Female <i>Loa loa</i> worm polyinfection in human hosts

Loiasis is widespread in Central Africa. Some acute symptoms are associated with high <i>Loa loa</i> microfilaraemia, but the relation between the latter and the adult worm burden infecting an individual with loiasis is still unclear. This study aims to determine whether polyinfection by several reproductive female worms could be assessed using genetic variation in the mitochondrial genome of microfilariae. Microfilariae were collected from the individuals' blood. An optimization of the DNA extraction method that provides enough genetic material and minimization of human host contamination was the first step of the study. Extracted DNA was sequenced using the Illumina platform. Genetic variation in the mitochondrial genome was assessed by identifying polymorphic Single Nucleotide Polymorphisms (SNPs) and estimating the number of haplotypes. Dedicated DNA extraction kits yielded more DNA extracted (mean: 530 ng; SD = 211) from dried blood smears than the in-house chloroform-isoamyl method (mean: 102.5 ng; SD = 118). Filtering the slide elution and venous blood with 5 µm pore size microfilters improved parasite DNA mapping rates (54.64-79.65%). Analysis of polymorphism in the microfilariae mitochondrial genome from three individuals revealed 50, 207 and 332 polymorphic SNPs, respectively. A total of 7 to 20 mitochondrial DNA haplotypes were identified, representing the minimum number of fertile female worms. This study presents the first approach to estimating the <i>L. loa</i> female worm burden and highlights female parent polyinfection in individuals with loiasis.