Detection of β-Tubulin Single Nucleotide Polymorphisms (SNPs) Associated with Drug Resistance among School-Aged Children Infected with Soil-Transmitted Helminths in Bungoma, Kenya

Abstract

Soil-transmitted helminths (STHs) are parasitic nematodes associated with poor sanitation. These parasites are transmitted to humans by ingesting infective eggs or skin penetration by larvae found in feces-contaminated soil. Approximately 1.5 billion people across the globe are infected with STHs with another 4 billion people being at risk of infection. Although STH infections may occur in different demographic groups, the adverse effects of these infections are most significant among children, resulting in malnutrition, anemia, stunted growth and impaired cognitive development. STHs are part of the 20 neglected tropical diseases (NTDS) identified by the WHO as priority diseases for enhanced control and elimination. In Kenya, STH infections remain a significant public health concern, mainly among vulnerable populations, such as children. Despite efforts to control STH infections through mass drug administration (MDA) programs which have been running in Kenya since 2012, the prevalence still persists, posing a risk to the health and well-being of millions of individuals across the country This highlights the need for continuous monitoring of the prevalence and surveillance of the genetic diversity of the helminthes targeting the mechanisms known to cause drug resistance. Assessing the prevalence of STH infections pre and post MDA is crucial for evaluating the effectiveness of the MDA program. This can further inform the policy makers on the future policies concerning the STH control including the resource allocation and control approaches. Incorporating genetic surveillance into STH control programs allows for early detection of drug resistance early enough and facilitates timely modifications to treatment strategies. This study aimed to compare the prevalence of STH infections pre- and post-deworming in school-aged children in Bungoma County, Western Kenya and to detect SNPs associated with drug resistance in these infections. A longitudinal study was conducted in 414 school-aged children. Stool samples were collected one month pre and two weeks post-MDA with MBZ. STH prevalence and infection intensities were determined using the Kato-Katz technique. Genomic DNA was extracted from eggs obtained from the stool samples pre- and post-deworming that were positive for A. lumbricoides infection. Standard PCR generated 564bp amplicons surrounding the target codons 167, 198, and 200 of the β-tubulin gene of A. lumbricoides, which have been associated with drug resistance phenotype in various helminth species. The amplicons were sequenced using Sanger sequencing to detect the presence of SNPs at these loci. Two hundred and two and 212 pre- and post-MDA stool samples were collected, respectively. The overall pre-MDA and post-MDA STH prevalence was 33% and 6%, respectively. A. lumbricoides was the dominant infecting helminth species with a prevalence of 31% and 4%, pre and post-MDA respectively. However, a few samples (<5) had hookworms, T. trichiura or mixed infections. Sequences were generated covering codons 167, 198, and 200 of the β-tubulin gene of A. lumbricoides. Sequencing of the DNA samples revealed that no β-tubulin SNPs were present in the A. lumbricoides infections. The significant reduction in prevalence post-MDA underscores the importance of continued investment in MDA interventions as part of integrated control strategies. The absence of detectable SNPs highlights the need for further research to understand the genetic diversity and population dynamics of STH in Bungoma County.