In this study, the complete mitogenomes of three Diaporthe species (Diaporthe eres 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 phytopathogenic fungal strain B. dothidea ZM200473, and tentatively named “Botryosphaeria dothidea polymycovirus 2” (BdPmV2). The genome of BdPmV2 have five genomic dsRNAs, ranging from 1224 to 2407 bp, encoding five putative open reading frames (ORFs), of which ORF1 encodes a conserved RNA-dependent RNA polymerase (RdRp) consisting of 763 amino acids (aa) with a molecular mass of 84.03 kDa, ORF3 encodes a putative methyltransferase (Met) consisting of 627 amino acids (aa) with a molecular mass of 68.41 kDa, ORF4 encodes a P-A-S-rich protein behaving as coat protein (CP) consisting of 261 amino acids (aa) with a molecular mass of 27.60 kDa. ORF2 and ORF5 encode putative proteins with unknown functions which consisting of 697 amino acids (aa) with a molecular mass of 76.49 kDa and consisting of 323 amino acids (aa) with a molecular mass of 33.92 kDa, respectively. BLASTp analysis revealed that the RdRp protein of BdPmV2 had the highest similarity 56.52% to a virus previously identified as " RNA dependent RNA polymerase of Aspergillus fumigatus polymycovirus 1". Phylogenetic analysis based on the RdRp aa sequence indicated that BdPmV2 is a new member of the family Polymycoviridae.

Xinjiang, a major apple-producing region in, is renowned for its high-quality apples in China. In May 2024, symptomatic branches showing black rot canker were collected from commercial 'Golden Delicious' (Malus domestica) and Xinjiang Wild Apple (Malus. sieversii) trees in orchards located in Ili Kazakh Autonomous Prefecture (sites: 43.57°N, 81.92°E and 44.43°N, 80.78°E, respectively). The prevalence of symptomatic trees ranged from 30% to 40% (five-point sampling; n=50). External symptoms included charcoal-like cankers with bark peeling on primary and secondary branches, while internal examination revealed dark brown xylem with firm consistency. To isolate the causal pathogen, symptomatic branch samples (n = 14) were cut into 5 mm length pieces, surface-disinfected in 75% ethanol and rinsed with sterile water, and then transferred onto potato dextrose agar (PDA). Plates were incubated at 25°C for 3 days. Colonies consistent with Botryosphaeriaceae species (Phillips et al. 2013) (n = 20) were consistently obtained, and pure cultures were obtained by transferring single hyphal tips to fresh PDA. Colonies exhibited light gray coloration, irregular margins, and abundant aerial mycelium. To induce pycnidia formation, two isolates (ZM305-5 and ZM316-5) were cultured on pine needle agar (PNA) under near-ultraviolet light (340 nm) at 25°C for 20 days. Conidia (n = 50) were thick-walled, ovoid, initially hyaline, and turned dark brown at maturity, mostly aseptate, with the size of mature conidia ranging from 24.99 to 29.07 (27.14) × 13.76 to 15.78 (14.71) μm. Molecular identification was performed by sequencing the internal transcribed spacer (ITS) using primers ITS1/ITS4, results showing 100% identity with the Diplodia bulgarica holotype (CBS 124136). To further confirm the identity, the translation elongation factor 1-alpha (tef1) and the beta-tubulin (tub2) gene were sequenced using EF1-688F/EF1-1251R and Bt2a/Bt2b, respectively (Phillips et al. 2012). Sequences were deposited in GenBank (accession nos. PV362629-PV362631 for ITS, PV388909-PV388912 for tub2, and PV419584-PV419587 for tef1). A maximum likelihood (ML) multi-locus phylogenetic analysis clustered Chinese isolates with reference strains of D. bulgarica. Pathogenicity tests were conducted on 1-year-old shoots (n = 5) of 5-year-old saplings trees by wounding the bark of test shoots and inoculating mycelial plugs of these isolates. Control branches were mock-inoculated with sterile PDA plugs. After 30 days, dark-brown necrotic lesions developed in inoculated branches with length ranging from 31.5 to 45.5 mm, whereas the negative controls remained asymptomatic. The experiment was performed twice with similar results. Koch's postulates were fulfilled by re-isolating the pathogen from lesion margins and confirming its identify via morphology and ITS sequence. D. bulgarica was first described affecting M. sylvestris in Bulgaria (Phillips et al. 2012) and later associated with branch cankers on M. domestica in Iran (Abdollahzadeh 2015), India (Nabi et al. 2020), Germany (Hinrichs-Berger et al. 2021), Türkiye (Eken 2021), and California (Elfar et al. 2023). The pathogen has also been identified as the cause of postharvest fruit rot (Eken 2022). To our knowledge, this is the first report of D. bulgarica causing black rot canker on apple in China, which providing important information for developing detection and disease control strategies in apple industry.