The Cultural Characteristic And Ontogeny Of Three Species Of Phallus

In order to better carry on the culture characteristic and the ontogeny research to the Phallus fungi,we collected two wild strains of Phallus ultraduplicatus and P.dongsun from Shandong province,and cultivated strain of P.rubrovolvatus from Guizhou province.Firstly,morphological identification of mycelium,spore and fruit body and molecular identification based on ITS sequence were carried out for the three species.Then,the cultural character and domestication of those three strains were carried out from the preliminary screening of stock culture medium,single factor experiment,orthogonal experiment,response surface method optimization,cultivation material formula experiment,cultivation material C/N ratio experiment and cultivation.Finally,the ontogeny of button at different growth stages was studied by paraffin sectioning during the cultivation process.The results showed that:1.The morphology of mycelia of the three species were all dendritic,with obvious clamp connection,and the differences in mycelial morphology was slightly.The spores of the three species were all elliptic cylinders,the spores of P.ultraduplicatus were elongated,the spores of P.dongsun and P.rubrovolvatus were coarser and shorter.The fruit bodies of the three species were mainly different from each other in the indusium.The indusium of P.ultraduplicatus was thinner,about 2-4 cm long,with irregular mesh.The indusium of P.rubrovolvatus was thicker,about 6-8 cm long and has circular mesh,while P.dongsun had no indusium.2.The species relationship of the three species was clear.The P.ultraduplicatus clustered in the same group with sequences from Liaoning Province,and the P.rubrovolvatus clustered in the same group with sequences from Guizhou and Hunan provinces.The P.dongsun clustered in the same group as those from Guizhou and Hunan of China,Korea and Japan.The P.ultraduplicatus was relatively closely related to P.dongsun,and was far related to P.rubrovolvatus.3.The optimal culture conditions of P.ultraduplicatus were glucose 15.3 g·L^-1,peptone 2.3 g·L^-1,p H 6.5 and temperature 26℃.The addition of 50-150 mg·L^-1 Vitamin B₁ and 2.0-2.5 g·L^-1 KH₂PO₄could promote the growth of mycelia,while the addition of more than 0.5 g·L^-1 Mg SO₄ inhibited the growth of mycelia.The optimal cultivation formula was 65%sawdust,33%wheat bran,1%gypsum powder,1%sucrose.Buttons began to appear after casing soil cultivation for 25 days,and fruit bodies began to mature after 55 days,with biological efficiency of 34%.4.The optimal culture conditions of P.dongsun were glucose 23 g·L^-1,yeast extract power 5.3 g·L^-1,p H 5.5 and temperature 26℃.The addition of 1.0-1.5 g·L^-1 KH₂PO₄ in the medium could promote the growth of mycelia,while the addition of Vitamin B₁ and Mg SO₄ could not.The optimal cultivation formula was 68%sawdust,30%wheat bran,1%gypsum powder,1%sucrose.Buttons began to appear after casing soil cultivation for40 days,and fruit bodies began to mature after 80 days,and the biological efficiency was17%.5.The optimal culture conditions of P.rubrovolvatus were sucrose,yeast extract,p H5,and temperature 28℃.The optimal cultivation material formula was 58%sawdust,40%wheat bran,1%gypsum powder,1%sucrose.Buttons began to appear after casing soil cultivation for 18 days,and fruit bodies began to mature after 48 days.The biological efficiency was 40%.6.The button’s inner development of the three species was the same,and the development time was different.The first development was the colloid cavity,then the hymenium,the stipe cavity,the stipe,the pileus,the indusium（veil）followed.During the development of button,the morphology of mycelium changes from loose to compact and from interwoven to organized.Under the same diameter of button,the development of P.dongsun was the fastest,when the diameter is 2 cm,all organs had basically developed,the development of P.rubrovolvatus had completed when the diameter was 3 cm,P.ultraduplicatus was the slowest,all organs had developed when the diameter was 4 cm.The research results of cultural character and domestication of those three species can provide theoretical guidance for the cultivation of Phallus fungi in northern China.The research results of morphology and fruit body ontogeny of the three species will be helpful to the identification and genetic relationship exploration of Phallus fungi.