Calibration of human and dog molecular markers for direct quantification of faecal pollution in marine ecosystems

Aquatic environments face increasing contamination pressures, threatening ecosystems and human health. Microbial Source Tracking (MST) enables identification of pollution sources such as sewage and animal faeces, yet translating MST data into quantitative contamination levels remains challenging. We calibrated gene copy numbers from common MST assays-HF183 and Lachno3 (sewage) and DG3 (dog faeces)-against measured faecal concentrations. Dilution series of sewage and dog faeces were analysed using quantitative PCR (qPCR) and digital PCR (dPCR), spanning raw sewage to 1:10¹⁰ seawater dilution and dog faeces from 2 g/L to 2 × 10^-9 g/L. Strong linear relationships were observed (R² > 0.98 for sewage; R² > 0.94 for dog faeces). These data underpin an online Faecal Pollution Calculator that converts qPCR and dPCR results into contamination levels, providing researchers and water managers with a practical tool to assess source contributions to poor water quality in aquatic ecosystems.