Genetic Characterization of Biomphalaria pfeifferi from Asao Stream, Kisumu County Kenya: in Relation to Seasonal Variation, Selfing and Refractory Properties

Abstract

Biomphalaria pfeifferi, a predominantly self-fertilizing freshwater snail, is the world’s most important intermediate host for Schistosoma mansoni, one of the causative agents of schistosomiasis, which affects about 200 million people in sub-Saharan Africa. Understanding the genetic variation, selfing rates, and refractory traits of B. pfeifferi is important for improving snail-based strategies to interrupt S. mansoni transmission. The use of multilocus genotypes (MLGs) is important in identifying unique lineages within the snail population. This study sought to determine: (i) if lineages of B. pfeifferi within Asao stream could persist through different seasons of the year, (ii) if infections by S. mansoni or other trematodes occur within the persistent lineages, (iii) if B. pfeifferi from the Asao stream are selfers and finally (iv) if there are refractory genotypes among snails from Asao stream. B. pfeifferi snails were collected from Asao Stream in six bi monthly (every other month) sampling times between November 2018 and September 2019. Snails were isolated and screened for S. mansoni and other trematode infections using the shedding method, cercarial morphology was examined microscopically. Successive infection and breeding experiments was done to determine refractory genotypes. Using 14 microsatellite markers in a multiplex Polymerase Chain Reaction (PCR), B. pfeifferi snails were genotyped. Data analysis was done using GENALEX software and R studio software; poppr and coin packages. For the first three objectives, a total of 502 B. pfeifferi snails were collected. Genetic differentiation among the snails was low, ranging from FST = 0.019, p = <0.05 (November and March) to FST = 0.372, p = <0.001) (July and March). Analysis of molecular variance (AMOVA) showed a small variation (8%) among sampling times. A total of 319 MLGs were identified, 26 of which were present at two or more sampling times. Four MLGs persisted across the 10 month sampling period, one of which was represented by 17 individuals. Of the trematode species S. mansoni was the most common at 64.79%, other trematodes included; Echinostomes (3.29%), Xiphidiocercariae (6.10%), Paramphistomoids (23.47), Sangunicolids (0.94%) and Strigeids (1.41%). The persistent MLGs had more trematode infections compared to those found only at a single sampling time. There was high inbreeding coefficients (Mean FIS =0.639) and low heterozygosity (Mean Ho=0.145), with selfing rates estimated at 77.77%. Objective four began with 300 non shedding snails, the breeding experiment generated 10 Fillial 3(F3) B. pfeifferi non-shedding snails. These findings suggest that B. pfeifferi in Asao stream is predominantly self-fertilizing, with persistent genotypes that may sustain S. mansoni transmission across seasons. The identification of 10 F3 non-shedding snails suggests the possible presence of refractory snails. This study highlights the need for year-round snail control and monitoring of potentially refractory snail genotypes should be considered in control strategies.