Application of 16S rRNA Gene Sequencing in Assessing Pharmaceutical Impact on Soil Microbiomes: A Review

Soil microbial communities are vital for ecosystem health, yet they face increasing pressure from pharmaceutical contamination entering soils through wastewater, agricultural practices, and landfill leachate. This review synthesizes the application of 16S rRNA gene sequencing in assessing the impact of these emerging contaminants on soil microbiomes. We detail the entire sequencing pipeline, from experimental design and laboratory workflows to bioinformatics analysis, while critically evaluating the technical biases and limitations inherent to the method. Our analysis confirms that pharmaceuticals, including antibiotics, NSAIDs, antidepressants, and complex mixtures, induce significant shifts in microbial community structure, reducing alpha diversity and enriching for resistant taxa and antimicrobial resistance (AMR) genes. However, while 16S sequencing is a powerful tool for taxonomic profiling, its limitations in functional prediction and resolution beyond genus level necessitate a more holistic approach. We conclude that the future of the field lies in integrating multi-omics technologies (metagenomics, metatranscriptomics) with standardized methodological frameworks to move beyond census-based data and truly understand the functional resilience of soil ecosystems. This integrated approach is essential for accurate environmental risk assessment and developing effective strategies to mitigate the ecological impact of pharmaceutical pollution.