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Background: KwaZulu-Natal ranked second highest among South African provinces for the number of laboratory-confirmed cases during the second wave of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic. The seroprevalence of SARS-CoV-2 among certain vulnerable groups, such as people living with HIV in KwaZulu-Natal, is unknown. Objective: The study aimed to determine the prevalence of SARS-CoV-2 immunoglobulin G (IgG) in HIV-positive versus HIV-negative patients. Methods: This was a retrospective analysis of residual clinical blood specimens unrelated to coronavirus disease 2019 (COVID-19) submitted for diagnostic testing at Inkosi Albert Luthuli Central Hospital, Durban, from 10 November 2020 to 09 February 2021. Specimens were tested for SARS-CoV-2 immunoglobulin G on the Abbott Architect analyser. Results: A total of 1977/8829 (22.4%) specimens were positive for SARS-CoV-2 antibodies. Seroprevalence varied between health districts from 16.4% to 37.3%, and was 19% in HIV-positive and 35.3% in HIV-negative specimens. Seroprevalence was higher among female patients (23.6% vs 19.8%; p 0.0001) and increased with increasing age, with a statistically significant difference between the farthest age groups ( 10 years and 79 years; p 0.0001). The seroprevalence increased from 17% on 10 November 2020 to 43% on 09 February 2021 during the second wave. Conclusion: Our results highlight that during the second COVID-19 wave in KwaZulu-Natal a large proportion of people living with HIV were still immunologically susceptible. The reduced seropositivity in people with virological failure further emphasises the importance of targeted vaccination and vaccine response monitoring in these individuals. What the study adds: This study contributes to data on SARS-CoV-2 seroprevalence before and during the second wave in KwaZulu-Natal, South Africa, which has the highest HIV prevalence globally. Reduced seropositivity was found among people living with HIV with virological failure, highlighting the importance of targeted booster vaccination and vaccine response monitoring.