Abstract

Smokeless tobacco products (STPs) carry assorted microbial population that contributes to carcinogens synthesis like tobacco-specific nitrosamines (TSNAs). Extensive exploration of microbiota-harboring STPs is required to understand their full carcinogenic potential. Here, we applied 16S rRNA gene sequencing to investigate bacteriome present in moist STPs immensely consumed in India (Khaini, Moist-snuff, Qiwam, and Snus). Further, the functional metagenome was speculated by PICRUSt (Phylogenetic Investigation of Communities by Reconstruction of Unobserved States) to assign the abundance of genes related to nitrogen metabolism, bacterial toxins, antibiotic drug resistance and other pro-inflammatory molecules. Highly diverse bacterial communities were observed in all moist STPs. Taxonomic analysis revealed a total of 549 genera belonging to four major phyla Proteobacteria, Firmicutes, Bacteroidetes and Actinobacteria. Overall, the core bacterial genera Acinetobacter, Bacillus, Prevotella, Acetobacter, Lactobacillus, Paracoccus, Flavobacterium, and Bacteroides were significantly abundant in moist STPs. Elevated moisture-holding products like Moist-snuff and Qiwam harbor rich bacterial species diversity and showed similar bacteriome composition. Furthermore, Qiwam products showed the highest level of genes associated with nitrogen metabolism, antibiotic resistance, toxins, and pro-inflammation (predicted by PICRUSt) which can contribute to the synthesis of TSNAs and induction of oral cancer. The present broad investigation of moist STPs-associated bacteriome prevalence and their detailed metabolic potential will provide novel insight into the oral carcinogenesis induced by STPs.

Highlights

  • Non-combustible form of tobacco is used by >300 million people worldwide and majorly (>85%) in South Asian countries (Siddiqi et al, 2020)

  • The bacteriome of moist STPs was established by high throughput sequencing of 16S rDNA (V3-V4 region) sequence present in genomic DNA isolated from the STPs and first time expansively analyzed by MicrobiomeAnalyst tool (Dhariwal et al, 2017)

  • We focused on the moist STPs because high moisture content plays a significant role in the increased level of tobacco-specific nitrosamines (TSNAs) by facilitating the growth of the microorganisms (Gholizadeh et al, 2016; Han et al, 2016)

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Summary

Introduction

Non-combustible form of tobacco (smokeless tobacco) is used by >300 million people worldwide and majorly (>85%) in South Asian countries (Siddiqi et al, 2020). A report on global tobacco consumption documented that there are 199.4 million smokeless tobacco (SLT) users in India (GATS-2, 2017). Characterization of Moist STP-Linked Bacteriome addiction, diabetes mellitus, oral diseases, neuronal and reproductive defects (Carlsson et al, 2017; Centers for Disease Control and Prevention, 2020). These pathological effects are generally assigned to carcinogens present in STPs essentially tobacco-specific nitrosamine (TSNAs), benzo(a)pyrene and heavy metals (Critchley and Unal, 2003; Stepanov et al, 2008). The STPs consumed in the South Asian region have higher TSNAs content in contrast to STPs available in Western countries (Nasrin et al, 2020)

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