Isolation And Identification Of Dominant Fungi From Different Origins Of Fu Brick Tea And Its Effects On Volatile Flavor Components Of Fu Brick Tea

Fu brick tea is a post-fermented black tea,which is loved by consumers for its unique flavor quality and bioactive function.Microbial participation in the post-fermentation process is an important factor in the formation of Fu brick tea flavor quality,in which the "Jinhua Fungus" as the dominant fungus in the post-fermentation process of Fu brick tea,plays a key role in the formation of unique flavor quality of Fu brick tea.Because of the differences in raw materials,processes and environments,there are some differences in the dominant fungi and flavor qualities in Fu brick tea from different origins.In order to elucidate the effect of dominant fungi on the formation of flavor quality of Fu brick tea in different production areas,this study was conducted by collecting samples of Fu brick tea from five main production areas of Fu brick tea,namely Hunan,Guangxi,Shaanxi,Zhejiang and Guizhou,Firstly,based on high-throughput sequencing technology,we analyzed the composition and structure of fungi in Fu brick tea of different origins to determine the composition and species of the dominant fungi,and then isolated the dominant fungi from Fu brick tea samples of different origins and identified them by combining morphological and ITS sequencing methods,Finally,the dominant fungi from different origins were artificially inoculated into Fu brick tea for enhanced fermentation,and GC-MS combined with multivariate data statistical analysis was used to elucidate the effect of dominant fungi on volatile flavor components of Fu brick tea from different origins,which provides a reference basis for enriching the quality formation mechanism of Fu brick tea from different regions in China and improving the processing theory of Fu brick tea.The main results of this study are as follows:(1)There is some variability in the fungal composition of Fu brick tea from different origins.In this study,a total of 69 operational taxonomic units(OUTs)were obtained by sequencing all samples based on the variable region of the internal transcribed inter-region(ITS2)of the fungus,which were classified into 5 phyla,15 orders,20 families,23 families,27 genera and 33 species.The results of Alpha diversity analysis showed that there were differences in the diversity and richness of Fu brick tea fungi among five different sources,with the highest Fu brick tea samples from Hunan region,the second highest from Guangxi region and the lowest from Zhejiang region.Among them,the genus Eurotium was the dominant species in all samples,with an average relative abundance of 88.84%.The relative abundance of Aspergillus in Fu brick tea samples from Guangxi and Hunan regions was only between 1%,except for 34.81% and 18.27% in other samples,respectively.(2)Isolation,identification and differential analysis of dominant fungi in Fu brick tea from different origins.In this study,one dominant fungal strain,numbered HNJ,GXJ,SXJ,ZJJ and GZJ,was isolated from Fu brick tea samples from Hunan,Guangxi,Shaanxi,Zhejiang and Guizhou,respectively,and then incubated with M40 Y,Bengal red and 40%sucrose peptone medium for 5 days in a single spot to observe the morphological characteristics of the colonies,respectively.The results showed that there were significant differences in the colony morphology of the five dominant fungal strains,among which the differences between strain HNJ and the other four dominant microorganisms were obvious,ZJJ and SXJ had similar colony morphology on M40 Y medium,and ZJJ and GZJ had similar colony morphology on Bengal red medium.It was observed by light microscopy and scanning electron microscopy that the ascospores of the five dominant fungi were all subspherical in shape,yellow in color,with wrinkles and roughness on the surface,and there were obvious differences in ascospore size,conidial head structure and mycelial entanglement of different dominant fungi.ITS sequencing of the five dominant fungi showed that the ITS gene fragments of the different dominant fungi differed,but all had high homology with Aspergillus cristatus(ACCESSION: MW512851.1).Therefore,GXJ,ZJJ,SXJ,GZJ and HNJ were tentatively identified as Aspergillus cristatus based on morphological observations and ITS sequence comparison results and according to the latest nomenclature rules.(3)The enhanced fermentation of different "Jinhua Fungus" has a certain effect on the volatile aroma components of Fu brick tea.Five strains of "Jinhua Fungus" were artificially inoculated into raw black tea for enhanced fermentation,and Fu brick tea from Hunan(HNE),Guizhou(GZE),Zhejiang(ZJE),Guangxi(GXE)and Shaanxi(SXE)were obtained from different sources of "Jinhua Fungus" for enhanced fermentation for 22 days,and compared with unfermented raw black tea(NNF)and naturally fermented Fu brick tea(NF).The results showed that there were significant differences in the effects of different "Jinhua Fungus" on the flavor of Fu brick tea.A total of 63 volatile components were measured in all Fu brick tea samples,with the main volatile components being alcohols,ketones,aldehydes,esters and hydrocarbons.In terms of volatile component fractions,compared to NF,alcohols were 25.9% lower,ketones were 14.8% higher,aldehydes were 31.7% higher,esters were65.2% lower and hydrocarbons were 23.4% higher in HNE.The alcohols in GZE were elevated by 10.6%,ketones by 22.4%,aldehydes by 40.9%,esters by 55.0% and hydrocarbons by 59.5%.The alcohols in ZJE were elevated by 11.3%,aldehydes by 33.4%,esters by 59.1% and hydrocarbons by 41.1%.Alcohols in GXE decreased by 27.7%,ketones decreased by 22.0%,aldehydes decreased by 34.0%,esters decreased by 64.6% and hydrocarbons increased by 123.1%.The alcohols in SXE were elevated by 12.2%,aldehydes by 30.9%,esters by 55.1% and hydrocarbons by 35.8%.Comparing the OAV values of volatile components in Fu brick tea samples from 22 days of enhanced fermentation with NF and different sources of "Jinhua Fungus",the results showed that D-Limonene and beta-Ionone with floral properties and(E)-3-hexen-1-ol with green properties in HNE contributed significantly to the flavor of Fu brick tea.Linalool with floral properties in GZE and SXE contributed significantly to the flavor of Fu brick tea.The D-Limonene of floral property and(E)-3-hexen-1-ol of green property in ZJE contributed significantly to the flavor of Fu brick tea.Styrene with floral attributes in GXE contributed significantly to the flavor of Fu brick tea.In addition,comparing the aroma composition of Fu brick tea for 22 days of intensive fermentation with five strains of "Jinhua Fungus",it was found that the "Jinhua Fungus" isolated from Shaanxi strains contributed significantly to the methyl salicylate of holly oil aroma property.