The impact of HBV or HCV infection in a cohort of HIV-infected pregnant women receiving a nevirapine-based antiretroviral regimen in Malawi.

BACKGROUND: Coinfection with the hepatitis viruses is common in the HIV population in sub-Saharan Africa. The aim of this study was to assess, in a cohort of HIV-infected pregnant women receiving antiretroviral drugs (ARVs), the prevalence of HBV and HCV infections and to determine the impact of these infections on the occurrence of liver toxicity and on the viro-immunological response. METHODS: Women were screened for HBsAg and HCV-RNA before starting, at week 25 of gestational age, an antiretroviral regimen consisting of lamivudine and nevirapine plus either stavudine or zidovudine. Women with CD4+ < 350/mm3 continued ARVs indefinitely, while the other women interrupted treatment 6 months postpartum (end of breastfeeding period). Both groups were followed for 2 years after delivery. Liver function was monitored by alanine aminotransferase (ALT) measurement. The Cox proportional hazards model was used to identify factors associated with the emergence of liver toxicity. RESULTS: A total of 28 women out of the 309 enrolled in the study (9.1%) were coinfected with HBV (n. 27), or HCV (n. 1). During follow-up 125 women (40.4%) developed a grade ≥ 1 ALT elevation, 28 (9.1%) a grade ≥ 2 and 6 (1.9%) an elevation defining grade 3 toxicity. In a multivariate model including age, baseline CD4+ count and hemoglobin level, the presence of either HBV or HCV infection was significantly associated with the development of an ALT increase of any grade (P = 0.035). Moderate or severe liver laboratory toxicity (grade ≥ 2) was more frequent among women with baseline CD4+ > 250/mm3 (P = 0.030). In HBV-infected women a baseline HBV-DNA level above 10,000 IU/ml was significantly associated to the development of liver toxicity of grade ≥ 1 (P = 0.040). Coinfections had no impact on the immunological and virological response to antiretroviral drugs up to 2 years after delivery. CONCLUSIONS: In this cohort of nevirapine-treated women the presence of HBV or HCV was associated only to the development of mild liver toxicity, while the occurrence of moderate or severe hepatoxicity was correlated to a baseline CD4+ count > 250/mm3. No statistically significant effect of the coinfections was observed on the efficacy of antiretroviral therapy.

Quantification of HIV-RNA from dried blood spots using the Siemens VERSANT® HIV-1 RNA (kPCR) assay.

OBJECTIVES: Simplified methods for virological monitoring in resource-limited settings are increasingly needed. We evaluated the performance of the VERSANT(®) HIV-1 RNA (kPCR) assay for the determination of HIV-1 viral load from dried blood spots (DBS). Assay sensitivity and correlation with plasma quantification values were assessed. METHODS: A total of 98 DBS were prepared from fresh blood samples of HIV-infected patients. DBS were kept at room temperature for 6 weeks or 7 months before processing while the corresponding plasma samples were stored at -80°C. DBS were first pre-treated in a special DBS buffer. The DBS extracts and the plasma samples were then purified and amplified using the VERSANT assay reagents. RESULTS: In the first series of tests, performed after 6 weeks of storage, there was good correlation between quantification of viral load in plasma and in DBS (r = 0.95, P < 0.001). The detection rate in DBS was 100% when plasma levels were >1000 copies/mL. The sensitivity and specificity of the DBS assay were 88.2% [95% confidence interval (CI) 79.4-93.6] and 69.2% (95% CI 42.0-87.4), respectively. Using the 5000 copies/mL threshold (defining virological failure in resource-limited settings), both positive and negative predictive values were high (95.2% and 87.5%, respectively). After 7 months of storage there was a modest decrease in the detection rate and less significant correlations for samples with HIV-RNA <5000 copies/mL. CONCLUSIONS: Quantification of HIV-RNA from DBS by the VERSANT automated sample preparation and detection method can be used to diagnose virological failure in HIV-positive patients.

A simplified way for the urgent treatment of somatic pain in patients admitted to the emergency room: the SUPER algorithm.

Somatic pain is one of the most frequent symptoms reported by patients presenting to the emergency department (ED), but, in spite of this, it is very often underestimated and under-treated. Moreover, pain-killers prescriptions are usually related to the medical examination, leading to a delay in its administration, thus worsening the patient's quality of life. With our study, we want to define and validate a systematic and homogeneous approach to analgesic drugs administration, testing a new therapeutic algorithm in terms of earliness, safety, and efficacy. 442 consecutive patients who accessed our ED for any kind of somatic pain were enrolled, and then randomly divided into two groups: group A follow the normal process of access to pain-control drugs, and group B follow our SUPER algorithm for early administration of drugs to relieve pain directly from triage. We excluded from the study, patients with abdominal pain referred to the surgeon, patients with headache, recent history of trauma, history of drug allergies, and life-threatening conditions or lack of cooperation. Drugs used in the study were those available in our ED, such as paracetamol, paracetamol/codeine, ketorolac-tromethamine, and tramadol-hydrochloride. Pain level, risk factors, indication, and contraindication of each drug were taken into account in our SUPER algorithm for a rapid and safe administration of it. The Verbal Numeric Scale (VNS) and the Visual Analog Scale (VAS) were used to verify the patient's health and perception of it. Only 59 patient from group A (27.1 %) received analgesic therapy (at the time of the medical examination) compared to 181 patients (100 %) of group B (p < 0.001). Group B patients, received analgesic therapy 76 min before group A subjects (p < 0.01), resulting in a significant lower VNS (7.31 ± 1.68 vs 4.75 ± 2.3; p < 0.001), and a superior VAS after discharge (54.43 ± 22.16 vs 61.30 ± 19.13; p < 0.001) compared to group A subjects. No significant differences concerning side effects were observed between group A and group B patients. Early administration of a pain-control therapy directly from triage is safe and effective, and significantly improves patients perceptions of their own health.

Correlation between HIV-1 viral load quantification in plasma, dried blood spots, and dried plasma spots using the Roche COBAS Taqman assay.

BACKGROUND: The use of simplified methods for viral load determination could greatly increase access to treatment monitoring of HIV patients in resource-limited countries. OBJECTIVE: The aim of the present study was to optimize and evaluate the performance of the Roche COBAS Taqman assay in HIV-RNA quantification from dried blood spots (DBS) and dried plasma spots (DPS). STUDY DESIGN: EDTA blood samples from 108 HIV-infected women were used to prepare 129 DBS and 76 DPS on Whatman 903 card. DBS and DPS were stored at -20 degrees C. HIV-1 RNA was extracted from DBS/DPS using the MiniMAG system (bioMerieux). Amplification and detection were performed using the Roche COBAS TaqMan assay. Plasma viral load results were used as standard. RESULTS: There was a high correlation between measures of viral load in plasma and in DBS/DPS (r=0.96 and 0.85 respectively, P<0.001). Overall, viral load values in DBS and DPS tended to be lower than in plasma with mean (SD) differences of 0.32 log(0.22) for DBS and of 0.35 (0.33) for DPS. Detection rates were 96.4% for DBS and 96.1% for DPS in samples with corresponding plasma values >3.0 log copies/ml. Samples with HIV-RNA below 50 copies/ml were correctly identified in 18/19 DBS and in 7/7 DPS. CONCLUSIONS: Both DBS and DPS provided results highly correlated to the plasma values. High detection rate was obtained with both DBS and DPS when HIV-RNA was >3.0 log copies/ml. Our results support the use of DBS/DPS to detect virologic failure in resource-limited settings.

Microbial translocation is associated with residual viral replication in HAART-treated HIV+ subjects with <50copies/ml HIV-1 RNA.

BACKGROUND: Recent data have shown that plasma levels of lipopolysaccharide (LPS) are a quantitative indicator of microbial translocation in HIV infected individuals. OBJECTIVES: To assess the impact of residual viral replication on plasma LPS in HAART-treated HIV+ subjects with <50copies/ml HIV-1 RNA and to evaluate LPS changes during repeated HAART interruptions not exceeding 2-month duration. STUDY DESIGN: LPS was measured in 44 HIV+ subjects at T0 (during HAART) and at day 15 of the first and fourth HAART interruption. Ten uninfected, healthy donors were studied as well. Residual plasma HIV-1 RNA was measured at T0 by an ultra-ultrasensitive method with limit of detection of 2.5copies HIV-1 RNA/ml. Subjects with less than 2.5copies/ml (fully suppressed - FS) were compared to those with 2.5-50copies/ml (partially suppressed - PS). RESULTS: At T0, plasma LPS levels were comparable in FS and uninfected subjects, whereas in PS they were higher than in uninfected subjects (p=0.049). After 4 HAART interruptions, they did not change significantly. However, LPS values were lower in FS than in PS (p=0.020). An inverse correlation was found between CD4 and LPS levels (p=0.044) in PS group only. CONCLUSIONS: A reduced degree of microbial translocation was seen in subjects with a more complete suppression of viral replication. Repeated HAART interruptions had no significant impact on plasma LPS levels.

Seroprevalence of hepatitis B and C viruses among HIV-infected pregnant women in Uganda and Rwanda.

A retrospective survey to estimate the prevalence of hepatitis B (HBV) and C (HCV) infections was conducted on the samples of 247 African HIV-1 positive pregnant women who had participated to a mother-to-child prevention trial carried out in urban settings in Kampala, Uganda and Kigali, Rwanda. Hepatitis B markers studied were HBs antigen (HBsAg) and, if positive after confirmatory testing, HBe antigen/anti-HBe antibodies and HBV DNA. A fourth generation HCV enzyme immunoassay (EIA) was used for primary HCV screening. Positive samples were analyzed further with a second different EIA. Both for HBV and for HCV the use of confirmatory tests allowed the removal of frequent false-positive screening results. HBsAg was found in 10/246 women (seroprevalence 4.1%, 95% confidence interval (95%CI) 1.7-6.8): 8/164 (4.9%) in Uganda and 2/82 (2.4%) in Rwanda. HBe Ag was found in 33% of HBsAg-positive patients and HBV DNA was quantifiable in 71%. Anti-HCV antibodies were found in 5/247 women (seroprevalence 2.0% 95%CI 0.3-3.9): 1/165 (0.6%) in Uganda and 4/82 (4.9%) in Rwanda. There was no interrelation between HCV and HBV markers. There was no difference between patients with and without co-infection with HBV or HCV with regards to CD4+ cell count. Overall, hepatitis B and C co-infection was relatively infrequent in this group of pregnant women. However, since approximately 6% of HIV-positive women in these countries had a co-infection with one hepatitis virus, caution should be used in the monitoring of possible hepatotoxicity related to antiretroviral drugs in these populations.

Triple antiretroviral prophylaxis administered during pregnancy and after delivery significantly reduces breast milk viral load: a study within the Drug Resource Enhancement Against AIDS and Malnutrition Program.

BACKGROUND: The administration of antiretroviral therapy to lactating women could represent a possible strategy to reduce postnatal HIV transmission. In this study, we assessed the effect of antiretroviral treatment on breast milk viral load and determined plasma and breast milk drug concentrations in pregnant women receiving highly active antiretroviral therapy (HAART). METHODS: We studied 40 women receiving zidovudine, lamivudine, and nevirapine from 28 weeks of gestation to 1 month postpartum (group A) and 40 untreated pregnant women (group B). Blood and breast milk samples were collected at delivery and 7 days postpartum. RESULTS: Women in group A had received a median of 85 days of therapy before delivery. Median breast milk concentrations of nevirapine, lamivudine, and zidovudine were 0.6, 1.8, and 1.1 times, respectively, those in maternal plasma. HIV RNA levels in breast milk were significantly lower in group A than in group B (median of 2.3 vs. 3.4 log at delivery and 1.9 vs. 3.6 log at day 7; P < 0.001 for both comparisons). CONCLUSIONS: Antiretroviral drugs administered during the last trimester of pregnancy and after delivery reach levels similar to or higher than plasma concentrations in breast milk and can significantly reduce HIV RNA levels. Our data support the potential role of maternal HAART prophylaxis in reducing the risk of breast-feeding-associated transmission.

Determinants of virologic and immunologic outcomes in chronically HIV-infected subjects undergoing repeated treatment interruptions: the Istituto Superiore di Sanita-Pulsed Antiretroviral Therapy (ISS-PART) study.

BACKGROUND: Factors influencing the outcome of structured treatment interruptions (STIs) in HIV chronic infection are not fully elucidated. METHODS: In ISS-PART, 273 subjects were randomly assigned to arm A (137 assigned to continuous highly active antiretroviral therapy [HAART]) and arm B (136 assigned to 5 STIs of 1, 1, 2, 2, and 3 months' duration, each followed by 3 months of therapy). Main outcome measures were the proportion of subjects with a CD4 count >500 cells/mm3, the rate of virologic failure, and the emergence of resistance at 24 months. RESULTS: The proportion of subjects with a CD4 count >500 cells/mm3 was higher in arm A than in arm B (86.5% vs. 69.1%; P = 0.0075). Pre-HAART CD4 cell count and male gender were independent predictors of a CD4 count >500 cells/mm3 in arm B. The overall risk of virologic failure was not increased in arm B; however, it was higher in the 38 subjects who had resistance mutations in the rebounding virus. Archived mutations at baseline and the use of a regimen that included an unboosted protease inhibitor (PI), compared with nonnucleoside reverse transcriptase inhibitor-based HAART, independently predicted the emergence of plasma mutations during STI (P = 0.002 for DNA mutations and P = 0.048 for PI-based HAART). CONCLUSIONS: Our results suggest that patients with preexisting mutations and treated with unboosted PI-based HAART should not be enrolled in studies of time-fixed treatment interruptions, being at higher risk of developing plasma mutations during STI and virologic failure at therapy reinstitution.

Development of a human immunodeficiency virus vector-based, single-cycle assay for evaluation of anti-integrase compounds.

Therapeutic strategies aimed at inhibiting human immunodeficiency virus type 1 (HIV-1) replication employ a combination of drugs targeted to two viral enzymes (reverse transcriptase and protease) and to the viral entry/fusion step. However, the high propensity of HIV-1 to develop resistance makes the development of novel compounds targeting different steps of the HIV-1 life cycle essential. Among these, integrase (IN) inhibitors have successfully passed the early phases of clinical development. By preventing integration, IN inhibitors preclude viral replication while allowing production of extrachromosomal forms of viral DNA (E-DNA). Here, we describe an improved and standardized assay aimed at evaluating IN inhibitors by taking advantage of the transcriptional activity of E-DNA produced by HIV-derived vectors in the absence of replication-competent virus. In this context, the use of the firefly luciferase gene as a reporter gene provides a rapid and quantitative measure of viral-vector infectivity, thus making it a safe and cost-effective assay for evaluating novel IN inhibitors.