Viral genotyping gave a positive result for single infections in 360 patients, 387 patients were positive for multiple infections, 127 women were negative for the viral genotypes present in the kit used. Thirty-six samples analyzed, from among all the samples, were inadequate; these patients and the negative women were excluded from the study group.
At the histological exam, carried out after biopsy, 458/747 patients (61.3 %) presented a low-grade preneoplastic lesion (CIN 1); 64 women (8.6 %) had a moderate-grade lesion (CIN 2), 105 women (14 %) had a sever-grade lesion (CIN 3); and 5 women (0.7%) had squamosa carcinomas (SCC). Finally, 115 (15.4%) patients had a negative histological diagnosis (Table 1). In total we had 174 CIN2+ (23.3%) lesions.
1. Single HPV infection
There were 360 (48.2%) single infections in our group with 123 lesions ( 34.1%) CIN2+. The typology of lesions with the highest percentage of a single infection was SCC with 100%, followed by CIN3 with 78%, CIN2 with 56.2% and finally negative and CIN1 with 48.6% and 39.5%, respectively. The genotype most frequently found was genotype 16, present in 142/360 cases (39.4%), and was responsible for 76.4 % (94/123) of the cases of CIN2+, 21.6% of the cases of CIN1 and 16 % of negatives (Table 2).
In the present study the patients with a single infection from HPV 16 had the highest incidence (76.4 %) of CIN2+ with respect to the other HR genotypes (23.5%).
In particular, the presence of genotype HPV16 in our study was associated with a 12 times greater risk of developing a high-grade lesion (CIN2+), OR = 12.70; (IC 95% = 7.52-21.42). After genotype 16, the genotype most represented was 31 with 36 cases, of which 11 CIN2+, while the genotypes 18 and 45 were poorly represented with 17 cases of which 8 CIN2+ and 6 cases of 1 CIN3. Other HR HPV found were 33 with 8 cases, of which 2 CIN3, 35 with 4 cases, of which 2 CIN3 and 52 and 58 both with 1 case of CIN2 . (Table 3). It should be noted that the most frequent genotypes were 16, 31, 33, 35, 52 and 58 belonging to the same group alfa9.
Particular importance needs to be given to genotype 51 that demonstrated with 3 cases of CIN2 and 1 squamous carcinoma to have an oncogenic capacity and also genotype 35, responsible for 2 cases of CIN3. Both 51 and 35 genotypes have not been assembled among the genotypes of the nonovalent vaccines.
2. Multiple HPV infections
There were 387 (51.8%) multiple infections in our study with an incidence of high-grade lesions of 10.6 % (41/387). The typology of the lesion with the highest percentage of multiple infections was CIN1 with a frequency of 60.4% and the negative cases were 50.4%. The percentage decreases more in CIN2 (43.7%) and progressively in CIN3 (22.1 %), arriving at zero in carcinomas, characterized exclusively by single infections (Fig.1). We had no cases of SCC with multiple genotypes.
The four most diffused HPV genotypes were HPV 16, 51, 59 and 31. Also in multiple infections the most frequent genotype was 16 (151/387) with a prevalence of 74.5% (38/51), this was significatively higher in patients with CIN2/CIN3, OR = 5.77 (IC 95% = 2.95-11.26 ). We studied in particular genotype 16 (Table 4). We analyzed the difference between infections of single and multiple types with HPV16 and the different impact that both the infections have on severe dysplasia and carcinomas: the patients with single infections had the highest incidence of CIN2+ (70.1%) with respect to those with multiple infections (29,3%).
OR of the single infections from HPV16 associated with CIN2+ was greater than the OR of multiple HPV16 infections (Table 5).
We divided multiple infection by the number of genotypes present (strains) into 2 strains, 3 strains, 4 strains, 5 strains or more. We found that increasing the histological grade of the lesion, decreases the number of HR genotypes present; in fact, CIN3 lesion has two high-risk genotypes in 82.6% of cases (19/23). the remaining 4 cases (4/23) had three genotypes. We did not have any cases of CIN3 lesion with 4 or 5 genotypes.
We correlated genotype 16 with the number of strains and the histological diagnosis (Table 6). CIN 3 other than representing 82.6 % of two high-risk genotypes, one of these genotypes is almost always genotype 16 (22/23).
When we studied the prevalence of the combinations between the genotypes, we found that the combinations HPV16, 18 and HPV16, 31 were the most frequent (55.5%) in CIN3 (Figure 2).
From the OR analysis, multiple HPV16 infections with 2 high-risk genotypes, with respect to infections with 3 or more genotypes, were significant with an OR = 3.92 (IC% 1.70-9.03) for CIN2+