Congress report: Online workshop on assessment of technical files for blood screening in vitro diagnostics for sub-Sahara African countries.

OBJECTIVES: The online workshop on IVD regulation was performed to broaden the understanding of the technical documentation needed for IVD licensing and the strategies to asses it. BACKGROUND: Testing of blood donors and donations significantly reduces the risk of transmitting transfusion-transmissible infections. Many test systems are commercially available, but not all meet the recommended sensitivity and specificity standards. Many African countries either lack functional structures for the regulation of IVDs this poses a threat to the quality of the blood supply. MATERIALSAND METHODS: The Paul-Ehrlich-Institut BloodTrain organised an online workshop in September 2021 to introduce staff from several National Regulatory Authorities (NRAs) in Africa to the regulation of IVD and the technical information that need to be provided by the manufacturers of blood screening IVD. Their evaluation was trained in practical exercises. RESULTS: This online workshop brought together over hundred participants from NRAs of 12 African countries. Speakers from PEI, Blood Train, WHO and academia, with experience in IVD regulation trained participants in the various topics addressed during this workshop. CONCLUSIONS: This workshop presented a great starting point for most participating NRAs to set up and/or strengthen their regulatory structures for IVDs.

Report on the GHPP-BloodTrain online workshop on "Authorisation and Licensing of Blood Establishments - Review of quality documentation of blood and blood components", 5th-8th of July 2021.

The GHPP BloodTrain team developed an e-learning concept in response to the COVID-19 travel restrictions, providing training formats with virtual stages that can be completed during the pandemic (and beyond) and on-site stages, where practical exercises and case reports in smaller groups can lead to a deeper understanding of the content. The virtual training workshop on "Authorisation and Licensing of Blood Establishments", hosted by the PEI GHPP BloodTrain from the 5th to the 8th of July 2021, was the first application of this concept. The number of participants could be substantially increased compared to an on-site event thanks to the virtual setting. Participants came mainly from national regulatory authorities and national blood transfusion services. There were also some Ministry of Health representatives from 19 countries from the WHO regions of AFRO, EMRO, and from Indonesia in attendance. The virtual workshop focused on reviewing and evaluating the quality documentation required for approval of processes used by blood establishments to prepare blood components. Presentations were given by members of the GHPP BloodTrain team as well as by representatives of the German Red Cross. The program was complemented by contributions from the WHO HQ and presentations on country-specific experiences from Ghana and Zimbabwe.

Prevalence of integrase strand transfer inhibitor resistance mutations in antiretroviral-naive HIV-1-infected individuals in Cameroon.

OBJECTIVES: In Cameroon, the integrase (IN) strand transfer inhibitor (INSTI) dolutegravir was recently introduced for the treatment of HIV-1 infection. Since pretreatment HIV-1 drug resistance can jeopardize the success of ART, and considering the high heterogeneity of circulating HIV-1 subtypes in Cameroon, we investigated the prevalence of pretreatment HIV-1 resistance to INSTIs. METHODS: Fingerprick dried blood spot samples were collected from 339 newly diagnosed HIV-1-infected individuals between 2015 and 2016 in four hospitals in Cameroon. Universal primers were designed to amplify the HIV-1 IN region from amino acid 1 to 276. Amplicons were sequenced with Illumina next-generation sequencing and analysed with the Polymorphism Analysis Sequencing (PASeq) platform, using the Stanford HIV Drug Resistance Database to interpret HIV-1 drug resistance mutations (DRMs). RESULTS: The amplification/sequencing success rate was 75.2% with 255/339 sequences obtained. Applying a cut-off of 1%, major DRMs to INSTIs were detected in 13 (5.1%) individuals, but only 1 individual harboured an INSTI DRM (E92G) at a nucleotide frequency ≥15%. However, 140/255 (54.9%) individuals harboured polymorphic accessory INSTI DRMs, mainly at high frequencies. In line with that observation, HIV-1 subtype diversity among individuals was high. CONCLUSIONS: Pretreatment HIV-1 resistance to INSTIs was low in the study sites, which supports the use of INSTIs in Cameroon. Nevertheless, further studies are necessary to assess the impact of polymorphic accessory INSTI DRMs on INSTI-based ART regimens.

Low-Abundance Drug-Resistant HIV-1 Variants in Antiretroviral Drug-Naive Individuals: A Systematic Review of Detection Methods, Prevalence, and Clinical Impact.

BACKGROUND: The presence of high-abundance drug-resistant HIV-1 jeopardizes success of antiretroviral therapy (ART). Despite numerous investigations, the clinical impact of low-abundance drug-resistant HIV-1 variants (LA-DRVs) at levels <15%-25% of the virus population in antiretroviral (ARV) drug-naive individuals remains controversial. METHODS: We systematically reviewed 103 studies assessing prevalence, detection methods, technical and clinical detection cutoffs, and clinical significance of LA-DRVs in antiretroviral drug-naive adults. RESULTS: In total, 14 919 ARV drug-naive individuals were included. Prevalence of LA-DRVs (ie, proportion of individuals harboring LA-DRVs) was 0%-100%. Technical detection cutoffs showed a 4 log range (0.001%-10%); 42/103 (40.8%) studies investigating the impact of LA-DRVs on ART; 25 studies included only individuals on first-line nonnucleoside reverse transcriptase inhibitor-based ART regimens. Eleven of those 25 studies (44.0%) reported a significantly association between preexisting LA-DRVs and risk of virological failure whereas 14/25 (56.0%) did not. CONCLUSIONS: Comparability of the 103 studies is hampered by high heterogeneity of the studies' designs and use of different methods to detect LA-DRVs. Thus, evaluating clinical impact of LA-DRVs on first-line ART remains challenging. We, the WHO HIVResNet working group, defined central areas of future investigations to guide further efforts to implement ultrasensitive resistance testing in routine settings.

Changing Trends in International Versus Domestic HCV Transmission in HIV-Positive Men Who Have Sex With Men: A Perspective for the Direct-Acting Antiviral Scale-Up Era.

BACKGROUND: Scale-up of direct-acting antiviral therapy is expected to abate hepatitis C virus (HCV) incidence among human immunodeficiency virus (HIV)-positive men who have sex with men (MSM). International transmission could influence this process. We classified HCV infections in HIV-positive MSM as either domestically or internationally acquired, and estimated how this classification changed over time. METHODS: HCV subtype 1a (the most frequent subtype among MSM) genomes from 99 persons enrolled in the Swiss HIV Cohort Study and diagnosed with replicating HCV infections, were sequenced. Sixty-six of these sequences were from MSM. We inferred maximum-likelihood phylogenetic trees and time trees containing a fragment of the NS5B region of these and 374 circulating strains. We inferred transmission clusters from these trees and used the country composition of such clusters to attribute infections to domestic or international transmission. RESULTS: Of HCV transmissions, 50% to 80% were classified as domestic depending on the classification criterion. Between 2000 and 2007, the fraction attributable to domestic transmission was 54% (range 0-75%). It increased to 85% (range 67%-100%) between 2008 and 2016. CONCLUSIONS: International and domestic transmission have played major roles in this epidemic. While international transmission persists, local transmission has established as the main source of infections.

Low prevalence of transmitted HIV-1 drug resistance detected by a dried blood spot (DBS)-based next-generation sequencing (NGS) method in newly diagnosed individuals in Cameroon in the years 2015-16.

Objectives: To determine the most recent prevalence, transmission patterns and risk factors of transmitted drug-resistance mutations (TDRMs) in Cameroon, we initiated a multicentre study monitoring HIV-1 drug resistance in newly HIV-1-diagnosed individuals using a novel next-generation sequencing (NGS) assay applicable to fingerprick dried blood spot (DBS) samples. Methods: Fingerprick DBS samples and questionnaires were collected from 360 newly HIV-1-diagnosed individuals in four hospitals in urban areas in Cameroon in the years 2015-16. We developed an HIV-1 protease and reverse transcriptase drug resistance genotyping assay applicable to DBS samples and HIV-1 genomes of groups M, N and O. The WHO 2009 list of mutations for surveillance of transmitted drug-resistant HIV strains was used to analyse TDRMs. Results: Applying our 'DBS-NGS-genotypic resistance test', baseline HIV-1 drug resistance data were successfully obtained from 82.8% (298/360) of newly diagnosed individuals. At nucleotide frequencies >15%, TDRMs to NRTIs were observed in 3.0% (9/298), to NNRTIs in 4.0% (12/298) and to PIs in 1.3% (3/240). The NNRTI mutation K103N was most commonly detected (2.7%). Expanding the analysis to low-abundance TDRMs, i.e. 3%-15%, 12 additional individuals (4.0%) harbouring TDRMs were identified. Having unprotected sex with a known HIV-1-positive person was significantly associated with the transmission of DRMs (adjusted OR 9.6; 95% CI 1.79-51.3). Conclusions: The prevalence of transmitted HIV-1 drug resistance is currently low in the study sites in Cameroon. Evidence of some risky sexual behaviours depicts a public health problem with possible implications for the prevention of new HIV-1 infections.

Emergence of Human Immunodeficiency Virus-1 Drug Resistance During the 3-Month World Health Organization-Recommended Enhanced Adherence Counseling Period in the CART-1 Cohort Study.

BACKGROUND: In resource-limited settings, the World Health Organization recommends enhanced adherence counseling (EAC) for individuals with an unsuppressed human immunodeficiency virus (HIV)-1 viral load (VL) and to remeasure VL after 3 months to avoid unnecessary regimen switches. In cases in which this follow-up VL remains unsuppressed, a regimen switch is indicated. We aimed to assess levels of HIV-1 drug resistance before and after the EAC period among people with ongoing viremia (≥80 c/mL) after EAC. METHODS: We included adult participants of the CART-1 cohort study conducted in Lesotho who had a VL ≥80 c/mL after EAC. Paired plasma samples (before and after EAC) were analyzed by next-generation sequencing. We assessed the prevalence of resistance-associated mutations and viral susceptibility scores to each participant's antiretroviral therapy (ART) regimen (range, 0-3; 3 indicates complete susceptibility). RESULTS: Among 93 participants taking nonnucleoside reverse-transcriptase inhibitor-based ART with an initial VL ≥1000 copies/mL who received a follow-up VL test after EAC, 76 still had a VL ≥80 copies/mL after EAC, and paired samples were available for 57 of 76. The number of individuals without full susceptibility to any drug in their regimen increased from 31 of 57 (54.4%) before to 36 of 57 (63.2%) after EAC. Median susceptibility scores dropped from 0.5 (interquartile range [IQR] = 0.25-) to 0.25 (IQR = 0.25-1) during the EAC period (P = .16). CONCLUSIONS: Despite high levels of resistance before EAC, we observed a slight decline in susceptibility scores after EAC. The risk of further accumulation of resistance during EAC has to be balanced against the benefit of avoiding unnecessary switches in those with spontaneous resuppression after EAC.

HCV Genetic Diversity Can Be Used to Infer Infection Recency and Time since Infection.

HIV-1 genetic diversity can be used to infer time since infection (TSI) and infection recency. We adapted this approach for HCV and identified genomic regions with informative diversity. We included 72 HCV/HIV-1 coinfected participants of the Swiss HIV Cohort Study, for whom reliable estimates of infection date and viral sequences were available. Average pairwise diversity (APD) was calculated over each codon position for the entire open reading frame of HCV. Utilizing cross validation, we evaluated the correlation of APD with TSI, and its ability to infer TSI via a linear model. We additionally studied the ability of diversity to classify infections as recent (infected for <1 year) or chronic, using receiver-operator-characteristic area under the curve (ROC-AUC) in 50 patients whose infection could be unambiguously classified as either recent or chronic. Measuring HCV diversity over third or all codon positions gave similar performances, and notable improvement over first or second codon positions. APD calculated over the entire genome enabled classification of infection recency (ROC-AUC = 0.76). Additionally, APD correlated with TSI (R2 = 0.33) and could predict TSI (mean absolute error = 1.67 years). Restricting the region over which APD was calculated to E2-NS2 further improved accuracy (ROC-AUC = 0.85, R2 = 0.54, mean absolute error = 1.38 years). Genetic diversity in HCV correlates with TSI and is a proxy for infection recency and TSI, even several years post-infection.