results from a cross-sectional study using respondent-driven sampling in eight German cities (2011–14)

Background People who inject drugs (PWID) are at increased risk of acquiring and transmitting HIV and Hepatitis C (HCV) due to sharing injection paraphernalia and unprotected sex. To generate seroprevalence data on HIV and HCV among PWID and related data on risk behaviour, a multicentre sero- and behavioural survey using respondent driven sampling (RDS) was conducted in eight German cities between 2011 and 2014. We also evaluated the feasibility and effectiveness of RDS for recruiting PWID in the study cities. Methods Eligible for participation were people who had injected drugs within the last 12 months, were 16 years or older, and who consumed in one of the study cities. Participants were recruited, using low-threshold drop-in facilities as study sites. Initial seeds were selected to represent various sub-groups of people who inject drugs (PWID). Participants completed a face-to-face interview with a structured questionnaire about socio-demographics, sexual and injecting risk behaviours, as well as the utilisation of health services. Capillary blood samples were collected as dried blood spots and were anonymously tested for serological and molecular markers of HIV and HCV. The results are shown as range of proportions (min. and max. values (%)) in the respective study cities. For evaluation of the sampling method we applied criteria from the STROBE guidelines. Results Overall, 2,077 PWID were recruited. The range of age medians was 29–41 years, 18.5–35.3 % of participants were female, and 9.2–30.6 % were foreign born. Median time span since first injection were 10–18 years. Injecting during the last 30 days was reported by 76.0–88.4 % of participants. Sharing needle/syringes (last 30 days) ranged between 4.7 and 22.3 %, while sharing unsterile paraphernalia (spoon, filter, water, last 30 days) was reported by 33.0–43.8 %. A majority of participants (72.8–85.8 %) reported incarceration at least once, and 17.8–39.8 % had injected while incarcerated. Between 30.8 and 66.2 % were currently in opioid substitution therapy. Unweighted HIV seroprevalence ranged from 0–9.1 %, HCV from 42.3–75.0 %, and HCV-RNA from 23.1–54.0 %. The implementation of RDS as a recruiting method in cooperation with low-threshold drop in facilities was well accepted by both staff and PWID. We reached our targeted sample size in seven of eight cities. Conclusions In the recruited sample of mostly current injectors with a long duration of injecting drug use, seroprevalence for HIV and HCV varied greatly between the city samples. HCV was endemic among participants in all city samples. Our results demonstrate the necessity of intensified prevention strategies for blood-borne infections among PWID in Germany

Emergence and Persistence of Minor Drug-Resistant HIV-1 Variants in Ugandan Women after Nevirapine Single-Dose Prophylaxis

BACKGROUND: Nevirapine (NVP) single-dose is still a widely used antiretroviral prophylaxis for the prevention of vertical HIV-1 transmission in resource-limited settings. However, the main disadvantage of the Non-nucleoside Reverse Transcriptase Inhibitor (NNRTI) NVP is the rapid selection of NVP-resistant virus with negative implications for subsequent NNRTI-based long-term antiretroviral therapy (ART). Here, we analysed the emergence of drug-resistant HIV-1 including minor variants in the early phase after NVP single-dose prophylaxis and the persistence of drug-resistant virus over time. METHODS AND FINDINGS: NVP-resistant HIV-1 harbouring the K103N and/or Y181C resistance mutations in the HIV-1 reverse transcriptase gene was measured from 1 week up to 18 months after NVP single-dose prophylaxis in 29 Ugandan women using allele-specific PCR assays capable of detecting drug-resistant variants representing less than 1% of the whole viral population. In total, drug-resistant HIV-1 was identified in 18/29 (62%) women; rates increased from 18% to 38% and 44% at week 1, 2, 6, respectively, and decreased to 18%, 25%, 13% and 4% at month 3, 6, 12 and 18, respectively. The proportion of NVP-resistant virus of the total viral population was significantly higher in women infected with subtype D (median 40.5%) as compared to subtype A (median 1.3%; p = 0.032, Mann-Whitney U test). 33% of resistant virus was not detectable at week 2 but was for the first time measurable 6-12 weeks after NVP single-dose prophylaxis. Three (10%) women harboured resistant virus in proportions >10% still at month 6. CONCLUSIONS: Current WHO guidelines recommend an additional postnatal intake of AZT and 3TC for one week to avoid NVP resistance formation. Our findings indicate that a 1-week medication might be too short to impede the emergence of NVP resistance in a substantial proportion of women. Furthermore, subsequent NNRTI-based ART should not be started earlier than 12 months after NVP single-dose prophylaxis

Prevalence of Transmitted Drug Resistance and Impact of Transmitted Resistance on Treatment Success in the German HIV-1 Seroconverter Cohort

BACKGROUND: The aim of this study is to analyse the prevalence of transmitted drug resistance, TDR, and the impact of TDR on treatment success in the German HIV-1 Seroconverter Cohort. METHODS: Genotypic resistance analysis was performed in treatment-naïve study patients whose sample was available 1,312/1,564 (83.9% October 2008). A genotypic resistance result was obtained for 1,276/1,312 (97.3%). The resistance associated mutations were identified according to the surveillance drug resistance mutations list recommended for drug-naïve patients. Treatment success was determined as viral suppression below 500 copies/ml. RESULTS: Prevalence of TDR was stable at a high level between 1996 and 2007 in the German HIV-1 Seroconverter Cohort (N = 158/1,276; 12.4%; CI(wilson) 10.7-14.3; p(for trend) = 0.25). NRTI resistance was predominant (7.5%) but decreased significantly over time (CI(Wilson): 6.2-9.1, p(for trend) = 0.02). NNRTI resistance tended to increase over time (NNRTI: 3.5%; CI(Wilson): 2.6-4.6; p(for trend)= 0.07), whereas PI resistance remained stable (PI: 3.0%; CI(Wilson): 2.1-4.0; p(for trend) = 0.24). Resistance to all drug classes was frequently caused by singleton resistance mutations (NRTI 55.6%, PI 68.4%, NNRTI 99.1%). The majority of NRTI-resistant strains (79.8%) carried resistance-associated mutations selected by the thymidine analogues zidovudine and stavudine. Preferably 2NRTI/1PIr combinations were prescribed as first line regimen in patients with resistant HIV as well as in patients with susceptible strains (susceptible 45.3%; 173/382 vs. resistant 65.5%; 40/61). The majority of patients in both groups were treated successfully within the first year after ART-initiation (susceptible: 89.9%; 62/69; resistant: 7/9; 77.8%). CONCLUSION: Overall prevalence of TDR remained stable at a high level but trends of resistance against drug classes differed over time. The significant decrease of NRTI-resistance in patients newly infected with HIV might be related to the introduction of novel antiretroviral drugs and a wider use of genotypic resistance analysis prior to treatment initiation

Emergence of Minor Drug-Resistant HIV-1 Variants after Triple Antiretroviral Prophylaxis for Prevention of Vertical HIV-1 Transmission

Background: WHO-guidelines for prevention of mother-to-child transmission of HIV-1 in resource-limited settings recommend complex maternal antiretroviral prophylaxis comprising antenatal zidovudine (AZT), nevirapine single-dose (NVP-SD) at labor onset and AZT/lamivudine (3TC) during labor and one week postpartum. Data on resistance development selected by this regimen is not available. We therefore analyzed the emergence of minor drug-resistant HIV-1 variants in Tanzanian women following complex prophylaxis. Method: 1395 pregnant women were tested for HIV-1 at Kyela District Hospital, Tanzania. 87/202 HIV-positive women started complex prophylaxis. Blood samples were collected before start of prophylaxis, at birth and 1–2, 4–6 and 12–16 weeks postpartum. Allele-specific real-time PCR assays specific for HIV-1 subtypes A, C and D were developed and applied on samples of mothers and their vertically infected infants to quantify key resistance mutations of AZT (K70R/T215Y/T215F), NVP (K103N/Y181C) and 3TC (M184V) at detection limits of,1%. Results: 50/87 HIV-infected women having started complex prophylaxis were eligible for the study. All women took AZT with a median duration of 53 days (IQR 39–64); all women ingested NVP-SD, 86 % took 3TC. HIV-1 resistance mutations were detected in 20/50 (40%) women, of which 70 % displayed minority species. Variants with AZT-resistance mutations were found in 11/50 (22%), NVP-resistant variants in 9/50 (18%) and 3TC-resistant variants in 4/50 women (8%). Three wome

Minor Drug-Resistant HIV Type-1 Variants in Breast Milk and Plasma of HIV Type-1-Infected Ugandan Women after Nevirapine Single-Dose Prophylaxis

Background Nevirapine single-dose (NVP-SD) reduces mother-to-child transmission of HIV type-1 (HIV-1), but frequently induces resistance mutations in the HIV-1 genome. Little is known about drug-resistant HIV-1 variants in the breast milk of women who have taken NVP-SD. Methods Blood and breast milk samples of 39 HIV-1-infected Ugandan women were taken 6–12 weeks after NVP-SD intake. Samples were analysed by population sequencing and allele-specific real-time PCR (AS-PCR) with detection limits for NVP-resistant HIV-1 variants (K103N and Y181C) of &lt;1% of the total viral population. Results AS-PCR results for both plasma and breast milk were obtained for 19 women who constituted the final study group (HIV-1 subtype frequencies were A1 n=11, D n=5, G n=2 and C n=1). A total of 7 (37%) and 10 (53%) women carried NVP-resistant virus in breast milk and plasma, respectively. Overall, 71% (5/7) women with NVP-resistant HIV-1 in breast milk displayed &gt;1 drug-resistant variant. Resistance in breast milk was higher at week 6 (6/13 samples [46%]) compared with week 12 (1/6 samples [17%]). In total, 10 drug-resistant populations harbouring the K103N and/or Y181C mutation were detected in the 19 breast milk samples; 7 (70%) were caused by resistant minorities (&lt;5% of the total HIV-1 population). In the four women with drug-resistant virus in both plasma and breast milk, the mutation patterns differed between the two compartments. Conclusions Minor populations of drug-resistant HIV-1 were frequently found in breast milk of Ugandan women after exposure to NVP-SD. Further studies need to explore the role of minor drug-resistant variants in the postnatal transmission of (resistant) HIV-1. </jats:sec

Detection and Quantification of Minor Human Immunodeficiency Virus Type 1 Variants Harboring K103N and Y181C Resistance Mutations in Subtype A and D Isolates by Allele-Specific Real-Time PCR▿

Nevirapine (single dose), commonly used to prevent the mother-to-child transmission of human immunodeficiency virus (HIV) in developing countries, frequently induces viral resistance. Even mutations which occur only in a minor population of the HIV quasispecies (<20%) are associated with subsequent treatment failure but cannot be detected by population-based sequencing. We developed sensitive allele-specific real-time PCR (ASPCR) assays for two key resistance mutations of nevirapine. The assays were specifically designed to analyze HIV-1 subtype A and D isolates accounting for the majority of HIV infections in Uganda. Assays were evaluated using DNA standards and clinical samples of Ugandan women having preventively taken single-dose nevirapine. Lower detection limits of drug-resistant HIV type 1 (HIV-1) variants carrying reverse transcriptase mutations were 0.019% (K103N [AAC]), 0.013% (K103N [AAT]), and 0.29% (Y181C [TGT]), respectively. Accuracy and precision were high, with coefficients of variation (the standard ratio divided by the mean) of 0.02 to 0.15 for intra-assay variability and those of 0.07 to 0.15 (K103N) and 0.28 to 0.52 (Y181C) for inter-assay variability. ASPCR assays enabled the additional identification of 12 (20%) minor drug-resistant HIV variants in the 20 clinical Ugandan samples (3 mutation analyses per patient; 60 analyses in total) which were not detectable by population-based sequencing. The individual patient cutoff derived from the clinical baseline sample was more appropriate than the standard-based cutoff from cloned DNA. The latter is a suitable alternative since the presence/absence of drug-resistant HIV-1 strains was concordantly identified in 92% (55/60) of the analyses. These assays are useful to monitor the emergence and persistence of drug-resistant HIV-1 variants in subjects infected with HIV-1 subtypes A and D

Impact of HIV-1 Subtype on CD4 Count at HIV Seroconversion, Rate of Decline, and Viral Load Set Point in European Seroconverter Cohorts

Background. Human immunodeficiency virus type 1 (HIV-1) subtype may influence disease progression. We compared CD4 lymphocyte cell count levels at seroconversion, decline rates and viral load set point in individuals infected with different HIV-1 subtypes. Methods. We used data from the Concerted Action on SeroConversion to AIDS and Death in Europe (CASCADE) collaboration, restricted to those infected since 1996, aged &gt;= 15 years, and applied mixed effects models for CD4 cell count decline and median regression for viral load set point (mean level 6-24 months from seroconversion). Results. The analysis included 3364 seroconverters with known HIV-1 subtypes. Compared with subtype B, CD4 at seroconversion was significantly higher for subtype CRF01 and lower for subtype C. Subsequent CD4 decline was significantly slower for subtypes A and CRF02 and marginally slower for subtype C compared with B. Mean CD4 loss at 2 years of seroconversion for white men exposed through sex between men, aged 30-39 years, having seroconverted since 2006, enrolled within 6 months of seroconversion, and without acute infection was 88, 142, 100, 130, 103, and 167 cells/mu L for subtypes A, B, C, CRF01_AE, CRF02_AG, and G, respectively. In adjusted analysis, median viral load set point and time to clinical AIDS/death did not differ significantly by subtype, although all subtypes, except C, tended to have lower levels compared with B. Conclusions. HIV-1 subtype significantly influences seroconversion CD4 cell levels and decline rates but not viral load set point. These findings may be helpful to HIV-positive individuals and their attending physicians in understanding disease progression