Successful application of indirect in-situ polymerase chain reaction to tissues fixed in Bouin's solution.

AIMS: To evaluate the value of polymerase chain reaction-in situ hybridization (PCR-ISH) for the detection of human papillomaviruses (HPV) in paraffin sections of cervical biopsies fixed either in 10% formalin or in Bouin's solution. METHODS AND RESULTS: We analysed 40 biopsies from Italian women infected with the human immunodeficiency virus type 1 (HIV 1). In-situ hybridization techniques were performed with commercial biotinylated probes. The PCR-ISH was carried out by the 'hot start modification'. Cervical intraepithelial neoplasia (CIN) was found in 23 of 40 patients (57. 5%); eight cases showed condylomatous features. Human papillomavirus was detected in 42.5% by ISH and in 65% by PCR-ISH. Sixty-nine per cent of positive biopsies contained HPV 16, 18, 31 and 33. HPV 6 and 11 were found only in condylomata acuminata samples. CONCLUSIONS: The results point to a high incidence of HPV infection as well as of CIN in HIV-positive patients. Human papillomavirus type 16 appears to be most frequently associated with CIN. Polymerase chain reaction-ISH is more sensitive than ISH in the detection and typing of HPV DNA both in clinical and in 'latent' infections. The two techniques yielded the same results with either formalin- or Bouin's-fixed material.

The use of carbon-dioxide laser surgery in the treatment of intraepithelial neoplasia of the uterine cervix.

From October 1989 to June 1997, 1841 patients (pts) suffering from different diseases of the lower genital tract have been treated with CO2 laser surgery in our Institution: among them, 782 were affected by cervical intraepithelial neoplasia (CIN). All pts underwent CO2 laser procedure for CIN after adequate colposcopic evaluation of the entire lower genital tract, colposcopic guided biopsy of the lesion, adequate pre-surgical work-up for possible infectious and coagulation associated disease. In 736 (94.1%) pts, the procedure was performed on an ambulatory basis while 46 pts (5.9%) were admitted for 1 or 2 days. A CO2 laser Sharplan 55 associated to a Zeiss operative colposcope was employed. The preoperative diagnosis of the 782 pts treated for CIN was 297 CIN1, 255 CIN2, 171 CIN3 and 59 CIS. Mean age was 33.6 years without statistical difference among the grade of disease: 605 pts underwent laser vaporization according to specific selection criteria. The depth of cervical destruction was less than 6 mm in 26 cases, between 6 and 10 mm in 549, between 11 and 15 mm in 157, more than 15 mm in 50 pts. 742 procedures were performed under local anesthesia and pain was absent in 667 pts. (89.9%), mild in 51 (6.8%), moderate in 19 (2.5%) and severe in 5 (0.7%). Intraoperative bleeding was severe in 30 pts. (3.8%), moderate in 77 (9.8%), mild in 204 (26.1%) and absent in 471 (60.2%). The conization procedure was shown to have a higher risk of bleeding but no direct relation was observed with the depth of cervical destruction. Late complications were scarce: 1.3% of late hemorrhages, 1.4% of stenosis of cervical external orifice and cervical endometriosis in 0.3%. In 76 pts (42%) of the 177 conizations the final pathology report was in accordance with the previous biopsy, in 56 (30.9%) a lower grade of CIN was observed, in 53 (29.3%) a worse grade of the lesion was retrieved. Among these latter pts. 10 showed a microinvasive and 2 an invasive cancer: both the invasive but only 3 of the 10 microinvasive cancer pts underwent a surgical procedure (2 radical and 3 extrafascial hysterectomies, respectively). After a mean follow up of 37 months our incidence of recurrence is 2.3% (18 pts): 5 CIN1, 7 CIN2, 3 CIN3, 2 CIS and 1 microinvasive disease. In 78% of the instances the recurrence has occurred within the first year of follow up. All 18 recurrences were successfully treated with further vaporization in 8 cases, conization in 9 and hysterectomy in 1. 93 term pregnancies occurred in 83 pts after CO2 laser treatment of CIN. No cervical incompetence occurred (no cervical cerclage employed) while the incidence of spontaneous abortion was not statistically significant. 82 pregnancies were delivered vaginally without significant variation of labor phase duration. The incidence of caesarian section (11.8%) was lower than the mean incidence in our Institution. These data confirm the successful complete restitutio ad integrum of the cervix after an adequate CO2 laser surgical procedure without any further risk of cervical incompetence, premature delivery or premature rupture of membranes. The use of CO2 laser surgery is recommended as modality treatment of choice in the management of cervical intraepithelial neoplasia.

Influence of maternal CD4 levels on the predictive value of virus load over mother-to-child transmission of human immunodeficiency virus type 1 (HIV-1). Study Group for Vertical Transmission.

Forty-four anti-HIV seropositive pregnant women were enrolled in a study of maternal factors related to mother-to-infant human immunodeficiency virus type 1 (HIV-1) transmission. HIV-1 infection was documented in 11 of 45 infants (24.4%). Obstetric factors, maternal CD4 counts, and disease stage were not related to the risk of transmission. HIV-1 RNA levels at delivery were significantly higher in mothers who transmitted the infection (P = .024). A strong relationship between viral load and risk of transmission was observed in women with stage A1 (P= .006), but not in those with stages A2-A3. These results suggest that vertical transmission of HIV-1 is multifactorial and that viral load plays a major role in mothers with early-stage HIV-1 infection.

Prospective study of mother-to-infant transmission of hepatitis C virus (HCV) infection. Study Group for Vertical Transmission.

Seventy-five women with anti-hepatitis C virus (HCV) antibody were enrolled prospectively during pregnancy or at delivery for study of mother-to-child transmission of HCV. Twenty-three women were coinfected with the human immunodeficiency virus (HIV). Seventy babies were monitored for at least 6 months. HCV infection was diagnosed in six infants (8.6%), four of whom were born to anti-HIV-positive mothers. HCV RNA was first detected between 2 and 6 months, and the genotypes of infected babies matched those of their mothers (type 1: n = 4; type 3: n = 2). Identical master sequences of the hypervariable region (HVR1) were detected in a mother-infant pair. In three babies coinfected with HCV and HIV, anti-HCV disappeared between 2 and 7 months, being persistently negative in two cases monitored for 11 and 26 months. Transmitting mothers did not differ significantly from those who did not transmit the infection with anti-HIV, HCV genotypes, and viral load at delivery, but had lower rate of reactivity to C100 by the recombinant immunoblot assay (RIBA) (P < .01). This prospective study confirms transmission of HCV from anti-HIV-negative mothers (4.4% in this series). Absence of anti-C100 antibodies at delivery is apparently related to increased risk of vertical transmission. Seronegative HCV infection can be observed in children coinfected with HIV.