Study Design and Population

We conducted a cross-sectional study primarily dedicated to establishing the seroprevalence of arboviral and hemorrhagic viral infections in Mali during October–November 2016 in 7 sites. In Bamako only, we also used 637 serum samples collected in 2013 from eligible volunteer blood donors for studying the seroprevalence of arboviral infections.

The Institutional Review Board of the Faculty of Medicine and Odonto-Stomatology, University of Sciences, Techniques and Technologies, Bamako, Mali, reviewed and approved this study (IRB letter no. 2016/113/CE/FMPOS). First, we visited all sites to meet health professionals, administrative authorities, and local community to explain the study context; after obtaining community permission, the study staff visited participant families. We conducted the study in alignment with institutional procedures and guidelines.

We selected 7 districts representing the different ecoclimatic areas of Mali: Diéma, Kita, Bougouni, Kadiolo, Niono, Bandiagara, and Commune IV of Bamako (Figure 1). Those districts are spread over the different administrative regions of Mali, excluding the northern region, which could not be investigated for security reasons. The selected districts are also those used by the Ministry of Health for infectious diseases surveillance.

The main climatic zones of Mali by Köppen climate classification and sites of study of Zika virus seroprevalence.

We selected villages or city areas randomly from an exhaustive list in each district. We randomly selected families and recruited volunteers with the objective of including 100–150 participants per site (Table 1), which corresponds to the estimate of the recruitment capacity by field teams for each site investigated. This sample size allows establishing seroprevalence in each site and globally. For significance level α = 0.05 and seroprevalence values 5%–30%, the precision varies from ≈4% to 9% (sample size = 100), and from ≈1.5% to 3% (sample size = 800).

*α = 0.05.

We enrolled only healthy, nonfebrile male and female volunteers >15 years of age. We provided study information in the most familiar language of the volunteer and in the presence of an observer designated by the village authorities who ensured that volunteers understood the information completely and helped obtain answers to any question. All volunteers signed the informed consent form before enrollment and received a copy of the signed form.

We used the Open Data Kit (ODK) platform on tablets to collect data, including sociodemographic information, weight, and history of travels. The same day, we drew venous blood (5 mL) in SST vacutainer tubes (Becton Dickinson, https:/www.bd.com), kept them at 2°C –8°C, and centrifuged (1,500–2,000 × g) within 12 h before aliquoting and storing at −80°C.

Eligibility criteria for blood donors in Bamako included acceptance and signature of the consent form, age (18–55 y), a normal blood pressure, a bodyweight >50 kg, the absence of recent fever or history of fever, and negative testing for HIV, hepatitis B and C, and syphilis. We excluded persons with known chronic disease or sensitivity to blood collection or those who were pregnant or breastfeeding. We heat inactivated samples collected in Mali, packed them at the Malaria Research and Training Center in Bamako, and sent them to the National Reference Centre for Arboviruses (Unité des Virus Émergents, Marseille, France) for serologic investigations, in compliance with national regulations in both countries.