key: cord-0838949-323zq9co authors: Snyman, Jumari; Hwa, Shi-Hsia; Krause, Robert; Muema, Daniel; Reddy, Tarylee; Ganga, Yashica; Karim, Farina; Leslie, Alasdair; Sigal, Alex; Ndung’u, Thumbi title: Similar antibody responses against SARS-CoV-2 in HIV uninfected and infected individuals on antiretroviral therapy during the first South African infection wave date: 2021-09-02 journal: Clin Infect Dis DOI: 10.1093/cid/ciab758 sha: dc79141dc7115283dd12e1fea9957b13f7c1f137 doc_id: 838949 cord_uid: 323zq9co BACKGROUND: There is limited understanding of SARS-CoV-2 pathogenesis in African populations with a high burden of infectious disease comorbidities such as HIV. The kinetics, magnitude and duration of virus-specific antibodies and the underlying B cell responses in people living with HIV (PLWH) in sub-Saharan Africa have not been fully characterized. METHODS: We longitudinally followed SARS-CoV-2 infected individuals in Durban, KwaZulu-Natal, South Africa and characterized SARS-CoV-2 receptor binding domain-specific IgM, IgG and IgA antibodies weekly for a month, and then at 3 months post diagnosis. 7/30 (41.7%) were PLWH, 83% (25/30) of which were on ART and with full HIV suppression. Potency of convalescent plasma neutralization was determined using a live virus neutralization assay and antibody secreting cell population frequencies were determined by flow cytometry. RESULTS: Similar seroconversion rates, time to peak antibody titer, peak magnitude and durability of anti-SARS-CoV-2 IgM, IgG, IgA, were observed in HIV uninfected and PLWH with complete HIV suppression on ART. In addition, similar neutralization potency against an isolate of SARS-CoV-2, circulating at the time of sampling in the first wave of SARS-CoV-2 infections in South Africa was observed in both groups. Loss of IgA was significantly associated with age (p=0.023) and a previous diagnosis of TB (p=0.018). CONCLUSIONS: Similar antibody response kinetics and neutralization potency in HIV negative and PLWH on stable ART in an African setting suggests that COVID-19 natural infections may confer comparable antibody immunity in these groups. This provides hope that COVID-19 vaccines will be effective in PLWH on stable ART. The COVID-19 pandemic caused by the SARS-CoV-2 virus has significantly impacted global health. Yet, there is relatively limited understanding of its impact in sub-Saharan African populations, with most studies conducted in developed countries. The heterogeneity in severity of COVID-19 has shown the importance of characterization of clinical outcomes and the corresponding immune responses across different populations. This is particularly important for sub-Saharan Africa (sSA) as this region has a higher burden of infectious diseases when compared with other regions. It bears the greatest burden of malaria, tuberculosis and HIV among other endemic infectious diseases that could significantly modulate the immunological profiles of individuals [1] . Indeed, poor immunological responsiveness to Ebola and Yellow Fever vaccines have been reported in African populations when compared to European populations, and this has been attributed to high baseline inflammatory profiles in African populations, even though genetic differences cannot be ruled out [2, 3] . Further, sSA bears the greatest burden of HIV; of the 37.9 million people living with HIV (PLWH) globally, 25.7 million reside in sSA [4] . Some countries in the region have particularly high HIV prevalence, such as South Africa especially KwaZulu-Natal, with a prevalence rate of 19% [5] . Notably, 48.0% of PLWH in sSA remain viremic, suggesting that a large proportion of PLWH in the region could be immunosuppressed. Moreover, antiretroviral therapy reduces but does not fully eliminate HIV-induced inflammation and immune activation, suggesting that some immune defects may persist despite fully suppressive antiretroviral therapy [6, 7] . Population cohort studies report an association of HIV infection and higher disease severity and/or mortality in COVID-19 patients [8] [9] [10] . The neutralizing antibody response to SARS-CoV-2 is a key correlate of protection [11] , HIVinduced impairment of anti-SARS-CoV-2 neutralizing antibody responses could result in A c c e p t e d M a n u s c r i p t 4 higher disease severity and mortality in COVID-19 patients. The antibody response to SARS-CoV2, in PLWH has, however, not been well characterized to date with most studies focusing on clinical outcome. HIV is known to affect multiple components of the immune system, including the B-cell compartment [12] , and PLWH make poor antibody responses to routine vaccination or upon exposure to other natural infections [13] [14] [15] [16] . Effective control of HIV viremia with antiretroviral drugs improves responsiveness to routine vaccines, especially when antiretroviral therapy is initiated early in the infection [13, 15, 17] . Considering that anti-SARS-CoV-2 antibodies effectively prevent SARS-CoV-2 infection in COVID-19 convalescent individuals and COVID-19 vaccine recipients, understanding the impact of HIV on their elicitation will reveal possible interactions between the two diseases in sSA. Antibodies are also important diagnostic and surveillance tools [18, 19] and understanding whether there are differences in responses between PLWH and the HIV uninfected population may have implications for the development of diagnostic and surveillance algorithms particularly in regions with high HIV burden. To characterize the general B-cell response to a live SARS-CoV-2 isolate (B.1.1.117, referred to here as D614G to denote the only mutation of significance) representing a typical SARS-CoV-2 infecting virus in sSA at the time of sampling, we report on the levels of anti-SARS-CoV-2 IgM, IgG, IgA, neutralizing antibodies, and antibody secreting cells (ASC) in a South African cohort of SARS-CoV-2 confirmed cases. We also disaggregate the study population based on HIV status to assess the impact of HIV on anti-SARS-CoV-2 antibody responses in PLWH on antiretroviral treatment (ART). A c c e p t e d M a n u s c r i p t 5 The study protocol was approved by the University of KwaZulu-Natal Institutional Review Board (approval BREC/00001275/2020). Written informed consent was obtained for all participants. All study participants were over 18 years old, capable of giving informed consent, presented with a positive SARS-CoV-2 polymerase chain reaction (PCR)-based diagnosis, and were A separate blood sample per participant was sent to an accredited diagnostic laboratory Table S1 ). Plasma samples collected were tested for anti-SARS-CoV-2 IgM, IgG and IgA antibodies as antibodies (Jackson ImmunoResearch, West Grove, PA, USA) were added to the respective plates (100 µl/well) and incubated at RT for 1 hour. Bound secondary antibodies were detected using 1-step Ultra TMB substrate (100 μl/well) (ThermoFisher Scientific). Plates were incubated at RT for 3 (IgG) and 5 (IgM and IgA) min respectively in the dark before addition of 1 N sulphuric acid stop solution (100 μl/well). Plates were washed with high salt TBS containing 0.05% Tween-20 after each incubation. Standard curves were used to calculate the concentration (ng/mL) of anti-RBD expressed as IgG (anti-SARS-CoV-2 monoclonal, CR3022), IgM (anti-SARS-CoV-2 S1 RBD IgM, hIgM2001) or IgA (anti-SARS-CoV-2 S1 RBD IgA, hIgA2001) (Genscript Piscataway, NJ, USA) [20, 21] . We used pre-pandemic plasma samples as negative controls to define seroconversion cut-offs calculated as three times the standard deviation plus the mean of the negative samples for each of the isotypes. A live virus neutralization focus-forming assay (LVNA) using an isolate of SARS-CoV-2 (D614G) [22] with the following modifications: input virus was 100 focus-forming units Statistical analyses were conducted in Stata (version 16) and GraphPad Prism (version 9.01). Lowess curves were generated to indicate kinetics of antibodies over time. Kaplan Meier curves (multivariable analyses) were used to determine time to seroconversion and Pearson's chi square test to determine factors associated with the loss of antibodies, with 95% Confidence intervals (CI). Cox regression and Breslow method for ties, were used to determine any associations to antibody responses. Spearman's rank test was used to determine all correlations. P<0.05 were considered significant. A total of 72 SARS-CoV-2 infected patients with 294 clinical specimens were analyzed in this longitudinal study. Full genome SARS-CoV-2 sequence data was available for 16 participants only none of which was infected with a variant of concern (data not shown). while CD8 cell counts were significantly higher (p=0.0008) PLWH. Hypertension was the leading co-morbidity (19.4%) in participants followed by diabetes (18.1%) and having a history of TB were significantly more common in PLWH (p=0.008). Of the participants, 6/72 (12.5%) had no clinical manifestation, 45/72 (62.5%) had mild clinical symptoms with limitation of activities, and 18/72 (25.0%) had severe clinical symptoms and required supplemental oxygen (Table 1) . For all analyses, days from symptom onset were used or day of enrolment for asymptomatic cases. Figure S1 ). There was also no difference in time to seroconversion between PLWH and HIV negative participants for any isotype. HIV-associated parameters (HIV plasma viral load, CD4, CD8 cell counts and CD4:CD8 ratio) were also not associated with significant antibody losses in PLWH across all three isotypes. A history of TB (only 6 participants with a history of TB developed IgA) (p=0.0018) and age (34-45 years) (p=0.023) were associated with loss of IgA antibodies at 3-month post symptom onset (Figure 2a Overall, age was significantly associated with both production and loss of IgA. This trend, however, was not observed when stratified by HIV status, probably a limitation of the sample size. Interestingly, loss of IgA was also associated with a previous diagnosis of TB. Current and previous diagnoses of TB have also been associated with COVID-19 death in a A c c e p t e d M a n u s c r i p t 12 population cohort study in South Africa [9] . In other studies, active TB disease has been associated with a lower frequency of B cells and a higher frequency of atypical doublenegative B cells; and whereas the frequency of total B cells normalized after treatment, individuals with a history of TB treatment still had higher double-negative B cells [27] . Therefore, active TB might affect the humoral response to other pathogens including SARS-CoV-2. Since Mycobacterium tuberculosis and SARS-CoV-2 are both respiratory pathogens, existing TB-related damage to the mucous and respiratory membranes could also be further exacerbated by SARS-CoV-2 infection and affect mucosal immunity. These potential interactions between respiratory pathogens need further investigation. Antibody secreting cells at 0-13 days post symptom onset positively correlated with IgG and IgA production 35-68 days post symptom onset. Due to the small sample size, we could not stratify into PLWH and HIV negative groups. A recent study, however, reported no difference in ASC between these two groups [28] . Multiple studies have shown that most SARS-CoV-2 infected individuals produce Sand RBD-specific antibodies during the primary response, and RBD-specific monoclonal antibodies can neutralize the virus in vitro and in vivo [29] . A recent study had demonstrated that higher sensitivity of the S protein versus the nucleocapsid protein for both acute and post-infection phase with the anti-N IgG antibodies waning after acute infection [30] . Therefore, RBD-specific antibodies would likely contribute to protection against re-infection. These results are similar to previous reports. Whether the neutralization response is also similar in subsequent infection waves with variants that show multiple functional differences to the original circulating strains is yet to be determined. A c c e p t e d M a n u s c r i p t A c c e p t e d M a n u s c r i p t 27 Figure 5 GBD. Global Burden of Diseases. 2021(Accessed on 19th Dynamics of the Humoral Immune Response to a Prime-Boost Ebola Vaccine: Quantification and Sources of Variation Immune activation alters cellular and humoral responses to yellow fever 17D vaccine WHO. HIV/AIDS: Disease burden. 2021(Accessed on 19th UNAIDS. 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The authors declare no competing interests. A c c e p t e d M a n u s c r i p t A c c e p t e d M a n u s c r i p t