| Pochonia and Metapochonia,as two simple genera in Clavicipitaceae,have poor species diversity and narrow host range.Species from Pochonia and Metapochonia can infect nematodes for different periods and are widely used as nematode biocontrol agents.Species in Pochonia and Metapochonia have simple morphology and tiny distinction,which lead to confusion in understanding of the two genera.In this study,comprehensive morphology and phylogeny were conducted on the nine strains.With Pochonia and Metapochonia as the core branches,we reconstructed the phylogenetic framework of the family Clavicipitaceae using combined ITS and 5-gene（nr SSU,nr LSU,tef,rpb1 and rpb2）dataset.Combining morphological characteristics,species from Pochonia and Metapochonia were defined.Besides,varieties and some species classified into wrong systematic positions were revised.Based on the second generation genome sequencing data,nine mitochondrial genomes（mitogenomes）were assembled,annotated and analyzed.We also constructed the phylogenetic tree of Hypocreales based on the protein coding genes（PCGs）,the result proved that the PCGs could be a method to identify species.The main results were as follows:（1）Based on the evidence of morphology and molecular systematics,three variaties of P.chlamydosporia were revised and proposed to recognize at the rank of species,respectively.The five trains YFCC 806,YFCC 8606,YFCC 8607,YFCC 8608 and YFCC 8609 were defined as M.bulbillosa Kepler,S.A.Rehner ＆ Humber.The strain YFCC 810 was defined as M.rubescens Kepler,S.A.Rehner ＆ Humber.The strain YFCC 808 was defined as M.sinensis.The strain YFCC 802 was defined as P.chlamydosporia Zare ＆ W.Gams.The strain YFCC 809 was defined as P.spinulospora H.Yu,Y.Wang ＆ H.J.Wu stat.nov..（2）The topological structure of phylogenetic tree inferred from mitochondrial PCGs agreed with the nuclear multigene phylogenetic tree constructed by Bayesian tree and Maximum likelihood methodes.The seven strains YFCC 806,YFCC 8606,YFCC8607,YFCC 8608,YFCC 8609,YFCC 8608,YFCC 808 and YFCC 810 were clustered together in Clavicipitaceae.The two strains YFCC 802 and YFCC 809 were clustered together in Pochonia.The strain YFCC 802 was clustered with P.chlamydosporia（NCBI number: KF 479445）.（3）Comparative analysis of the mitogenomic structures of nine strains showed that they were similar in the same species.The five strains YFCC 806,YFCC 8606,YFCC8607,YFCC 8608 and YFCC 8609 were M.bulbillosa.Mitogenomes sizes of the four strains YFCC 806,YFCC 8606,YFCC 8607 and YFCC 8608 were 23590 bp,mitogenome size of strain YFCC 8609 was 23588 bp.Besides,the five strains had the same genetic compositions（a total of 42 genes with 25 t RNAs,15 PCGs,one rnl and one rns）,similar arrangement structure and same GC content（28.80%）.The length,gene size,and gene content of mitogenomes varied among different species.Mitogenome size of M.bulbillosa was the smallest（23588 bp）,while mitogenome size of M.sinensis was the largest（30796 bp）.Besides the basic 42 genes,M.sinensis,P.chlamydosporia and P.spinulosporia also contained ORFs.Eight ORFs were in M.sinensis,three ORFs were in P.chlamydosporia,and two ORFs were in P.spinulosporia.The length of nad2 gene of M.rubescens was 1119 bp,other species were 1668 bp.The length of cox1 gene of P.chlamydosporia was 1587 bp,the length of cox1 gene of P.spinulosporia was 1137 bp,other species were 1635 bp.The length of atp6 gene of P.chlamydosporia was 465 bp,the length of atp6 gene of P.spinulosporia was 786 bp,but other species were 801 bp. |
