The Species Diversity And Molecular Idendification Of The Enigmatic Species Of The Genus Conidiobolus In Anhui Guniujiang Nature Reserve

The genus Conidiobolus occupies a special important position not only in thesystematics of Entomophthorales and Fungi, but also plays an important role in theecological environment. Guniujiang Nature Reserve is one of the typical areas in Chineseeastern subtropical evergreen broad-leaved forest where protection species of animal, plantand fungi are extremely rich. Therefore, species diversity of Conidiobolus (belonging toFungi) from Gunniujiang Nature Reserve of Anhui were systematically studied.From April2010to March2012, soil and plant detritus samples were collected fromGunniujiang Nature Reserve in Shitai county, Chizhou city and Qimen county,Huangshang city of Anhui province. Strains isolated from these samples were identifiedmorphologically. Following the isolation and identification method of Drechsler and King,eighty-two Conidiobolus isolates were isolated from this area. According to themorphological taxonomic system of King and Ben-Ze’ev&Kenneth, eighty-twoConidiobolus strains were identified as ten Conidiobolus species, including one newspecies, one new variety species and one new record from China; They belong to threesubgenera, the subgenus Conidiobolus contains: Conidiobolus guniujiangensis sp. nov.(new species)(1strain), Conidiobolus osmodes (5strains), Conidiobolus iuxtagenitus (2strains), Conidiobolus macropapillatus (3strains); the subgenus Capillidium contains:Conidiobolus coronatus (12strains), Conidiobolus rhysosporus (new record)(1strain); thesubgenus Delacroixia contains: Conidiobolus coronatus (55strains), Conidiobolusbrefeldianus (1strain), Conidiobolus polytocus (1strain), Conidiobolus coronatus var.longvillosus var. nov.(new variety)(1strain). Through the investigation and research,Conidiobolus coronatus was the richest species and Conidiobolus heterosporus was thesecond common distribution species in Anhui Gunniujiang Nature Reserve.The genomic DNA from ten Conidiobolus species were extracted from mycelia byCTAB method. With the primer LROR and LR5,28S rDNA partial region was amplifiedby the polymerase chain reaction (PCR).1kb PCR products were purified and sequenced.The sequences were edited and aligned by mean of BIOEDIT and DNAMAN software.With27sequences of Conidiobolus, phylogenetic tree was constructed by PAUP*4.0b10software using Maximum Parsimony (MP). Batkoa apiculata and Entomophthora muscaewere selected as outgroup. Phylegenetic tree of Conidiobolus included3clades which wereconsistent with Ben-Ze’ev&Kenneth’s taxonomic system in the three sub-division of thegenus in1982. The molecular evidence supported the morphological results of10 Conidiobolus species.We found the Conidiobolus strain RCEF5347similar with Conidiobolus coronatusmorphologically, but it distinguished from Conidiobolus coronatus in the length of villoseaccording to the original description.28S rDNA and EF-1α partial region were amplifiedwith a enigmatic isolates RCEF5347and5strains of Conidiobolus coronatus which wereisolated from different areas in China. The alignments with sequences of C. coronatusfrom sequencing and NCBI were fed to DNAMAN software for calculating geneticsimilarities. The similarities of partial28S rDNA and EF-1α sequences between C.coronatus are98.4%—99.8%and98.1%—100%; the similarities of partial28S rDNA andEF-1α sequences between the strain RCEF5347and C. coronatus are96.4%—96.9%and94.6%—95.2%. The strain RCEF5347was identified as a new variety of Conidioboluscoronatus based on the morphological evidence.