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To describe baseline HIV-1 RNA viral load (VL) trends within South Africa's Early Infant Diagnosis program 2010–2016, with reference to prevention of mother-to-child transmission guidelines.HIV-1 total nucleic acid polymerase chain reaction (TNA PCR) and RNA VL data from 2010 to 2016 were extracted from the South African National Health Laboratory Service's central data repository. Infants with a positive TNA PCR and subsequent baseline RNA VL taken at age <7 months were included. Descriptive statistics were performed for quantified and lower-than-quantification limit (LQL) results per annum and age in months. Trend analyses were performed using log likelihood ratio tests. Multivariable linear regression was used to model the relationship between RNA VL and predictor variables, whereas logistic regression was used to identify predictors associated with LQL RNA VL results.Among 13,606 infants with a positive HIV-1 TNA PCR linked to a baseline RNA VL, median age of first PCR was 57 days and VL was 98 days. Thirteen thousand one hundred ninety-five (97.0%) infants had a quantified VL and 411 (3.0%) had an LQL result. A significant decline in median VL was observed between 2010 and 2016, from 6.3 log10 (interquartile range: 5.6–6.8) to 5.6 log10 (interquartile range: 4.2–6.5) RNA copies per milliliter, after controlling for age (P < 0.001), with younger age associated with lower VL (P < 0.001). The proportion of infants with a baseline VL <4 Log10 RNA copies per milliliter increased from 5.4% to 21.8%. Subsequent to prevention of mother-to-child transmission Option B implementation in 2013, the proportion of infants with an LQL baseline VL increased from 1.5% to 6.1% (P < 0.001).Between 2010 and 2016, a significant decline in baseline viremia within South Africa's Early Infant Diagnosis program was observed, with loss of detectability among some HIV-infected infants.