Molecular divergence and species delimitation of the cultivated oyster mushrooms: integration of IGS1 and ITS

Abstract

Identification of edible mushrooms particularly Pleurotus genus has been restricted due to various obstacles. The present study attempted to use the combination of two variable regions of IGS1 and ITS for classifying the economically cultivated Pleurotus species. Integration of the two regions proved a high ability that not only could clearly distinguish the species but also served sufficient intraspecies variation. Phylogenetic tree (IGS1+ITS) showed seven distinct clades, each clade belonging to a separate species group. Moreover, the species differentiation was tested by AMOVA and the results were reconfirmed by presenting appropriate amounts of divergence (91.82% among and 8.18% within the species). In spite of achieving a proper classification of species by combination of IGS1 and ITS sequences, the phylogenetic tree showed the misclassification of the species of P. nebrodensis and P. eryngii var. ferulae with other strains of P. eryngii. However, the constructed median joining (MJ) network could not only differentiate between these species but also offer a profound perception of the species' evolutionary process. Eventually, due to the sufficient variation among and within species, distinct sequences, simple amplification, and location between ideal conserved ribosomal genes, the integration of IGS1 and ITS sequences is recommended as a desirable DNA barcode.

Figure 1

Primer binding sites used to amplify IGS1 and ITS regions. The arrowheads show the 3′ end of each primer. LSU rDNA represents the large subunit ribosomal DNA and LSU rDNA represents the small subunit ribosomal DNA.

Figure 2

Phylogenetic tree constructed based on IGS1 sequences of experimental samples using UPGMA method. The sequences of AB234031 (P. pulmonarius); AB234030 (P. ostreatus); AB234045, AB234042, AB286142, and AB234047 (P. eryngii); AY463034 (P. nebrodensis); and AB286124 (P. eryngii var. ferulae) were downloaded from NCBI GenBank. Numbers close to branches indicate 1000 replication of bootstrap test and the codes refer to sample ID.

Figure 3

The median joining haplotype network of the combined IGS1 and ITS sequences. H and MV indicate haplotype and median vector, respectively.

Figure 4

Phylogenetic tree constructed by the combination of IGS1 and ITS sequences using UPGMA method. The IGS1 sequences of AB234031 (P. pulmonarius); AB234030 (P. ostreatus); AB234045, AB234042, and AB234047 (P. eryngii); AY463034 (P. nebrodensis); and AB286124 (P. eryngii var. ferulae) as well as ITS sequences of HM561973 (P. ostreatus); AB115050 (P. pulmonarius); AB286159 (P. eryngii var. ferulae); AB286173 and AB286174 (P. eryngii); EU424308 (P. nebrodensis); and AY315805 (P. cystidiosus) were downloaded from NCBI GenBank. Numbers close to branches indicate 1000 replications of bootstrap test and codes represent the sample ID.