Background:The objective of this study was to explore the impact of Erbium:Yttrium-Aluminum-Garnet (Er:YAG) laser therapy on the concentration of cariogenic species and the microbial load composition within therapeutic cavities. This investigation aimed to assess the potential reduction in microorganisms following Er:YAG laser treatment and make a comparative analysis with conventional manual and rotating therapies (CT).Methods:Conducting a clinical trial, adult participants presenting with active deep carious lesions on permanent teeth were divided into two groups: a control group undergoing conventional treatment (CT) and an intervention group receiving Er:YAG therapy. Before and after any conservative treatment, oral samples were collected using a sterile microbrush scrubbed within the base of the dentinal cavity tissue. The focus was on evaluating the percentage reduction and colony-forming units (CFUs) count for total microorganisms, including Candida spp., Streptococcus spp., and Lactobacillus spp., after Er:YAG and conventional treatments.Results: indicated a substantial microbial reduction ranging from 80.2% to 100%, notably observed for total microorganisms and Streptococcus spp. (p < 0.05) following Er:YAG laser therapy. This outcome underscored the potential clinical applications of Er:YAG laser treatment, particularly in cases involving pediatric and complicated patients. The minimally invasive properties of Er:YAG laser therapy, coupled with its demonstrated efficacy in reducing microbial load, suggest its viability as an alternative or adjunctive approach in conservative dental treatments.Conclusion: These findings contribute valuable insights into the potential benefits of incorporating Er:YAG laser therapy into clinical practice, offering a promising avenue for improving treatment outcomes, especially in challenging cases.