Background: High-risk human papillomavirus (HR-HPV) anal infection is a major problem among men who have sex with men (MSM) living in sub-Saharan Africa. The prevalence of anal HR-HPV infection and associated risk factors were estimated in a cross-sectional study in MSM living in Bamako, Mali. Methods: MSM consulting at sexual health center of the National NGO Soutoura, Bamako, were prospectively included. Sociodemographic and clinical-biological data were collected. HPV detection and genotyping were performed from anal swabs using multiplex real-time PCR. Risk factors associated with anal HPV infection were assessed by logistic regression analysis. Results: Fifty MSM (mean age, 24.2 years; range, 18-35) of which 32.0% were infected with HIV-1, were prospectively included. The overall prevalence of anal HPV infection of any genotypes was 70.0% (35/50) with 80.0% (28/35) of swabs positive for HR-HPV. HR-HPV-58 was the most detected genotype [13/35 (37.1%)], followed by HR-HPV-16 and low-risk (LR)-HPV-6 [12/35 (34.2%)], LR-HPV-40 [10/35 (28.6%)], LR-HPV-11 [9/35 (25.7%)], HR-HPV-51 [8/35 (22.8%)], HR-HPV types 18 and 39 [7/35 (20.0%)] and LR-HPV-43 [6/35 (17.1%)]. HR-HPV-52 and LR-HPV-44 were detected in lower proportions [5/35 (14.3%) and 4/35 (11.4%), respectively]. LR-HPV-42, LR-HPV-54, HR-HPV-31 and HR-HPV-35 were detected in very low proportions [3/35 (8.5%)]. Multiple HR-HPV infections were diagnosed in one-third of anal samples [16/50 (32.0%)], including around half of HR-HPV-positive anal swabs [16/35 (45.7%)]. More than half [27/50 (54.0%)] swabs were infected by at least one of HPV genotypes targeted by Gardasil-9® vaccine, including a majority of vaccine HR-HPV [22/50 (44.0%)]. In multivariate analysis, participation to sex in group was associated with anal infection by multiple HPV (aOR: 4.5, 95% CI: 1.1-18.1%; P = 0.032), and HIV-1 infection was associated with anal shedding of multiple HR-HPV (aOR: 5.5, 95% CI: 1.3-24.5%; P = 0.024). Conclusions: These observations indicate that the MSM community living in Bamako is at high-risk for HR-HPV anal infections, with a unique epidemiological HPV genotypes profile and high prevalence of anal HPV covered by the Gardasil-9® vaccine. Scaling up prevention strategies against HPV infection and related cancers adapted to this highly vulnerable MSM community should be urgently prioritized with innovative interventions.