It has been well established that human papilloma virus (HPV) is the major cause of cervical pre-cancerous lesions and cervical cancer. Extended HPV genotyping has pointed out that co-infections with multiple high-risk (HR)-HPV genotypes not only is possible and quite frequent, but also has different prognoses. The purpose of this study was to evaluate the prevalence of co-infections in women tested for HR-HPV in the national cervical cancer screening program of Lazio (Italy). From June 1st to November 30th 2022, we analyzed 30,445 samples of women aged between 30 and 64 years, using the Anyplex TM II HPV HR Detection test by Seegene (Arrow), which identifies 14 HPV genotypes: 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, 68. The data were analyzed using the SG STATS platform. In total, 4,244 (13.94%) were positive: 3,290 (77.52%) showed a single genotype infection and 954 (22.48%) an infection with 2 to 5 different genotypes. In 721 (75.60%) cases, two different genotypes were detected, in 191 (20.00%) there were three genotypes, in 41 (4.30%) cases there were four genotypes and in only one case (0.10%) five different genotypes were detected. HPV 16 (262 cases of co-infections) was associated in 27 cases with HPV 31 genotype, in 25 cases with HPV 68 and in 18 cases with HPV 58. HPV 16 was the most frequent genotype detected in co-infections. Immunity status, vaccination, lifestyle, and other possible risk factors, such as the combination of the HR-HPV genotype multiple infections, may influence the development and progression of the disease.