Background:The incidence of anal cancer is increasing globally. Evidence-based improvement in early detection and management of this morbid cancer is thus required. In other cancers associated with Human Papillomavirus (HPV), viral status and dynamics, including viral load (VL) has been shown to influence clinical outcome. Our aim was to determine the influence of HPV status and HPV16 VL on the clinical outcomes of anal cancer patients.Methods: A total of 185 anal cancer lesions were genotyped for HPV. Of the HPV16 positive component, VL was determined using a digital droplet PCR assay.The association of qualitative HPV status and VL (low (<12.3), medium (12.3-57) and high (>57 copies/cell)) on overall survival and hazard of death was assessed.Results: Of the 185 cases, 164 (88.6%) samples were HPV positive. HPV16 was detected in 154/185 samples (83.2%). HPV positive status was associated with improved overall survival in the univariate analysis [hazard ratio (HR) of 0.44, 0.23-0.82, p = 0.01]. When adjusted by age, sex, stage and response to treatment, the association of positive HPV status with improved survival remained (HR 0.24 [0.11-0.55] p < 0.001). High VL was associated with improved overall survival in the univariate analysis with a HR of 0.28 (0.11-0.71, p = 0.007). When adjusted only by age and sex, high VL was associated with better overall survival (HR 0.27, 0.11-0.68 p = 0.006).Conclusions: HPV status appears to be independently associated with improved outcomes in anal cancer patients. Moreover, HPV viral load quantification may be informative for further risk stratification and warrants further investigation.