Microbial Community Structure Of Cicada Flowers And Cultivation Of The Dominant Fungus

Cicada flower is a long-established edible fungus that contains a variety of polysaccharides,cordycepic acid,nucleosides,sterols,and peptides.However,as the natural environment changes,the resource of cicada flowers begins to be scarce.Artificial cultivation of cicada flowers has become a hot spot of concern.In this study,we explored the microbial community structure of cicada flowers to find the key strains,and then screened some strains to select the one with better fermentation traits.Finally,the simulated fermentation method by adding cicadae pupa powder was used to cultivate the worm complex.The main results are as follows:(1)The high-throughput sequencing method was used to explore the fungal and bacterial community structure of cicada flowers from different regions,different compartments and different growth stages.Isaria cicadae was dominant and widely distributed in the fungal community among different regions.Among them,OTU6782 was the most abundant OTU.For the community structure of different compartments(the soil adhering to the surface of cicada flowers,microhabitat including external mycelial cortices that cover larvae,coremia of cicada flowers,inside sclerotia of cicada flowers and the external mycelial cortices that cover larvae),Isaria cicadae dominated and OTU6782 was the main group.The OTU14723 and OTU5752 might have a symbiotic relationship with the dominant fungus.During the growth of cicada flowers,Isaria cicadae dominated quickly.OTU6782,which was rich in intraspecific diversity.Besides,OTU12109,OTU13152,OTU486,OTU2297,OTU11491,OTU246,OTU58,OTU2950,OTU8787,OTU73,OTU130 could grow together with dominant fungi.(2)The ITS and 18 S rRNA sequences of 8196,82542,81957 and TZC-3 strains were aligned.Liquid fermentation of different strains was performed.Through the molecular and morphological identification,8196,82542,81957 and TZC-3 strains had more than 99% similarities with Isaria cicadae,and 99.65% similarity with OTU6782.Thus,these strains and OTU6782 belong to the same species Isaria cicadae.The liquid fermentation culture was carried out,and biomass,extracellular polysaccharide content,intracellular polysaccharide content,extracellular nucleosides content and extracellular nucleosides contents of different strains were determined.It was found that different Isaria cicadae strains have some differences in the fermentation indexes.The TZC-3 strain was selected for the subsequent fermentation experiment for exerting obvious advantages in biomass,production of intracellular polysaccharide and adenosine content.(3)The effect of cicadae pupa powder on the fermentation process of Isaria cicadae TZC-3 was investigated,and the differences in composition between the cultured product and the wild fruiting body were compared.The contents of intracellular polysaccharides,adenosine and HEA were promoted to different degrees by adding cicadae pupa powder.We found that the worm complex with 4% added amount was better in yield of metabolites.In detail,the content of intracellular polysaccharide was 14.55 mg/g,74.78 mg/100 mL.The adenosine content was 1.22 mg/g,6.66 mg/100 mL,and HEA content was 1.58 mg/g,8.63 mg/100 mL.Compared with the cicadae pupa powder and mycelium,the composition in the enhanced mycelium(proximate ingredients,hydrolyzed amino acids,free amino acids,hydrolyzed monosaccharides and fatty acids)was more similar to wild fruiting bodies.Besides,the contents of some free amino acids(tonic acid,serine,threonine,phenylalanine,isoleucine,leucine and proline)and hydrolyzed monosaccharides(arabinose,galactose,glucose and xylose)had significantly increased in the enhanced mycelium.