Molecular divergence in early-stage speciation of cryptic crickets: Loxoblemmus campestris and L. sylvestris

Understanding the minimal genetic changes underlying the earliest stages of speciation remains a central challenge in evolutionary biology. However, most model systems used to study early speciation involve pronounced ecological divergence, making it difficult to isolate the initial genetic changes associated with speciation. Here, we examined genetic differences between the cryptic cricket species Loxoblemmus campestris and L. sylvestris, which inhabit the same geographic region without pronounced ecological or morphological divergence. Despite their morphological similarity, the two species exhibited small but consistent molecular phylogenetic divergence in both mitochondrial and nuclear genomes. To investigate the genetic differences between these species, we conducted population genomic and comparative genomic analyses. We identified amino acid substitutions that were fixed within species, including both homozygous and consistently maintained heterozygous variants, across single-copy orthologous genes. Notably, only three genes— Raf, Slit, and Sur-8—had amino acid mutations with low evolutionary probability under neutral expectations and these substitutions are fixed within each species. All three genes are key components of the Ras/MAPK signaling pathway. These findings suggest that early speciation in L. campestris and L. sylvestris may have involved a limited number of mutations with potentially large functional effects, rather than widespread genomic divergence, prior to pronounced phenotypic differentiation. Cryptic Loxoblemmus species provide a useful model for understanding the earliest genomic events of speciation in the absence of pronounced phenotypic divergence.