact and fumigant activity against Cryptolestes ferrugineus (Stephens) and higher than that their free form. In comparison, among the five essential oils, garlic (Allium sativum L.) essential oil nanoemulsion exhibited the lowest LC50 of 0.03 mu L/cm2 via contact toxicity against C. ferrugineus (Stephens), whereas cinnamon (Cinnamomum cassia Presl) essential oil nanoemulsion showed the lowest LC50 of 1.16 mu L/L via fumigant toxicity after 48 h of exposure. Considering the high stability of GEO nanoemulsion, it would be the best candidate for controlling C. ferrugineus. These findings suggest that that these nanoemulsion of garlic essential oil preparated by one-pot ultrasonic cavitational emulsification have the potential to be used as a sustainable and eco-friendly pest management application against C. ferrugineus (Stephens) in grain preservation and biopesticides formulations.

Fusarium crown rot (FCR), a cereal disease caused primarily by Fusarium pseudograminearum and other related Fusarium species, poses a major threat to wheat production in many regions worldwide. Driven by climate change and suboptimal farming practices, FCR has been spreading rapidly across China, with the affected area expanding from 2.66 million hectares in 2022 to approximately 4 million hectares in 2024. This rapid expansion underscores the urgent need to develop effective and sustainable management strategies. This review comprehensively summarizes the research progress in several key areas, including the evaluation and measurement of FCR severity, the identification of resistant germplasm, the discovery of quantitative trait loci (QTLs) and genes conferring FCR resistance, and the elucidation of their underlying mechanisms. Numerous methods for assessing FCR have been developed; however, their effectiveness in predicting field-level resistance or tolerance varies significantly. Screening over 3,000 wheat genotypes revealed that only 7% exhibit moderate FCR resistance, highlighting a critical shortage of highly resistant varieties. Resistance-associated genes or QTLs have been identified across all 21 wheat chromosomes. Their mechanisms primarily involve enhancing physical defenses through cell wall composition modification, improving reactive oxygen species (ROS) scavenging capacity, facilitating toxin detoxification, activating the expression of defense-related genes, and boosting the accumulation of protective metabolites. FCR resistance is generally non-specific to pathogen species and tends to be durable. Although fully immune cultivars may be unattainable, developing lines with high levels of resistance is feasible. Therefore, future research should prioritize the identification and characterization of novel resistance sources, and utilization of those with high breeding potential to develop cultivars with enhanced FCR resistance. This review offers valuable insights for improving FCR evaluation and measurement, and demonstrates that pyramiding well-characterized resistance QTLs, combined with genomics selection, constitutes an effective strategy for enhancing FCR resistance in wheat.

Botryosphaeria dothidea is a prevalent pathogen of woody plants with a global distribution. In this study, we identified a novel mycovirus from the B. dothidea strain ZM200473, which we have tentatively designated "Botryosphaeria dothidea non-segmented dsRNA virus" (BdNSRV1). The genome of BdNSRV1 is composed of dsRNA that spans 2,902 base pairs and contains two non-overlapping open reading frames (ORF1 and ORF2). ORF1 encodes a hypothetical protein consisting of 316 amino acids with a molecular weight of 35.0 kDa, and this protein shares similarity to the coat proteins of several mycoviruses. ORF2 encodes a protein of 561 amino acids with a molecular weight of 66.0 kDa, which includes a conserved RNA-dependent RNA polymerase (RdRp). Sequence comparisons and phylogenetic analysis indicated that BdNSRV1 establishes a well-supported independent clade alongside members of the recently established genus Unirnavirus, being most closely related to Lasiodiplodia pseudotheobromae mycovirus 1 (LpMyV1), with 70.74% amino acid sequence identity in the RdRp. Therefore, BdNSRV1 should be classified as a novel non-segmented dsRNA mycovirus of the genus Unirnavirus and is the first characterized non-segmented virus associated with B. dothidea.

Here, we describe a novel ourmia-like virus, Botryosphaeria dothidea ourmia-like virus 3 (BdOLV3), derived from the phytopathogenic fungus Botryosphaeria dothidea strain ZM211299-2 infecting apple shoots in Shandong province of China. The complete genome sequence of BdOLV3 consists of a positive-sense single-stranded RNA (+ ssRNA) segment with a length of 2,650 nucleotides (nt). The sequence contains a large open reading frame (ORF) encoding a putative RNA-dependent RNA polymerase (RdRp) consisting of 673 amino acids (aa) with a molecular mass of 75.53 kDa. This RdRp protein contains eight typical conserved motifs associated with ourmia-like viruses. BLASTp analysis revealed that the RdRp protein of BdOLV3 had the highest similarity (61.98%, and 51.47% identity) to viruses previously identified as Plasmopara viticola lesion associated ourmia-like virus 45, and Magnaporthe oryzae botourmiavirus 7, respectively. Phylogenetic analysis based on the RdRp aa sequence indicated that BdOLV3 is a new member of the genus Epsilonscleorulivirus in the family Botourmiaviridae . This study is the first to report the complete genomic sequence of the genus Epsilonscleorulivirus in B. dothidea .

In this study, the complete mitogenomes of three Diaporthe species (Diaportheeres ZM79-3, D.phaseolorum ZM33-4 and Diaporthe sp. ZM41-5) were sequenced, assembled and compared with the other three previously sequenced Diaporthe mitogenomes (D.caulivora VNIIKR SE Dcaul3, D.longicolla MSPL 10-6 and D.sojae VNIIKR SE Dps12). The six Diaporthe mitogenomes were found to be circular DNA molecules, with lengths ranging from 53,646 bp to 108,865 bp. The mitogenomes of the six Diaporthe species mainly comprised the same set of 15 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 15 PCGs in the six mitogenomes. The nad1 gene had the least K2P genetic distance of the 15 core PCGs among the 13 Diaporthales species, indicating that this gene was highly conserved. The Ka/Ks values for all 15 core PCGs were < 1, suggesting that these genes were all subject to purifying selection. Comparative mitogenome analysis showed that introns contributed the most to the size variation of Diaporthe mitogenomes. Frequent intron loss/gain events were detected to have occurred in the cox1 gene during the evolution of the Diaporthales mitogenomes. Although the mitogenomes of 13 species from Diaporthales had undergone large-scale gene rearrangements, six mitogenomes of Diaporthe species had identical gene arrangements. Phylogenetic analysis based on combined mitochondrial gene datasets showed that the six Diaporthe species formed well-supported topologies. To our knowledge, this study is the first report on the mitogenomes of D.phaseolorum ZM33-4 and Diaporthe sp. ZM41-5, as well as the first comparison of mitogenomes among Diaporthe species. Our findings will further promote investigations of the genetics, evolution and phylogeny of the Diaporthe species.

Apple Valsa canker, caused by Valsa mali and Valsa pyri, is a devastating disease of apple trees and poses a severe threat to the sustainable development of apple production. Although the two species’ whole genomes have been sequenced, their mitochondrial genomes are still uncharacterized. In this study, the complete mitochondrial genomes of V. mali and V. pyri were assembled, annotated, and compared by bioinformatic methods. The results indicate that the mitogenomes are both circular DNA molecules with sizes of 213,406 bp and 128,022 bp, respectively. The AT skew values of the two Valsa species’ mitogenomes were positive, while the GC skew values were negative. Comparative mitogenome analysis revealed that the length and base composition of protein-coding genes (PCGs), rRNA genes, and tRNA genes differed between the two Valsa species. It was found that the expansion of V. mali was primarily attributable to the intronic regions. There are large numbers of interspersed repetitive sequences (IRS) in both Valsa mitogenomes; however, the proportion of IRS in V. mali (43.56%) was much higher than that in V. pyri (2.41%). The alignment of large fragments between the mitochondrial and nuclear genomes of both V. mali (1.73 kb) and V. pyri (5.17 kb) indicates that gene transfer between mitochondrial and nuclear genomes occurred during evolution. The ka/ks ratios for 15 core PCGs were below one, suggesting that these genes were subjected to purifying selection pressure. Comparative mitogenomics revealed that the two fungi had significant mitogenomic collinearity and large-scale gene rearrangements. The results of phylogenetic analysis based on Bayesian inference (BI) and maximum likelihood (ML) using a combined mitochondrial gene set confirmed that V. mali and V. pyri were fully independent taxa with a high bootstrap value of 100 (ML) and a high posterior probability of 1.0 (BI). This is the first report on the mitogenomes within the genus Valsa. These results will pave the way to understanding the evolution and differentiation of mitogenomes in the genus Valsa.

Here, a novel dsRNA virus belonging to the family Polymycoviridae was identified in the phytopathogenic fungal strain B. dothidea ZM200473 and tentatively named "Botryosphaeria dothidea polymycovirus 2" (BdPmV2). The genome of BdPmV2 consists of five genomic dsRNA segments, ranging in size from 1224 to 2407 bp, each containing an open reading frame (ORF). ORF1 encodes a conserved RNA-dependent RNA polymerase (RdRP) consisting of 763 amino acids (aa) with a molecular mass (Mr) of 84.03 kDa, ORF3 encodes a putative methyltransferase (Met) consisting of 627 amino acids (aa) with an Mr of 68.41 kDa, ORF4 encodes a P-A-S-rich protein that serves as a coat protein (CP) consisting of 261 amino acids (aa) with an Mr of 27.60 kDa. ORF2 and ORF5 encode putative proteins with unknown functions, consisting of 697 amino acids (aa) with an Mr of 76.49 kDa and 323 amino acids (aa) with an Mr of 33.92 kDa, respectively. BLASTp analysis revealed that the RdRP protein of BdPmV2 had the highest similarity (56.52% identity) to that of a virus previously identified as "Aspergillus fumigatus polymycovirus 1". Phylogenetic analysis based on RdRP aa sequences indicated that BdPmV2 is a new member of the family Polymycoviridae.

A novel double-stranded RNA (dsRNA) virus was isolated and described from strain ZZZ210557 of plant endophyte Allocryptovalsa sichuanensis. The dsRNA virus contains four dsRNA segments, dsRNA1 to dsRNA4, with a size range of 3.8 to 5.1 kbp. Each possesses a single large ORF and is encapsulated in isometric particles approximately 42-47 nm in diameter. Notably, the dsRNA3 encoded sequence revealed modest similarities to the amino acid sequences of RdRps predicted from the nucleotide sequences of known and suspected members of the family Quadriviridae. Phylogenetic analysis of the putative RdRp with the corresponding proteins of other quadriviruses revealed that the dsRNA virus is a new member belonging to the family Quadriviridae, tentatively named Allocryptovalsa sichuanensis quadrivirus 1 (AsQV1). All four segments of AsQV1 could be successfully cured through ribavirin treatment, whereas it likely has no apparent impact on the morphologies or virulence of the host fungus. This study is the first report of a quadrivirus isolated from the fungus A. sichuanensis, and our results enhance the diversity of the quadrivirus.

Here, a novel mycovirus, Botryosphaeria dothidea narnavirus 5 (BdNV5), was discovered in the plant-pathogenic fungus Botryosphaeria dothidea strain ZM210167-1. The BdNV5 genome sequence is 2,397 nucleotides (nt) in length and contains a putative open reading frame (ORF) encoding an RNA-dependent RNA polymerase (RdRp) with a molecular mass of 72.77 kDa. A BLASTp search using the RdRp amino acid (aa) sequence showed that it was most similar to the RdRp of Botryosphaeria dothidea narnavirus 4 (42.35%). In a phylogenetic tree based on RdRp aa sequences, BdNV5 clustered with members of the family Narnaviridae. BdNV5 is thus a novel member of the family Narnaviridae infecting the phytopathogenic fungus B. dothidea.

Background 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. Results 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. Conclusions 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.

Laccases are the key enzymes responsible for plant lignin biosynthesis and responses to environment stress. However, the roles of LAC genes in plant disease resistance are still largely unknown, especially in grapevine, one of the most important horticultural crops in the world. Its quality and yield are very vulnerable to gray mold disease caused by Botrytis cinerea. In total, 30 VvLAC genes were identified and found to be unevenly distributed on seven chromosomes; they were classified into seven groups based on phylogenetic analysis according to the criteria applied in Arabidopsis thaliana. Collinearity and synteny analyses identified some orthologous gene pairs in Vitis vinifera and a few paralogous gene pairs among grape and peach. The VvLAC gene family has diverse gene structures and a highly conserved motif composition. The prominent presence of the MYB cis-elements in each VvLAC promoter highlighted MYB transcriptional factors as the main regulators of VvLAC genes. Furthermore, twenty-five VvLAC genes with functional redundancy are probably implicated in grape lignin biosynthesis. The expression patterns of the LAC genes in grape leaves of Chinese wild V. amurensis ‘Shuangyou’ (SY), a germplasm highly resistant to B. cinerea, were investigated through transcriptomic data and qRT-PCR verification. Combined with the phylogenetic analysis, with AtLACs participating in lignin metabolism, and the cis-element analysis, VaLAC14, VaLAC19, VaLAC24 and VaLAC30 were identified as key candidate genes for lignin biosynthesis in the grape response to B. cinerea. This study supplies a comprehensive understanding of the classification, evolution, structure and responses of the grape LAC genes against B. cinerea. It also provides valuable genetic resources for functional characterization towards enhancing grapevine disease resistance.

Lasiodiplodia is a widely distributed genus that is associated with a variety of diseases in many plant species, especially fruit trees. In this study, a disease survey of fruit trees growing in 12 orchards located in the Henan and Shandong provinces of China was conducted between 2020 and 2022. The symptoms observed included stem canker, branch dieback, and gummosis. Phylogenetic analyses of internal transcribed spacer, tub2, tef1, and rpb2 sequence data combined with morphological characteristics revealed that the 19 isolates collected during the survey belonged to five documented Lasiodiplodia species, namely, Lasiodiplodia citricola, L. chiangraiensis, L. huangyanensis, L. pseudotheobromae, and L. theobromae, and two previously undescribed species, L. xinyangensis and L. ziziphi. In addition, the survey identified three novel host-pathogen interactions: L. chiangraiensis on loquat, L. citricola on apple, and L. huangyanensis on grapevine. Furthermore, the detailed phylogenic analysis indicated that four previously described Lasiodiplodia species were genetically very closely related that they would be better classified as synonyms rather than distinct species, so L. paraphysoides and L. nanpingensis should be considered synonyms of L. citricola, L. fujianensis should be a synonym of L. iraniensis, and L. henanica should be a synonym of L. huangyanensis. Pathogenicity tests confirmed that representative isolates of the two novel species and three new host-pathogen interactions identified in the current study were pathogenic to their original hosts, thereby fulfilling Koch's postulates. Similarly, all of the isolates were found to be pathogenic on four alternative hosts, although a high degree of variation in virulence was observed.

In 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.