Hepatocellular carcinoma (HCC) from regions with high dietary exposure to aflatoxins and endemic for hepatitis B virus (HBV) often contain a specific mutation at codon 249 in TP53 (249(ser); AGG to AGT, Arg to Ser). This mutation is also detectable in circulating cell-free DNA from the plasma of HCC patients and healthy subjects in these regions. We have examined the joint effect of plasma 249(ser) and HBV infection in a case-control study design involving 348 control, 98 cirrhotic, and 186 HCC participants from The Gambia, West Africa, an area of high HCC incidence. The 249(ser) mutation was detected in 3.5% of controls, 15.3% of cirrhotics, and 39.8% of HCC cases (adjusted odds ratios (OR): 4.83, (95% confidence interval (CI): 1.71-13.7) for cirrhosis and 20.3 (8.19-50.0) for HCC). HBsAg positivity along with plasma 249(ser) was observed in 45/183 (24.6%) HCC cases compared to only one (0.3%) control. Risk for HCC was associated with markers of HBV alone (OR: 10.0, 95% CI: 5.16-19.6), 249(ser) alone (OR: 13.2, 95% CI: 4.99-35.0), and both markers present (OR: 399, 95% CI: 48.6-3270). These results suggest a multiplicative effect on HCC risk resulting from the mutational effect of aflatoxin on TP53, as monitored by detection of plasma 249(ser), with concomitant chronic infection with HBV.