Bacillus subtilis DES-59 Modulates Microbial Community Structure and Aroma Formation in Cigar Fermentation to Improve the Quality of Heat-Not-Burn (HnB) Tobacco.

Abstract

Cigar tobacco is characterized by high protein and low sugar contents, which limits the sensory performance of cigar-flavored heat-not-burn (HnB) products. In this study, Yunxue 6 cigar tobacco powder was fermented with Bacillus subtilis DES-59 for 7 days under solid-state culture at 25°C and 65% relative humidity. Compared with the control group, fermented tobacco exhibited a 10.55% reduction in protein, alongside 30.10% and 47.86% increases in total free amino acids and reducing sugars, respectively. Py-GC/MS analysis at 350°C showed increased release of pyrolytic aroma compounds, with the relative abundances of esters, aldehydes / ketones, and phenols increasing by 86.95%, 58.53%, and 35.69%, respectively. Sensory evaluation of HnB cigarettes manufactured from the fermented tobacco powder showed an overall sensory score improvement from 79.00 to 83.00, mainly driven by enhanced aroma and flavor. High-throughput sequencing demonstrated that inoculation reshaped the bacterial community, increasing the relative abundance of Bacillus from 12.20% to 52.40%. PICRUSt functional prediction suggested an increased functional potential for amino acid and carbohydrate metabolism in the treated group. Overall, inoculation with B. subtilis DES-59 was associated with changes in the chemical composition, bacterial community structure, pyrolysis-derived aroma profile, and sensory characteristics of cigar tobacco powder. These findings provide preliminary evidence supporting the potential application of B. subtilis DES-59 in the fermentation of cigar tobacco powder for the development of cigar-flavored raw materials for HnB products.