Comparative Analysis of Mitochondrial Genome Features among Four Clonostachys Species and Insight into Their Systematic Positions in the Order Hypocreales

Abstract

The mycoparasite fungi of Clonostachys have contributed to the biological control of plant fungal disease and nematodes. The Clonostachys fungi strains were isolated from Ophiocordyceps highlandensis, Ophiocordycepsnigrolla and soil, which identified as Clonostachyscompactiuscula, Clonostachysrogersoniana, Clonostachyssolani and Clonostachys sp. To explore the evolutionary relationship between the mentioned species, the mitochondrial genomes of four Clonostachys species were sequenced and assembled. The four mitogenomes consisted of complete circular DNA molecules, with the total sizes ranging from 27,410 bp to 42,075 bp. The GC contents, GC skews and AT skews of the mitogenomes varied considerably. Mitogenomic synteny analysis indicated that these mitogenomes underwent gene rearrangements. Among the 15 protein-coding genes within the mitogenomes, the nad4L gene exhibited the least genetic distance, demonstrating a high degree of conservation. The selection pressure analysis of these 15 PCGs were all below 1, indicating that PCGs were subject to purifying selection. Based on protein-coding gene calculation of the significantly supported topologies, the four Clonostachys species were divided into a group in the phylogenetic tree. The results supplemented the database of mitogenomes in Hypocreales order, which might be a useful research tool to conduct a phylogenetic analysis of Clonostachys. Additionally, the suitable molecular marker was significant to study phylogenetic relationships in the Bionectriaceae family.

Keywords: Clonostachys; mitochondrial genome; phylogenetic analysis; protein coding genes; repeat sequence.

Figure 1

Culture and micromorphological characteristics with asexual morphs of four Clonostachys species in this study. (A–D): Culture of C. compactiuscula, C. rogersoniana, C. solani and Clonostachys sp.; (E–H): Perithecia of C. compactiuscula, C. rogersoniana, C. solani and Clonostachys sp.; (I–L): Conidia of C. compactiuscula, C. rogersoniana, C. solani and Clonostachys sp. Scale bars (A–C) = 1.5 cm; (D) = 1 cm; (E–H) = 10 μm; (I) = 2.5 μm; (J–L) = 10 μm.

Figure 2

Phylogenetic tree of Clonostachys genus by the analysis of three genes (nrLSU + Tub + ITS) dataset.

Figure 3

Circular maps of four newly sequenced mitogenomes from different Clonostachys species. (Genes are represented by different colored blocks, as presented in the legend below the maps. Colored blocks outside of each ring indicate that the genes were on the direct strand, while colored blocks in the rings indicate that the genes were located on the reverse strand.).

Figure 4

The proportion of the mitogenomes comprised by protein-coding, intronic, intergenic, and RNA genes (tRNA and rRNA) for four Clonostachys species. ((A–D): C. compactiuscula, C. rogersoniana, C. solani, Clonostachys sp.).

Figure 5

Codon usage analysis of the mitochondrial genomes from four Clonostachys species. ((A–D): C. compactiuscula, C. rogersoniana, C. solani, Clonostachys sp.).

Figure 6

Variation in length and base composition of each of the 15 core protein-coding genes (PCGs) among four Clonostachys mitochondrial genomes. ((A), PCG length variation; (B), GC content across PCGs; (C), GC skew; (D), AT skew).

Figure 7

Genetic analysis of the 15 core protein-coding genes among four Clonostachys mitogenomes. (K2P, the Kimura-2-parameter distance; Ka, the number of nonsynonymous substitutions per nonsynonymous site; Ks, the number of synonymous substitutions per synonymous site.)

Figure 8

Mitogenome synteny among four Clonostachys species. (Six homologous regions were identified among the four mitogenomes, while the sizes and relative positions of the homologous fragments varied across the mitogenomes).

Figure 9

Molecular phylogenies of 54 species based on Bayesian inference and Maximum likelihood analysis of 14 protein-coding genes (PCGs).