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Blastocystis sp. is a widespread microorganism that colonizes the intestinal tract of several animals, including human beings, while its pathogenic role in humans is still under debate. The objective of the present study was to describe the frequency of Blastocystis sp. subtypes (STs) and their genetic variation within and among samples recovered from scholars inhabiting two rural villages with tropical climates and compare this information with previously documented data from arid and temperate zones in Mexico. Blastocystis sp. positive samples and ST identification were achieved by coprological analysis screening and Polymerase Chain Reaction-sequencing, respectively. Classical population genetics indexes (nucleotide diversity (π), haplotype polymorphism (θ), gene flow (Nm), genetic differentiation (ST), and Tajima's D) were calculated by comparing the sequences here obtained (n = 42) and those from previous studies from the arid (n = 80) and temperate (n = 61) climates from Mexico. Although Blastocystis sp. was the parasite most frequently found between 33% and 26% in both communities, only STs 1–3 were found. Haplotype network inference of Blastocystis sp. STs showed different haplotype profiles among STs vs. climate zones, although no specific haplotypes were identified for any particular climatic zone. Population genetics indexes showed different values within STs and climate zones (π and θ values ranged from 0.004 to 0.147; Nm > 4 and ST from 0.006 to 0.12). Our results show that Blastocystis sp. subtypes exhibit a different genetic variability profile according to the climate zone, suggesting a balancing process between the genetic variability within the Blastocystis sp. subtype and the number of haplotypes identified in each climate. • Blastocystis sp. subtypes (STs) 1–3 in rural communities of south-southeastern Mexico are reported for the first time. • Each Blastocystis sp. ST showed a different genetic variability depending on the climate zone. • A balancing process in the genetic variability within subtypes and their haplotypes in each climate zone may occur. [ABSTRACT FROM AUTHOR]