The Impact Of Heterogeneity Environment On Population Differentiation And Sexual Reproduction Of Arthrobotrys Musiformis

The maintenance and evolution of sexual reproduction has been concerned by biologists.A few evidences from laboratory experiment were shown that heterogeneity environments may accelerate the evolution of sex,but they were questioned because of limited timescales and the simplified nature of laboratory environments.Dianchi Lake is a famous freshwater lake in Yunnan.The lake is not only polluted by severe nitrogen and phosphorus,but also has a heavy metal content higher than that of Yunnan Province.It is a natural heterogeneous environment.Arthrobotrys musiformis belongs to the Ascomycota Orbiliaceae.It is the asexual state of Orbilia,which is widely distributed in various habitats,and have both sexual reproduction and asexual reproduction.It is a good material for studying the effects of habitats on fungal sexual reproduction.In this study,three A.musiformis populations isolated from Dianchi sediment(DC),Xishan farmland soil(XS)next to Dianchi Lake,and ascospores of Orbilia(S)were selected as research materials from mitochondrial genomes diversity.The perspective reveals the impact of heterogeneous environments on sexual reproduction.The main findings are as follows:1.Phylogenetic analysis of A.musiformisPhylogenetic analysis of 48 A.musiformis strains was carried out by concatenated three sequences of ITS,RPB2 and TEF.All the strains were clustered into one branch,and differ from the A.oligospora,a fungus of same genus.Phylogenetic analysis showed that the selected strains are identified correctly.Based on the result from the analysis of nuclear genetic,the strains from the sediments of Dianchi and ascospores are clustered one clade,while strains from farmland are separate from theem.2.Mating type distribution of A.musiformisAfter re-sequencing to obtain high-quality data(clean data),using the software CLC Genomics Workbench assembly,and then using the MAT gene of A.oligospora(MAT]-1/MAT1-2)as a reference,local blast,obtain the corresponding fragment,design Primers amplify the MAT gene of 48 strains of A.musiformis,and obtained 32 sequences of MAT1-1.There were 22 sequences with deletion mutations at 521-526 bp,of which 68.2%occurred in strains from Dianchi Lake,which is presumed to be related to the heterogeneous environment in Dianchi Lake.In this study,only MAT1-1 was obtained,so it is speculated that the mating type of A.musiformis is heterozygous.3.Overall comparison of mitochondrial genomesOne strain was randomly selected from the three populations for mitochondrial genomes sequencing,which was YMF1.01900 S isolated from ascospores,YMF1.03721 DC in Dianchi sediment,and YMF1.03753 XS isolated from farmland soil.The mitochondrial genomes of the three strains showed significant differences in size,intron and intergenic region.The size is 174085 bp,179060 bp,184443 bp,and 184443 bp,respectively,containing 41,42 and 43 introns,respectively,and the proportion of introns increases sequentially:37.52%,45.62%49.85%,while the proportion of gene spacers decreased in turn:48.95%,39.94%,38.26%.There are 17,18 and 17 tRNAs,respectively.The number and sequence of protein-coding genes are relatively conservative.Except NADH2,NADH3,NADH4L,ATP8,rns,ml,other protein-coding genes contain introns of varying numbers;GC content is also basically the same;there is no significant difference in codon preference among amino acids in the use of relative synonymous codons.4.Polymorphism analysis of protein coding genes and intronsBy comparing the polymorphisms of 14 protein coding genes,17 mutation sites were found in NADH1,although all of them were synonymous mutations.By comparing the NADH1 gene of 29 strains(nearly 10 strains per population),these mutations also existed in the population.The strains from ascospores and Dianchi Lake sediment had the same base,while the strains isolated from Xishan had mutations at the corresponding sites.Therefore,this gene can be used as one of the molecular markers for fungal population genetics research.In the NADH6 gene,YMF 1.01900 S and YMF 1.03721 DC contain one intron,while YMF 1.03753 XS does not contain the intron.By designing primers to detect the presence or absence of introns in 17 strains,this situation still exists at the population level.Therefore,the presence of introns in this gene can also be used as one of the markers of fungal genetic diversity.By comparing the mitochondrial genomes,the results showed that the genetic differentiation promoted by heterogeneous environment,regardless of the diversity of coding genes or the presence of introns,the ascospores source population and the sediment-derived population of Dianchi Lake had the same changes.However,the population from Xishan,which is located very close to each other,is obviously different from that from the conidia,suggesting that the heterogeneous environment in nature is also a driving force for fungi to evolve towards sexual reproduction..