BACKGROUND: This systematic review summarized current evidence regarding the role of upper aerodigestive tract microbiomes (UAM) in laryngeal squamous cell carcinoma (LSCC) development, progression, clinical, and oncological outcomes. METHODS: Two investigators systematically search PubMed, Scopus, and Cochrane Library databases for studies investigating microbiome characteristics, mechanistic roles, and associations with clinical and oncological outcomes in LSCC according to the Preferred Reporting Items For A Systematic Review And Meta-analysis statements. The bias analysis was conducted with the methodological index for nonrandomized studies. RESULTS: Ten studies were included, accounting for 491 LSCC patients. LSCC tissues demonstrated lower bacterial diversity compared with controls. Taxonomic analyses suggested an overrepresentation of Bacteroidetes (Prevotella) and Fusobacteriota (Fusobacterium) in LSCC, while Firmicutes (Stomatobaculum longum, Abiotrophia, Gemella, and Streptococcus) and Actinobacteria (Actinomyces, Corynebacterium, and Rothia mucilaginosa) were predominant in control tissues. Firmicutes demonstrated the largest compositional variation across studies, with 30.9%-63.6% abundance in LSCC compared with 13.9%-32% in controls. Two studies explored microbiome signatures: one for LSCC diagnosis and another for prognosis. Substantial methodological heterogeneity was observed across studies regarding confounding factor analysis, UAM assessment protocols, and control tissue selection. CONCLUSION: The current literature supports potential distinct UAM signatures between LSCC and noncancerous tissues, with Bacteroidetes and Fusobacteriota enriched in LSCC tissues. Although emerging evidence supporting the key role of UAM in the development of LSCC, substantial methodological heterogeneity across studies necessitates standardized protocols for future investigations.