Molecular Identification And Phylogenetic Relationships Of Four Periplaneta Species Based On CO? Gene Sequences And GBS Technology

Sampled 203 individuals from four species of Periplaneta australasiae,Periplaneta americana,Periplaneta brunnea,and Periplaneta fuliginosa for genetic investigation,which are commonly known as cockroaches in the Blattaria,Blattidae,Periplaneta.Molecular identification and phylogenetic analyses for the four species of Periplaneta were performed through mitochondrial cytochrome c oxidase subunit I(COI)sequences and genotyping by sequencing(GBS)technology,which may give new sights into the genetic relationships between the four species and fill in the gaps in traditional morphologcial identification,and lay the molecular basis for following basic application research.Our research results are listed as follows:(1)118 variable sites in COI gene 658bp sequences from 203 samples of four Periplaneta species were indentified,which defined 20 haplotypes.(2)The intraspecific genetic distances were 0-0.01,the intraspecific average genetic distance was 0.0025,the theinterspecific genetic distance was 0.08-0.13,and the interspecific average genetic distance was 0.09 for four Periplaneta species.(3)Four Periplaneta species separately clustered a monophyletic clade on the phylogenetic tree and haplotype network based on the COI gene sequences.On the phylogenetic tree,P.fuliginosa was at the base while P.americana was at the top,which indicates that the former differentiated earlier and the later differentiated later.(4)GBS data results showed that the genetic differentiation index of four Periplaneta species was 0.4242-0.7044,indicating that they have high genetic differentiation degree and can be clearly differentiated into different species.(5)Through searching for the GBS data of Simple Sequence Repeats(SSR)markers of four Periplaneta species,7 base repeat types were identified,with 3 being the dominant base repeat types.83 SSR marker loci which can be used for identification of any two species.were screened out through SSR clustering and polymorphhism assessment analysis.(6)Based on the Principal Component and phylogenetic analysis of GBS Data,the present study showed that P.australasiae,P.americana,P.brunnea and P.fuliginosa clustered as separately monophyletic cluster or clade,which indicated that GBS data can clearly distinguish these four Periplaneta species.In summary,COI gene sequence and GBS sequencing data can be effectively used for molecular identification and phylogenetic preliminary analysis of Periplaneta species.This study provides a scientific basis for subsequent studies on the species of the cockroach(commonly known as cockroach).