Population genetic structure of Schistosoma haematobium and Schistosoma haematobium × Schistosoma bovis hybrids among school-aged children in Côte d'Ivoire

Abstract

While population genetics of Schistosoma haematobium have been investigated in West Africa, only scant data are available from Côte d'Ivoire. The purpose of this study was to analyze both genetic variability and genetic structure among S. haematobium populations and to quantify the frequency of S. haematobium × S. bovis hybrids in school-aged children in different parts of Côte d'Ivoire. Urine samples were subjected to a filtration method and examined microscopically for Schistosoma eggs in four sites in the western and southern parts of Côte d'Ivoire. A total of 2692 miracidia were collected individually and stored on Whatman^® FTA cards. Of these, 2561 miracidia were successfully genotyped for species and hybrid identification using rapid diagnostic multiplex mitochondrial cox1 PCR and PCR Restriction Fragment Length Polymorphism (PCR-RFLP) analysis of the nuclear ITS2 region. From 2164 miracidia, 1966 (90.9%) were successfully genotyped using at least 10 nuclear microsatellite loci to investigate genetic diversity and population structure. Significant differences were found between sites in all genetic diversity indices and genotypic differentiation was observed between the site in the West and the three sites in the East. Analysis at the infrapopulation level revealed clustering of parasite genotypes within individual children, particularly in Duekoué (West) and Sikensi (East). Of the six possible cox1-ITS2 genetic profiles obtained from miracidia, S. bovis cox1 × S. haematobium ITS2 (42.0%) was the most commonly observed in the populations. We identified only 15 miracidia (0.7%) with an S. bovis cox1 × S. bovis ITS2 genotype. Our study provides new insights into the population genetics of S. haematobium and S. haematobium × S. bovis hybrids in humans in Côte d'Ivoire and we advocate for researching hybrid schistosomes in animals such as rodents and cattle in Côte d'Ivoire.

Keywords: Côte d’Ivoire; Microsatellites; Population genetics; Schistosoma haematobium; Schistosomiasis.

Figure 1

Sites in the southern and western parts of Côte d’Ivoire and number (percentage of total sample size) of Schistosoma-infected children, miracidia collected, and miracidia genotyped.

Figure 2

Principal component analysis (PCA) of microsatellite data by sites. Each miracidium is represented by one dot. The first two principal components (PCs) explain 41.1% and 33.7% of total inertia of the data set, respectively.

Figure 3

Principal component analysis (PCA) of the microsatellite data with each miracidium assigned its cox1-ITS2 genotype. Each miracidium is represented by a single dot. The first two principal components (PCs) explain 61.6% and 21.0% of total inertia of the data set, respectively.

Figure 4

Bar plot depicting the genetic population structure of 1966 Schistosoma miracidia from the four sampling sites in Côte d’Ivoire produced by Structure for K = 4. Each column represents one miracidium, with colors indicating the proportional contribution of each of the four identified genetic clusters.

Figure 5

Genetic clustering produced by Structure software for each population with K = number of patients. The patient codes are in abscises. *, ** and *** represent statistical difference from random repartition among the clusters at the 5%, 1% and 0.1% levels, respectively. The red line represents the null hypothesis of random repartition (1/K).

Supplementary Fig. S1:

S1a: Diagram of the 505 bp fragment and the Mbo1 cut sites for both species, S1b: Banding patterns of the restriction fragment length polymorphism (RFLP) analysis by MboI of the nuclear ITS2 region of Schistosoma. Schistosoma haematobium exhibits four bands at 281, 98, 82 and 44 bp. Schistosoma bovis exhibits three bands at 379, 82 and 44 bp. Hybrids between S. haematobium × S. bovis exhibit bands at 379, 281, 98, 82 and 44 bp.

Supplementary Fig. S2:

Bar plots depicting the percentage for each miracidium belonging to each cluster by child, using children hosting at least 15 miracidia in Adzopé (ADZ), Agboville (AGB), Duekoué (DUE), and Sikensi (SIK). This was done in Structure and each site K was set equal to the number of children.