Human Immunodeficiency Virus (HIV) remains a persistent global public health challenge. In 2020, approximately 37.7 million (30.2–45.1 million) individuals were living with HIV globally, including 1.7 million children <15 years old, with a global HIV prevalence of 0.8% among adults1. Almost 74.9 million individuals have been infected with HIV and 32.0 million have died from AIDS-related illnesses from the time when the epidemic started. A larger portion of people living with HIV(PLWH) are found in low- and middle-income countries, and Sub-Saharan Africa (SSA) is home to about 68% of people living with HIV in the world 2. Among this group, 20.6 million reside in East and Southern Africa which recorded 800,000 new HIV infections in 2018 (ref). The use of combination antiretroviral therapy (cART) in the treatment and management of HIV patients has lowered the risk of HIV-1 transmission, morbidity and mortality. Data from 2019 indicated that 25.33 million PLWH were receiving cART worldwide. However, mortality rates are still high in populations residing in resource-limited settings especially in the first months on cART as a result of late diagnosis and an already depleted. A meta-analysis of 63 studies conducted in low and middle income countries (LMICs) indicated that HIV drug resistance in persons before the start of cART (pre-treatment HIVDR or PDR) has steadily increased worldwide from 2000 to 2016, coincident with the wide-scale roll out of ART4.A systematic literature review on HIVDR resistance in 8 LMICs found that, in 60% of patients experiencing ART failure after 12 months of initiation, there were resistance mutations to at least 1 ART drug class5. Other studies reported that, in HIV-1–infected infants aged between 2.5 months - 8 years in SSA who were exposed to ART for prevention of mother-to-child transmission (PMCT), close to 32.4% had virus resistant to non- nucleoside reverse transcriptase inhibitors (NNRTIs)6. Around 86% of adolescents and young adults experiencing virologic failure had HIVDR in Zimbabwe 5678.HIVDR contributes to increased costs of HIV-1 care. Unfortunately, in Zimbabwe, HIVDR testing is not routinely done especially in children and adolescents and it might be one of the major causes of treatment failure among this special group.

Strikingly, with increased uptakes in PMTCT, challenges in cART programs, and high viremia among children and adolescents in SSA, the success rate of ART might be quickly compromised, with possible HIVDR emergence, particularly after years of paediatric cART exposure. Therefore, monitoring ART response in children and adolescents in terms of HIVDR patterns and socio-economic determinants of treatment failure might limit disease progression and guide on subsequent cART options for SSA especially Zimbabwean rural where there is paucity of information on HIVDR prevalence in children and adolescents.

We thus analysed baseline enrolment data collected through a randomized controlled study entitled Community Based ART (CBART) in order to determine the baseline characteristics and then genotyped plasma samples to determine the prevalence and distribution of HIVDR resistance among ART- experienced children and adolescents, aged 10 to 19 years, attending Chidamoyo Christian Hospital in Hurungwe, Zimbabwe

The study was nested in the CBART randomized control clinical trial .This was cohort study under Biomedical Research and Training Institute (BRTI), whose main objective as to determine if Implementation of point of care virus load differentiated care (POC virus load), targeted genotyping and Ministry of Health tools will result in improved virologic suppression among children, adolescents and young adults (<25 years) on ART.”The CBART study enrolled 600 cART experienced children and adolescents (10 to 19 years) over a period of two years (2018 to 2020).

In the original study, all eligible ART patients under the care of Chidamoyo Christian Hospital (< 25 years of age) were requested to consent to participate in the study at their regular cART visits. At the beginning of the study, the Ministry of Health and Child Care Zimbabwe (MOHCZ) was rolling out a new standard of care (SOC) which included annual virus load monitoring using the Roche Cobas HIV-1 RNA quantification (Roche Diagnostics, CA, USA) through the Provincial Laboratory at Chinhoyi (100 km from Chidamoyo). Four-hundred and fifty HIV infected children and adolescents and young adults (CAY) on cART were randomized (1:1) at enrolment to either SOC (n=300) or a near POC (n=300) VLDC monitoring test. All those who consented to CBART were offered the SOC VL performed by Roche COBAS at Chidamoyo and the results were returned to the hospital within 2-4 weeks of testing. Follow-up testing for HIV RNA > 1,000 copies/ml was offered using the same virologic monitoring system at the next drug/clinic visit within 3 months.

Participants recruitment, data collection and follow up were carried from January 2018–March 2020 in Hurungwe District, Mashonaland West Province, Zimbabwe. Hurungwe is comprised of mainly rural areas, with an economy centred on agriculture (mainly tobacco and cotton farming). Mashonaland West Provinces has the fourth highest HIV prevalence in Zimbabwe. A total of 3 223 new HIV infections were recorded in 2016 in Mashonaland West, out of which 4% were children below the age of 14. The study was conducted at Chidamoyo Christian Hospital in Hurungwe which serves about 30000 people and is an important referral centre for neighbouring areas such as Siakobvu, Kariba and Karoi. The hospital has over 18 outreach Opportunistic Infection (OI) clinics where they deliver ART drugs. However, 8 sites were used for this sub-study. The choice of the Institution for the study was based on its high HIV burden, its high coverage of cART, the long-term experience in ART provision of health facilities, hence need to assess HIVDR. In this sub-study,89 participants with viral load above 1000 copies per millilitre (VL>1000 copies/Ml) where enrolled and their samples genotyped for HIVDR.

Our study findings provide valuable data on HIVDR among adolescents and children with virologic failure in Hurungwe, Zimbabwe. The study participants had similar characteristics i.e. those who were successfully genotyped and those who were not.

We observed HIVDR in 32.5% of the participants who had VL≥1000 having at least one mutation. More than half (62%) of those with HIVDR were resistant to at least one of their current ART drugs. This indicates that approximately 1/3 of adolescents and children who were failing ART require a drug switch and over 2/3 are experiencing ART failure as a result of other factors other than HIVDR for example non-adherence and sub-optimal dosing.

There is also a possibility that some of the participants with poor adherence could have HIVDR but with reversion to wildtype. The phenomenon of reversion HIV to wildtype in the absence of optimal drug pressure may complicate HIVDR result interpretation for the patients who experienced virologic failure but with no HIVDR.

Medium-high levels of HIVDR mutations are not unique to Hurungwe or Zimbabwe. A number of studies done in adults from Zimbabwe have reported a range of HIVDR from 10-48%. In one study conducted in children and adolescents in Zimbabwe, found that 31% had HIVDR, 29% cases were resistant to NNRTIs,3% were resistant to NRTIs. Above that, our findings indicated that most participants who had resistance to a current regimen were 1^st line failures and required a drug switch to the 2^nd line regimen. These findings highlight the potential implications of delayed clinical decision making in this special group. Chidamoyo Opportunistic Infection (OI) patients have no enough access to recommended testing onsite and they rely on Chinhoyi Central Hospital laboratory which is overwhelmed most of the times with samples from a number of sites in the provinces and sometimes it takes a month or more to get the result back. Inaccessibility of routine viral load tests and delays in turnaround times for VL results contribute to delayed decision making which might cause the emergence of HIVDR. Therefore, there is urgent need to strengthen mechanisms to ensure availability of VL results, improving tracking mechanisms for results and clinical supervision of healthcare workers to support timely decision making.

We also found that the prevalence of HIVDR mutation to 1^st line NNRTI and NRTI was the same (65%). Only 5 patients (17.24 %) were resistant to PI. The high levels of HIVDR to NNRTI supports the 2018 Zimbabwean treatment guidelines on the use of high genetic barrier drugs such as DTG for all age groups and PIs (Lopinavir-ritonavir) for specific groups including children who are not eligible for DTG as 1^st treatment in place of NNRTIs. However, numerous studies which were conducted in Zimbabwe and SSA region have revealed an increasing rate of primary NNRTI among adult patients. Increase in primary NNRTI resistance together with the increase of adolescents and children experiencing NNRTI resistance in this current study indicates an urgent need for optimizing HIVDR testing as a way of determining the best drug option, more importantly in those who were infected perinatally.

Our results support the need for routine HIVDR testing surveillance to determine prevalence of primary and acquired HIVDR in both ART-naive and experienced children and adolescents to guide therapeutic options as they develop into adulthood.

When it comes to HIVDR to NRTI, the prevalence of any TAMs was at 32.8% and the most common TAM’s were common were M41L (24.1%), D67N (10.34%), T215Y (17.24%), and K70R (10.34%). TAM’s confer resistance to zidovudine, stavudine and other NRTI. When compared to earlier studies from Zimbabwe, results from this current study shows a higher prevalence of TAM’s among patients failing first line HIV treatment and with resistance.

Higher prevalence of TAMs (D67N and K70R) or TAMs in combination (T215Y with M41L) lessens susceptibility to Zidovudine (an important drug for most 2^nd line regimens). The TAMs of K70R/T/Q/N/E, K219Q/E, and D67N in combination effectively lowers the potency of zidovudine in a 2^nd line combination. The high prevalence of TAMs is a cause of concern although most of these participants (80.7%) have been on zidovudine or stavudine 1^stline ART at the time of enrolment into the study. Considering that these individuals were on abacavir or tenofovir, there is a chance that the TAMs observed emanated from TDR or to archived mutations from previous ART switches/exposures. Transmitted resistance and archived mutations have reported as potential causes of HIVDR. High NNRTI and NRTI resistance mutations may suggest the effect of previous policy driven first line ART switches over time that were done without any VL or HIVDR tests results. High prevalence of TAMs in our study brings to question the present-day practice especially in low income countries of switching patients failing first line regimens to second line regimens without HIVDR testing. With no HIVDR testing, we do not know what fraction of patients failing the first line ART where failing because of sub-optimal regimens or poor adherence. There is therefore an urgent need to provide HIV drug resistance testing at first line ART failure

Another major observation from our research was the high prevalence of the Y181C (10.3%) and Y188CE (3.44%) mutations, revealing resistance to 3^rd line therapy rilpivirine and etravirine, to which none of the participants had prior exposure to. Y181C and Y188C mutations has been demonstrated among 2^nd line failures earlier studies from SSA (5)(67). In our study, resistance to these drugs may be due in part to cross resistance with nevirapine and efavirenz which are widely used in PMTCT or first line ART. Regrettably, we were unable establish whether the HIVDR mutation to etravirine resulted from primary HIVDR transmission or developed as secondary HIVDR.

Findings from recent studies have showed the increasing outstanding role of integrase inhibitors over NNRTIs and PIs even in the presence of TAMs. These results may suggest that NRTIs are effective when used in together with integrase inhibitor dolutegravir (DTG)(68). Hence, the drive to introduce DTG as part of first and second line HIV care in Zimbabwe may be appropriate in the face of our evidence that third line ARV etravirine may potentially not be effective in approximately 3-5% of all youth failing on ART in Chidamoyo.

Gender differences in ARV treatment outcomes and drug-resistance mutations were also of interest in our study. Several studies from developed regions have not identified gender as a predictor for primary drug resistance. Differences between male and female participants were recorded in the levels of HIV-1 expressed mutations and HIVDR. In this study, we observed that the average number of HIV mutations was 4.9 in male and 4.5 in female participants respectively and this is in agreement with previous findings other studies, our sample size was low for us to conclude.

We also noticed that NNRTI-associated HIVDR was more common in individuals who had lower baseline CD4⁺ cell count.  Among the participants who had at least one HIVDR mutation, we noticed that any NNRTI resistance, the mutations G190A, K103N and K101E were associated with low CD4 cell (200–350cells/μl)(69). Though there is no scientific evidence from previous findings to support this observation, we suggest that this might be due to the fact that majority of our participants with HIVDR had NNRTI resistance.

This study was a cross-sectional study among ART experienced group and it was not possible to determine primary HIVDR and actual factors that would have contributed to the development of HIVDR. Our study sample size was low, but our findings makes valid contributions on prevalence, distribution and characteristics of HIVDR among children and adolescents.

HIVDR represents of the serious challenges in ART treatment in Zimbabwe and SSA. Majority of children and adolescents experiencing virologic failure may be resistant to their current ART regimens and require regimen switch. Use of HIVDR testing in guiding and monitoring development of HIVDR at the start of ART or at 1^st failure can be very important in treatment options and patient management.