Phylogenetic and drug-resistance analysis of HIV-1 sequences from an extensive paediatric HIV-1 outbreak in Larkana, Pakistan

Introduction: In April 2019, an HIV-1 outbreak among children occurred in Larkana, Pakistan, affecting more than a thousand children. It was assumed that the outbreak originated from a single source, namely a doctor at a private health facility. In this study, we performed subtype distribution, phylogenetic and drug-resistance analysis of HIV-1 sequences from 2019 outbreak in Larkana, Pakistan.Methods: A total of 401 blood samples were collected between April-June 2019, from children infected with HIV-1 aged 0-15 years recruited into a case-control study to investigate the risk factors for HIV-1 transmission. Partial HIV-1 pol sequences were generated from 344 blood plasma samples to determine HIV-1 subtype and drug resistance mutations (DRM). Maximum-likelihood phylogenetics based on outbreak and reference sequences was used to identify transmission clusters and assess the relationship between outbreak and key population sequences between and within the determined clusters. Bayesian analysis was employed to identify the time to the most recent common recent ancestor (tMRCA) of the main Pakistani clusters.Results: The HIV-1 circulating recombinant form (CRF) 02_AG and subtype A1 were most common among the outbreak sequences. Of the treatment-naïve participants, the two most common mutations were RT: E138A (8%) and RT: K219Q (8%). Four supported clusters within the outbreak were identified, and the median tMRCAs of the Larkana outbreak sequences were estimated to 2016 for both the CRF02_AG and the subtype A1 clusters. Furthermore, outbreak sequences exhibited no phylogenetic mixing with sequences from other high-risk groups of Pakistan.Conclusion: The presence of multiple clusters indicated a multi-source outbreak, rather than a single source outbreak from a single health practitioner as previously suggested. The multiple introductions were likely a consequence of ongoing transmission within the high-risk groups of Larkana, and it is possible that the so-called Larkana strain was introduced into the general population through poor infection prevention control practices in healthcare settings. The study highlights the need to scale up HIV-1 prevention programmes among key population groups and improving infection prevention control in Pakistan

Phylogenetic and Drug-Resistance Analysis of HIV-1 Sequences From an Extensive Paediatric HIV-1 Outbreak in Larkana, Pakistan.

INTRODUCTION: In April 2019, an HIV-1 outbreak among children occurred in Larkana, Pakistan, affecting more than a thousand children. It was assumed that the outbreak originated from a single source, namely a doctor at a private health facility. In this study, we performed subtype distribution, phylogenetic and drug-resistance analysis of HIV-1 sequences from 2019 outbreak in Larkana, Pakistan. METHODS: A total of 401 blood samples were collected between April-June 2019, from children infected with HIV-1 aged 0-15 years recruited into a case-control study to investigate the risk factors for HIV-1 transmission. Partial HIV-1 pol sequences were generated from 344 blood plasma samples to determine HIV-1 subtype and drug resistance mutations (DRM). Maximum-likelihood phylogenetics based on outbreak and reference sequences was used to identify transmission clusters and assess the relationship between outbreak and key population sequences between and within the determined clusters. Bayesian analysis was employed to identify the time to the most recent common recent ancestor (tMRCA) of the main Pakistani clusters. RESULTS: The HIV-1 circulating recombinant form (CRF) 02_AG and subtype A1 were most common among the outbreak sequences. Of the treatment-naïve participants, the two most common mutations were RT: E138A (8%) and RT: K219Q (8%). Four supported clusters within the outbreak were identified, and the median tMRCAs of the Larkana outbreak sequences were estimated to 2016 for both the CRF02_AG and the subtype A1 clusters. Furthermore, outbreak sequences exhibited no phylogenetic mixing with sequences from other high-risk groups of Pakistan. CONCLUSION: The presence of multiple clusters indicated a multi-source outbreak, rather than a single source outbreak from a single health practitioner as previously suggested. The multiple introductions were likely a consequence of ongoing transmission within the high-risk groups of Larkana, and it is possible that the so-called Larkana strain was introduced into the general population through poor infection prevention control practices in healthcare settings. The study highlights the need to scale up HIV-1 prevention programmes among key population groups and improving infection prevention control in Pakistan

Inferring transmission dynamics from HIV-1 genealogies

With a national prevalence of 4.9% in the adult population, the HIV-1 epidemic in Kenya is the fifth largest in the world. HIV-1 prevalence is more than three-fold higher among HIV key populations – including men who have sex with men (MSM), people who inject drugs (PWID), and female sex workers (FSW) than in the general heterosexual (HET) population. However, the contribution of different risk groups in the propagation of the epidemic has not been investigated. Also, theepidemic is geographically heterogeneous (65% of all new infections occur in nine out of the 47 counties in Kenya). Yet, the rates of HIV-1 transmission between geographic regions have not been described. Also, data are lacking on how levels and trends of HIV drug resistance (HIVDR) in Kenya compare among individuals of different risk groups, with or without antiretroviral therapy (ART) exposure. The primary objective was to phylogenetically describe virus transmission within and between risk groups (MSM, PWID, FSW, and HET) and geographic locations as well as to determine levels of HIV-1 drug resistance over time within and between risk groups in Kenya. A secondary objective was to phylogenetically characterise transmission patterns in a paediatric HIV-1 outbreak in Pakistan.In the first objective, clustering patterns in Kenya indicated that HIV-1 transmission between risk groups was rare – where most HIV-1 transmission occurs within-risk groups. In addition, when HIV-1 (infrequently) jumped between risk populations, virus jumps from HET to key populations were more common than vice-versa. There was significant West-to-East transmission (i.e. from high-to-low HIV-1 prevalence regions) in the mixed epidemic. Interestingly, Coast and Nairobi provinces were suggested to be important geographic hubs of HIV-1 dissemination in the MSM-specific HIV-1 sub-epidemic. HIVDR analysis revealed that overall pre-treatment HIVDR increased from 6.9% in 1986-2005 to 24.2% in 2016-2020. This was associated with increased non-nucleoside reverse transcriptase inhibitors (NNRTI) resistance in all risk groups. DRMs of any kind were found in treatment naïve HET (13.9%, 95% CI: 12.7-15.2), FSW (19.9%, 95% CI: 15.8-24.6), MSM (15.1%, 95% CI: 9.7-21.9), PWID (31.0%, 95% CI: 19.5-44.5), and children (41.3%, 95% CI: 30.1-53.3). PWID and children were more likely than HET to have DRMs (aOR, 3.5, 95% CI: 1.7-5.4, p<0.001, and aOR, 3.0, 95% CI: 1.8-4.8, p<0.001), respectively. No integrase strand transfer inhibitors (INSTI) drug resistance was detected. Hence, current INSTI-based ART regimens may remain effective in controlling HIV-1 in Kenya. In the secondary objective, clustering patterns in the Pakistani paediatric HIV-1 outbreak revealed multiple introductions of HIV-1 and no phylogenetic HIV-1 mixing between children andkey populations. Findings may be relevant for HIV-1 control in Kenya and Pakistan