Analysis of the Fungal Diversity and Community Structure in Sichuan Dark Tea During Pile-Fermentation

Abstract

The fungi present during pile-fermentation of Sichuan dark tea play a pivotal role in the development of its aroma and physical characteristics. Samples of tea leaves were collected on days 0 (YC-raw material), 8 (W1-first turn), 16 (W2-second turn), 24 (W3-third turn), and 32 (W4-out of pile) during pile-fermentation. High-throughput sequencing revealed seven phyla, 22 classes, 41 orders, 85 families, 128 genera, and 184 species of fungi. During fermentation, the fungal diversity index declined from the W1 to W3 stages and then increased exponentially at the W4 stage. A bar plot and heatmap revealed that Aspergillus, Thermomyces, Candida, Debaryomyces, Rasamsonia, Rhizomucor, and Thermoascus were abundant during piling, of which Aspergillus was the most abundant. Cluster analysis revealed that the W4 stage of fermentation is critical for fungal growth, diversity, and the community structure in Sichuan dark tea. This study revealed the role of fungi during pile-fermentation in the development of the essence and physical characteristics of Sichuan dark tea. This study comes under one of the Sustainable Development Goals of United Nations Organization (UNO) to “Establish Good Health and Well-Being.”

Comprehensive Analysis of Bacterial Community Structure and Diversity in Sichuan Dark Tea (Camellia sinensis)

“…In recent years, only fungi have been the focus of researchers exploring the composition of the microbial community in dark tea, such as Eurotium cristatum (Yan et al, 2021), but knowledge about the bacterial community is scarce (Zeng et al, 2020). Bacterial flora displayed a lagging growth phase at the early flowering stage of Camellia sinensis due to the exponential growth of E. cristatum, which transformed into bacterial exponential growth at the late flowering stage to establish a symbiotic relationship with E. cristatum (Liu et al, 2014).…”

“…Finally, DNA was purified using the E.Z.N.A. TM Soil DNA Kit to remove impurities, such as polyphenol, that may have a negative effect on the PCR reaction (Yan et al, 2021).…”

“…Pair reads were merged into a single sequence by following the overlap relationship between PE reads and filtered for quality control to a avoid splicing effect. Finally, reads obtained by sequencing were checked for their quality, and sequence direction was corrected (Mukherjee et al, 2014;Yan et al, 2021). According to primer use, proper barcoding/labeling was performed on each step to avoid any error.…”

Determination of Community Structure and Diversity of Seed-Vectored Endophytic Fungi in Alpinia zerumbet

“…To amplify the ITS1 sequence of the fungal ITS region using PCR, the following primer pairs were employed: ITS1F (5′-CTTGGTCATTTAGAGGAAGTAA-3′) and ITS2R (5′-GCTGCGTTCTTCATCGATGC-3′) ( Yan et al, 2021a ). Each reaction comprised a 1 × polymerase chain reaction (PCR) buffer, 1 mM of dNTPs, 0.2 mM of each forward and reverse primer, 1.25 U of FastPfu DNA Polymerase (TransStart ® , AP221-01, China), and 2 μl of DNA template, with a final volume of 25 μl achieved by adding ddH 2 O.…”

“…Endophytic symbiotic fungi establish an irreversible long-lasting association, namely, seed-vectored endophytic fungi, which travels from generation to generation ( Johnston-Monje and Raizada, 2011 ). In Eucalyptus, rice, tea, and maize, seed-derived endophytic microbes serve as a source of future microbiota of seedlings and develop a very pleasant aroma during pil fermentation of tea ( Johnston-Monje and Raizada, 2011 ; Yan et al, 2021a , b ). Therefore, research to disclose the complex diversity of the endophytes of plant seeds, especially for medicinal and spice plants species, is critical.…”