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Smokeless tobacco consumption induces dysbiosis of oral mycobiome: a pilot study


Smokeless tobacco (SLT) alters the oral microbiome of smokeless tobacco users. Dysbiosis of oral bacteriome has been determined; however, the mycobiome of SLT users has not been characterized. The oral mycobiome was assayed by amplification and sequencing of the fungal internal transcribed spacer (ITS1) region from oral swab samples of non-SLT users, SLT users (with or without oral lesions), and SLT with alcohol users. We observed that the richness and diversity of oral mycobiome were significantly decreased in SLT with oral lesions users than in non-users. The β-diversity analysis showed significant dissimilarity of oral mycobiome between non-users and SLT with oral lesions users. Linear discriminant analysis effect size and random forest analysis of oral mycobiome affirm that the genus Pichia was typical for SLT with oral lesions users. Prevalence of the fungal genus Pichia correlates positively with Starmerella, Mortierella, Fusarium, Calonectria, and Madurella, but is negatively correlated with Pyrenochaeta, Botryosporium, and Alternaria. Further, the determination of oral mycobiome functionality showed a high abundance of pathotroph-saprotroph-symbiotroph and animal pathogen-endophyte-epiphyte-undefined saprotroph at trophic and guild levels, respectively, indicating possibly major changes in normal growth repression of types of fungi. The oral mycobiome in SLT users was identified and comprehensively analyzed for the first time. SLT intake is associated with oral mycobiome dysbiosis and such alterations of the oral mycobiome may contribute to oral carcinogenesis in SLT users. This study will provide a basis for further large-scale investigations on the potential role of the mycobiome in SLT-induced oral cancer.

Key points

SLT induces dysbiosis of the oral microbiome that can contribute to oral cancer.

Oral mycobiome diversity is noticeably reduced in SLT users having oral lesions.

Occurrence of Pichia can be used as a biomarker for SLT users having oral lesions.

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Data availability

The metabarcoding data of the fungal ITS amplicon was submitted to NCBI Short Read Archive (SRA) under BioProject accession number PRJNA803602.

Code availability

Not applicable.


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We are grateful to the Director, ICMR-NICPR, for providing the facilities to carry out this work. The Biokart India Pvt. Ltd. Bengaluru, India, services of metagenomics sequencing and data processing are thankfully acknowledged. We thank Mr. Ravi Kumar Yadav and Miss. Lata Joshi for the necessary technical assistance in the study.


This work was carried out under a Task-Force project (ISRM/14(04)/TF/2018) of the Indian Council of Medical Research (ICMR), India, funding which was awarded to MB.

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Authors and Affiliations



MB conceived, designed, and guided the study. MS and RH collected the samples. Data collection, data analysis, literature review, and the first draft of the manuscript were performed and written by MS. PS and SS led the literature survey. PS, SS, RH, HS, and MB reviewed and gave comments and suggestions for the manuscript. All of the authors gave final approval for manuscript submission.

Corresponding author

Correspondence to Mausumi Bharadwaj.

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Ethics approval

The study was approved by the Institutional Ethics Committee, ICMR-National Institute of Cancer Prevention and Research, India (NICPR/123/DIR/Ethical/2019/06).

Consent to participate

The detailed clinical proforma was filled and consent was taken from participants before sample collection.

Conflict of interest

The authors declare no competing interests.

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Sajid, M., Sharma, P., Srivastava, S. et al. Smokeless tobacco consumption induces dysbiosis of oral mycobiome: a pilot study. Appl Microbiol Biotechnol 106, 5643–5657 (2022).

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  • Smokeless tobacco
  • Smokeless tobacco users
  • Oral mycobiome
  • Oral lesions
  • Alcohol
  • Fungal co-occurrence
  • Next-generation sequencing