Breadth of Fc-mediated effector function correlates with clinical immunity following human malaria challenge.

Malaria is a life-threatening disease of global health importance, particularly in sub-Saharan Africa. The growth inhibition assay (GIA) is routinely used to evaluate, prioritize, and quantify the efficacy of malaria blood-stage vaccine candidates but does not reliably predict either naturally acquired or vaccine-induced protection. Controlled human malaria challenge studies in semi-immune volunteers provide an unparalleled opportunity to robustly identify mechanistic correlates of protection. We leveraged this platform to undertake a head-to-head comparison of seven functional antibody assays that are relevant to immunity against the erythrocytic merozoite stage of Plasmodium falciparum. Fc-mediated effector functions were strongly associated with protection from clinical symptoms of malaria and exponential parasite multiplication, while the gold standard GIA was not. The breadth of Fc-mediated effector function discriminated clinical immunity following the challenge. These findings present a shift in the understanding of the mechanisms that underpin immunity to malaria and have important implications for vaccine development.

A retrospective analysis of P. falciparum drug resistance markers detects an early (2016/17) high prevalence of the k13 C469Y mutation in asymptomatic infections in Northern Uganda.

The emergence of drug-resistant Plasmodium falciparum parasites in sub-Saharan Africa will substantially challenge malaria control. Here, we evaluated the frequency of common drug resistance markers among adolescents from Northern Uganda with asymptomatic infections. We used an established amplicon deep sequencing strategy to screen dried blood spot samples collected from 2016 to 2017 during a reported malaria epidemic within the districts of Kitgum and Pader in Northern Uganda. We screened single-nucleotide polymorphisms within: kelch13 (Pfk13), dihydropteroate synthase (Pfdhps), multidrug resistance-1 (Pfmdr1), dihydrofolate reductase (Pfdhfr), and apical membrane antigen (Pfama1) genes. Within the study population, the median age was 15 years (14.3-15.0, 95% CI), and 54.9% (78/142) were Plasmodium positive by 18S rRNA qPCR, which were subsequently targeted for sequencing analysis. We observed a high frequency of resistance markers particularly for sulfadoxine-pyrimethamine (SP), with no wild-type-only parasites observed for Pfdhfr (N51I, C59R, and S108N) and Pfdhps (A437G and K540E) mutations. Within Pfmdr1, mixed infections were common for NF/NY (98.5%). While for artemisinin resistance, in kelch13, there was a high frequency of C469Y (34%). Using the pattern for Pfama1, we found a high level of polygenomic infections with all individuals presenting with complexity of infection greater than 2 with a median of 6.9. The high frequency of the quintuple SP drug-resistant parasites and the C469Y artemisinin resistance-associated mutation in asymptomatic individuals suggests an earlier high prevalence than previously reported from symptomatic malaria surveillance studies (in 2016/2017). Our data demonstrate the urgency for routine genomic surveillance programs throughout Africa and the value of deep sequencing.

Seroprevalence of Antibodies to Severe Acute Respiratory Syndrome Coronavirus 2 Among Healthcare Workers in Kenya.

BACKGROUND: Few studies have assessed the seroprevalence of antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) among healthcare workers (HCWs) in Africa. We report findings from a survey among HCWs in 3 counties in Kenya. METHODS: We recruited 684 HCWs from Kilifi (rural), Busia (rural), and Nairobi (urban) counties. The serosurvey was conducted between 30 July and 4 December 2020. We tested for immunoglobulin G antibodies to SARS-CoV-2 spike protein, using enzyme-linked immunosorbent assay. Assay sensitivity and specificity were 92.7 (95% CI, 87.9-96.1) and 99.0% (95% CI, 98.1-99.5), respectively. We adjusted prevalence estimates, using bayesian modeling to account for assay performance. RESULTS: The crude overall seroprevalence was 19.7% (135 of 684). After adjustment for assay performance, seroprevalence was 20.8% (95% credible interval, 17.5%-24.4%). Seroprevalence varied significantly (P < .001) by site: 43.8% (95% credible interval, 35.8%-52.2%) in Nairobi, 12.6% (8.8%-17.1%) in Busia and 11.5% (7.2%-17.6%) in Kilifi. In a multivariable model controlling for age, sex, and site, professional cadre was not associated with differences in seroprevalence. CONCLUSION: These initial data demonstrate a high seroprevalence of antibodies to SARS-CoV-2 among HCWs in Kenya. There was significant variation in seroprevalence by region, but not by cadre.

Malaria attributable fractions with changing transmission intensity: Bayesian latent class vs logistic models.

BACKGROUND: Asymptomatic carriage of malaria parasites is common in high transmission intensity areas and confounds clinical case definitions for research studies. This is important for investigations that aim to identify immune correlates of protection from clinical malaria. The proportion of fevers attributable to malaria parasites is widely used to define different thresholds of parasite density associated with febrile episodes. The varying intensity of malaria transmission was investigated to check whether it had a significant impact on the parasite density thresholds. The same dataset was used to explore an alternative statistical approach, using the probability of developing fevers as a choice over threshold cut-offs. The former has been reported to increase predictive power. METHODS: Data from children monitored longitudinally between 2005 and 2017 from Junju and Chonyi in Kilifi, Kenya were used. Performance comparison of Bayesian-latent class and logistic power models in estimating malaria attributable fractions and probabilities of having fever given a parasite density with changing malaria transmission intensity was done using Junju cohort. Zero-inflated beta regressions were used to assess the impact of using probabilities to evaluate anti-merozoite antibodies as correlates of protection, compared with multilevel binary regression using data from Chonyi and Junju. RESULTS: Malaria transmission intensity declined from over 49% to 5% between 2006 and 2017, respectively. During this period, malaria attributable fraction varied between 27-59% using logistic regression compared to 10-36% with the Bayesian latent class approach. Both models estimated similar patterns of fevers attributable to malaria with changing transmission intensities. The Bayesian latent class model performed well in estimating the probabilities of having fever, while the latter was efficient in determining the parasite density threshold. However, compared to the logistic power model, the Bayesian algorithm yielded lower estimates for both attributable fractions and probabilities of fever. In modelling the association of merozoite antibodies and clinical malaria, both approaches resulted in comparable estimates, but the utilization of probabilities had a better statistical fit. CONCLUSIONS: Malaria attributable fractions, varied with an overall decline in the malaria transmission intensity in this setting but did not significantly impact the outcomes of analyses aimed at identifying immune correlates of protection. These data confirm the statistical advantage of using probabilities over binary data.

Controlled human malaria infection (CHMI) outcomes in Kenyan adults is associated with prior history of malaria exposure and anti-schizont antibody response.

BACKGROUND: Individuals living in endemic areas acquire immunity to malaria following repeated parasite exposure. We sought to assess the controlled human malaria infection (CHMI) model as a means of studying naturally acquired immunity in Kenyan adults with varying malaria exposure. METHODS: We analysed data from 142 Kenyan adults from three locations representing distinct areas of malaria endemicity (Ahero, Kilifi North and Kilifi South) enrolled in a CHMI study with Plasmodium falciparum sporozoites NF54 strain (Sanaria® PfSPZ Challenge). To identify the in vivo outcomes that most closely reflected naturally acquired immunity, parameters based on qPCR measurements were compared with anti-schizont antibody levels and residence as proxy markers of naturally acquired immunity. RESULTS: Time to endpoint correlated more closely with anti-schizont antibodies and location of residence than other parasite parameters such as growth rate or mean parasite density. Compared to observational field-based studies in children where 0.8% of the variability in malaria outcome was observed to be explained by anti-schizont antibodies, in the CHMI model the dichotomized anti-schizont antibodies explained 17% of the variability. CONCLUSIONS: The CHMI model is highly effective in studying markers of naturally acquired immunity to malaria. Trial registration Clinicaltrials.gov number NCT02739763. Registered 15 April 2016.

Antigenic cartography of immune responses to Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1).

Naturally acquired clinical immunity to Plasmodium falciparum is partly mediated by antibodies directed at parasite-derived antigens expressed on the surface of red blood cells which mediate disease and are extremely diverse. Unlike children, adults recognize a broad range of variant surface antigens (VSAs) and are protected from severe disease. Though crucial to the design and feasibility of an effective malaria vaccine, it is not yet known whether immunity arises through cumulative exposure to each of many antigenic types, cross-reactivity between antigenic types, or some other mechanism. In this study, we measured plasma antibody responses of 36 children with symptomatic malaria to a diverse panel of 36 recombinant proteins comprising part of the DBLα domain (the 'DBLα-tag') of PfEMP1, a major class of VSAs. We found that although plasma antibody responses were highly specific to individual antigens, serological profiles of responses across antigens fell into one of just two distinct types. One type was found almost exclusively in children that succumbed to severe disease (19 out of 20) while the other occurred in all children with mild disease (16 out of 16). Moreover, children with severe malaria had serological profiles that were narrower in antigen specificity and shorter-lived than those in children with mild malaria. Borrowing a novel technique used in influenza-antigenic cartography-we mapped these dichotomous serological profiles to amino acid sequence variation within a small sub-region of the PfEMP1 DBLα domain. By applying our methodology on a larger scale, it should be possible to identify epitopes responsible for eliciting the protective version of serological profiles to PfEMP1 thereby accelerating development of a broadly effective anti-disease malaria vaccine.

Vaccine candidate discovery for the next generation of malaria vaccines.

Although epidemiological observations, IgG passive transfer studies and experimental infections in humans all support the feasibility of developing highly effective malaria vaccines, the precise antigens that induce protective immunity remain uncertain. Here, we review the methodologies applied to vaccine candidate discovery for Plasmodium falciparum malaria from the pre- to post-genomic era. Probing of genomic and cDNA libraries with antibodies of defined specificities or functional activity predominated the former, whereas reverse vaccinology encompassing high throughput in silico analyses of genomic, transcriptomic or proteomic parasite data sets is the mainstay of the latter. Antibody-guided vaccine design spanned both eras but currently benefits from technological advances facilitating high-throughput screening and downstream applications. We make the case that although we have exponentially increased our ability to identify numerous potential vaccine candidates in a relatively short space of time, a significant bottleneck remains in their validation and prioritization for evaluation in clinical trials. Longitudinal cohort studies provide supportive evidence but results are often conflicting between studies. Demonstration of antigen-specific antibody function is valuable but the relative importance of one mechanism over another with regards to protection remains undetermined. Animal models offer useful insights but may not accurately reflect human disease. Challenge studies in humans are preferable but prohibitively expensive. In the absence of reliable correlates of protection, suitable animal models or a better understanding of the mechanisms underlying protective immunity in humans, vaccine candidate discovery per se may not be sufficient to provide the paradigm shift necessary to develop the next generation of highly effective subunit malaria vaccines.

CD4+ T cell responses to the Plasmodium falciparum erythrocyte membrane protein 1 in children with mild malaria.

The immune response against the variant surface Ag Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) is a key component of clinical immunity against malaria. We have investigated the development and maintenance of CD4(+) T cell responses to a small semiconserved area of the Duffy binding-like domain (DBL)α-domain of PfEMP1, the DBLα-tag. Young children were followed up longitudinally, and parasites and PBMCs were isolated from 35 patients presenting with an acute case of uncomplicated malaria. The DBLα-tag from the PfEMP1 dominantly expressed by the homologous parasite isolate was cloned and expressed as recombinant protein. The recombinant DBLα-tag was used to activate PBMCs collected from each acute episode and from an annual cross-sectional survey performed after the acute malaria episode. In this article, we report that CD4(+) T cell responses to the homologous DBLα-tag were induced in 75% of the children at the time of the acute episode and in 62% of the children at the following cross-sectional survey on average 235 d later. Furthermore, children who had induced DBLα-tag-specific CD4(+)IL-4(+) T cells at the acute episode remained episode free for longer than children who induced other types of CD4(+) T cell responses. These results suggest that a wide range of DBLα-tag-specific CD4(+) T cell responses were induced in children with mild malaria and, in the case of CD4(+)IL-4(+) T cell responses, were associated with protection from clinical episodes.

Bi-isotype immunoglobulins enhance antibody-mediated neutrophil activity against Plasmodium falciparum parasites.

Background: Malaria remains a major global health priority, and monoclonal antibodies (mAbs) are emerging as potential new tools to support efforts to control the disease. Recent data suggest that Fc-dependent mechanisms of immunity are important mediators of protection against the blood stages of the infection, but few studies have investigated this in the context of mAbs. We aimed to isolate mAbs agnostic to cognate antigens that target whole merozoites and simultaneously induce potent neutrophil activity measured by the level of reactive oxygen species (ROS) production using an antibody-dependent respiratory burst (ADRB) assay. Methods: We used samples from semi-immune adults living in coastal Kenya to isolate mAbs that induce merozoite-specific ADRB activity. We then tested whether modifying the expressed IgG1 isotype to an IgG-IgA Fc region chimera would enhance the level of ADRB activity. Results: We isolated a panel of nine mAbs with specificity to whole merozoites. mAb J31 induced ADRB activity in a dose-dependent fashion. Compared to IgG1, our modified antibody IgG-IgA bi-isotype induced higher ADRB activity across all concentrations tested. Further, we observed a negative hook effect at high IgG1 mAb concentrations (i.e., >200 µg/mL), but this was reversed by Fc modification. We identified MSP3.5 as the potential cognate target of mAb J31. Conclusions: We demonstrate an approach to engineer mAbs with enhanced ADRB potency against blood-stage parasites.

Full-length MSP1 is a major target of protective immunity after controlled human malaria infection.

The merozoite surface protein 1 (MSP1) is the most abundant protein on the surface of the invasive merozoite stages of Plasmodium falciparum and has long been considered a key target of protective immunity. We used samples from a single controlled human malaria challenge study to test whether the full-length version of MSP1 (MSP1FL) induced antibodies that mediated Fc-IgG functional activity in five independent assays. We found that anti-MSP1FL antibodies induced complement fixation via C1q, monocyte-mediated phagocytosis, neutrophil respiratory burst, and natural killer cell degranulation as well as IFNγ production. Activity in each of these assays was strongly associated with protection. The breadth of MSP1-specific Fc-mediated effector functions was more strongly associated with protection than the individual measures and closely mirrored what we have previously reported using the same assays against merozoites. Our findings suggest that MSP1FL is an important target of functional antibodies that contribute to a protective immune response against malaria.

Antibody-Dependent Respiratory Burst against Plasmodium falciparum Merozoites in Individuals Living in an Area with Declining Malaria Transmission.

Malaria transmission intensity affects the development of naturally acquired immunity to malaria. An absolute correlate measure of protection against malaria is lacking. However, antibody-mediated functions against Plasmodium falciparum correlate with protection against malaria. In children, antibody-mediated functions against P. falciparum decline with reduced exposure. It is unclear whether adults maintain antibody-mediated functions as malaria transmission declines. This study assessed antibody-dependent respiratory burst (ADRB) in individuals from an area with declining malaria transmission. In an age-matched analysis, we compare ADRB activity during high versus low malaria transmission periods. Age significantly predicted higher ADRB activity in the high (p < 0.001) and low (p < 0.001) malaria transmission periods. ADRB activity was higher during the high compared to the low malaria transmission period in older children and adults. Only older adults during the high malaria transmission period had their median ADRB activity above the ADRB cut-off. Ongoing P. falciparum infection influenced ADRB activity during the low (p = 0.01) but not the high (p = 0.29) malaria transmission period. These findings propose that naturally acquired immunity to P. falciparum is affected in children and adults as malaria transmission declines, implying that vaccines will be necessary to induce and maintain protection against malaria.

Characterization of a novel Plasmodium falciparum merozoite surface antigen and potential vaccine target.

Introduction: Detailed analyses of genetic diversity, antigenic variability, protein localization and immunological responses are vital for the prioritization of novel malaria vaccine candidates. Comprehensive approaches to determine the most appropriate antigen variants needed to provide broad protection are challenging and consequently rarely undertaken. Methods: Here, we characterized PF3D7_1136200, which we named Asparagine-Rich Merozoite Antigen (ARMA) based on the analysis of its sequence, localization and immunogenicity. We analyzed IgG and IgM responses against the common variants of ARMA in independent prospective cohort studies in Burkina Faso (N = 228), Kenya (N = 252) and Mali (N = 195) using a custom microarray, Div-KILCHIP. Results: We found a marked population structure between parasites from Africa and Asia. African isolates shared 34 common haplotypes, including a dominant pair although the overall selection pressure was directional (Tajima's D = -2.57; Fu and Li's F = -9.69; P < 0.02). ARMA was localized to the merozoite surface, IgG antibodies induced Fc-mediated degranulation of natural killer cells and strongly inhibited parasite growth in vitro. We found profound serological diversity, but IgG and IgM responses were highly correlated and a hierarchical clustering analysis identified only three major serogroups. Protective IgG and IgM antibodies appeared to target both cross-reactive and distinct epitopes across variants. However, combinations of IgG and IgM antibodies against selected variants were associated with complete protection against clinical episodes of malaria. Discussion: Our systematic strategy exploits genomic data to deduce the handful of antigen variants with the strongest potential to induce broad protection and may be broadly applicable to other complex pathogens for which effective vaccines remain elusive.

Serum immunoglobulin G and mucosal immunoglobulin A antibodies from prepandemic samples collected in Kilifi, Kenya, neutralize SARS-CoV-2 in vitro.

OBJECTIVES: Many regions of Africa have experienced lower COVID-19 morbidity and mortality than Europe. Pre-existing humoral responses to endemic human coronaviruses (HCoV) may cross-protect against SARS-CoV-2. We investigated the neutralizing capacity of SARS-CoV-2 spike reactive and nonreactive immunoglobulin (Ig)G and IgA antibodies in prepandemic samples. METHODS: To investigate the presence of pre-existing immunity, we performed enzyme-linked immunosorbent assay using spike antigens from reference SARS-CoV-2, HCoV HKU1, OC43, NL63, and 229E using prepandemic samples from Kilifi in coastal Kenya. In addition, we performed neutralization assays using pseudotyped reference SARS-CoV-2 to determine the functionality of the identified reactive antibodies. RESULTS: We demonstrate the presence of HCoV serum IgG and mucosal IgA antibodies, which cross-react with the SARS-CoV-2 spike. We show pseudotyped reference SARS-CoV-2 neutralization by prepandemic serum, with a mean infective dose 50 of 1: 251, which is 10-fold less than that of the pooled convalescent sera from patients with COVID-19 but still within predicted protection levels. The prepandemic naso-oropharyngeal fluid neutralized pseudo-SARS-CoV-2 at a mean infective dose 50 of 1: 5.9 in the neutralization assay. CONCLUSION: Our data provide evidence for pre-existing functional humoral responses to SARS-CoV-2 in Kilifi, coastal Kenya and adds to data showing pre-existing immunity for COVID-19 from other regions.

Anti-merozoite antibodies induce natural killer cell effector function and are associated with immunity against malaria.

Natural killer (NK) cells are potent immune effectors that can be activated via antibody-mediated Fc receptor engagement. Using multiparameter flow cytometry, we found that NK cells degranulate and release IFN-γ upon stimulation with antibody-opsonized Plasmodium falciparum merozoites. Antibody-dependent NK (Ab-NK) activity was largely strain transcending and enhanced invasion inhibition into erythrocytes. Ab-NK was associated with the successful control of parasitemia after experimental malaria challenge in African adults. In an independent cohort study in children, Ab-NK increased with age, was boosted by concurrent P. falciparum infections, and was associated with a lower risk of clinical episodes of malaria. Nine of the 14 vaccine candidates tested induced Ab-NK, including some less well-characterized antigens: P41, P113, MSP11, RHOPH3, and Pf_11363200. These data highlight an important role of Ab-NK activity in immunity against malaria and provide a potential mechanism for evaluating vaccine candidates.

Characterization of Anopheles gambiae D7 salivary proteins as markers of human-mosquito bite contact.

BACKGROUND: Malaria is transmitted when infected Anopheles mosquitoes take a blood meal. During this process, the mosquitoes inject a cocktail of bioactive proteins that elicit antibody responses in humans and could be used as biomarkers of exposure to mosquito bites. This study evaluated the utility of IgG responses to members of the Anopheles gambiae D7 protein family as serological markers of human-vector contact. METHODS: The D7L2, D7r1, D7r2, D7r3, D7r4 and SG6 salivary proteins from An. gambiae were expressed as recombinant antigens in Escherichia coli. Antibody responses to the salivary proteins were compared in Europeans with no prior exposure to malaria and lifelong residents of Junju in Kenya and Kitgum in Uganda where the intensity of malaria transmission is moderate and high, respectively. In addition, to evaluate the feasibility of using anti-D7 IgG responses as a tool to evaluate the impact of vector control interventions, we compared responses between individuals using insecticide-treated bednets to those who did not in Junju, Kenya where bednet data were available. RESULTS: We show that both the long and short forms of the D7 salivary gland antigens elicit a strong antibody response in humans. IgG responses against the D7 antigens reflected the transmission intensities of the three study areas, with the highest to lowest responses observed in Kitgum (northern Uganda), Junju (Kenya) and malaria-naïve Europeans, respectively. Specifically, the long form D7L2 induced an IgG antibody response that increased with age and that was lower in individuals who slept under a bednet, indicating its potential as a serological tool for estimating human-vector contact and monitoring the effectiveness of vector control interventions. CONCLUSIONS: This study reveals that D7L2 salivary antigen has great potential as a biomarker of exposure to mosquito bites and as a tool for assessing the efficacy of vector control strategies such as bednet use.

Distinct kinetics of antibodies to 111 Plasmodium falciparum proteins identifies markers of recent malaria exposure.

Strengthening malaria surveillance is a key intervention needed to reduce the global disease burden. Reliable serological markers of recent malaria exposure could improve current surveillance methods by allowing for accurate estimates of infection incidence from limited data. We studied the IgG antibody response to 111 Plasmodium falciparum proteins in 65 adult travellers followed longitudinally after a natural malaria infection in complete absence of re-exposure. We identified a combination of five serological markers that detect exposure within the previous three months with >80% sensitivity and specificity. Using mathematical modelling, we examined the antibody kinetics and determined that responses informative of recent exposure display several distinct characteristics: rapid initial boosting and decay, less inter-individual variation in response kinetics, and minimal persistence over time. Such serological exposure markers could be incorporated into routine malaria surveillance to guide efforts for malaria control and elimination.

Phagocytosis of Plasmodium falciparum ring-stage parasites predicts protection against malaria.

Ring-infected erythrocytes are the predominant asexual stage in the peripheral circulation but are rarely investigated in the context of acquired immunity against Plasmodium falciparum malaria. Here we compare antibody-dependent phagocytosis of ring-infected parasite cultures in samples from a controlled human malaria infection (CHMI) study (NCT02739763). Protected volunteers did not develop clinical symptoms, maintained parasitaemia below a predefined threshold of 500 parasites/µl and were not treated until the end of the study. Antibody-dependent phagocytosis of both ring-infected and uninfected erythrocytes from parasite cultures was strongly correlated with protection. A surface proteomic analysis revealed the presence of merozoite proteins including erythrocyte binding antigen-175 and -140 on ring-infected and uninfected erythrocytes, providing an additional antibody-mediated protective mechanism for their activity beyond invasion-inhibition. Competition phagocytosis assays support the hypothesis that merozoite antigens are the key mediators of this functional activity. Targeting ring-stage parasites may contribute to the control of parasitaemia and prevention of clinical malaria.

Comparative performance of WANTAI ELISA for total immunoglobulin to receptor binding protein and an ELISA for IgG to spike protein in detecting SARS-CoV-2 antibodies in Kenyan populations.

Many SARS-CoV-2 antibody detection assays have been developed but their differential performance is not well described. In this study we compared an in-house (KWTRP) ELISA which has been used extensively to estimate seroprevalence in the Kenyan population with WANTAI, an ELISA which has been approved for widespread use by the WHO. Using a wide variety of sample sets including pre-pandemic samples (negative gold standard), SARS-CoV-2 PCR positive samples (positive gold standard) and COVID-19 test samples from different periods (unknowns), we compared performance characteristics of the two assays. The overall concordance between WANTAI and KWTRP was 0.97 (95% CI, 0.95-0.98). For WANTAI and KWTRP, sensitivity was 0.95 (95% CI 0.90-0.98) and 0.93 (95% CI 0.87-0.96), respectively. Specificity for WANTAI was 0.98 (95% CI, 0.96-0.99) and 0.99 (95% CI 0.96-1.00) while KWTRP specificity was 0.99 (95% CI, 0.98-1.00) and 1.00 using pre-pandemic blood donors and pre-pandemic malaria cross-sectional survey samples respectively. Both assays show excellent characteristics to detect SARS-CoV-2 antibodies.

Epidemiology of COVID-19 infections on routine polymerase chain reaction (PCR) and serology testing in Coastal Kenya.

Background: There are limited studies in Africa describing the epidemiology, clinical characteristics and serostatus of individuals tested for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. We tested routine samples from the Coastal part of Kenya between 17 th March 2020 and 30 th June 2021. Methods: SARS-CoV-2 infections identified using reverse transcription polymerase chain reaction (RT-PCR) and clinical surveillance data at the point of sample collection were used to classify as either symptomatic or asymptomatic. IgG antibodies were measured in sera samples, using a well validated in-house enzyme-linked immunosorbent assay (ELISA). Results: Mombasa accounted for 56.2% of all the 99,694 naso-pharyngeal/oro-pharyngeal swabs tested, and males constituted the majority tested (73.4%). A total of 7737 (7.7%) individuals were SARS-CoV-2 positive by RT-PCR. The majority (i.e., 92.4%) of the RT-PCR positive individuals were asymptomatic. Testing was dominated by mass screening and travellers, and even at health facility level 91.6% of tests were from individuals without symptoms. Out of the 97,124 tests from asymptomatic individuals 7,149 (7%) were positive and of the 2,568 symptomatic individuals 588 (23%) were positive. In total, 2458 serum samples were submitted with paired naso-pharyngeal/oro-pharyngeal samples and 45% of the RT-PCR positive samples and 20% of the RT-PCR negative samples were paired with positive serum samples. Symptomatic individuals had significantly higher antibody levels than asymptomatic individuals and become RT-PCR negative on repeat testing earlier than asymptomatic individuals. Conclusions: In conclusion, the majority of SARS-CoV-2 infections identified by routine testing in Coastal Kenya were asymptomatic. This reflects the testing practice of health services in Kenya, but also implies that asymptomatic infection is very common in the population. Symptomatic infection may be less common, or it may be that individuals do not present for testing when they have symptoms.

SARS-CoV-2 seroprevalence in three Kenyan health and demographic surveillance sites, December 2020-May 2021.

BACKGROUND: Most of the studies that have informed the public health response to the COVID-19 pandemic in Kenya have relied on samples that are not representative of the general population. We conducted population-based serosurveys at three Health and Demographic Surveillance Systems (HDSSs) to determine the cumulative incidence of infection with SARS-CoV-2. METHODS: We selected random age-stratified population-based samples at HDSSs in Kisumu, Nairobi and Kilifi, in Kenya. Blood samples were collected from participants between 01 Dec 2020 and 27 May 2021. No participant had received a COVID-19 vaccine. We tested for IgG antibodies to SARS-CoV-2 spike protein using ELISA. Locally-validated assay sensitivity and specificity were 93% (95% CI 88-96%) and 99% (95% CI 98-99.5%), respectively. We adjusted prevalence estimates using classical methods and Bayesian modelling to account for the sampling scheme and assay performance. RESULTS: We recruited 2,559 individuals from the three HDSS sites, median age (IQR) 27 (10-78) years and 52% were female. Seroprevalence at all three sites rose steadily during the study period. In Kisumu, Nairobi and Kilifi, seroprevalences (95% CI) at the beginning of the study were 36.0% (28.2-44.4%), 32.4% (23.1-42.4%), and 14.5% (9.1-21%), and respectively; at the end they were 42.0% (34.7-50.0%), 50.2% (39.7-61.1%), and 24.7% (17.5-32.6%), respectively. Seroprevalence was substantially lower among children (<16 years) than among adults at all three sites (p≤0.001). CONCLUSION: By May 2021 in three broadly representative populations of unvaccinated individuals in Kenya, seroprevalence of anti-SARS-CoV-2 IgG was 25-50%. There was wide variation in cumulative incidence by location and age.