Abstract
Bidirectional best hits (BBH), which entails identifying the pairs of genes in two different genomes that are more similar to each other than either is to any other gene in the other genome, is a simple and widely used method to infer orthology. A recent study has analyzed the link between BBH and orthology in bacteria and archaea and concluded that, given the very high consistency in BBH they observed among triplets of neighboring genes, a high proportion of BBH are likely to be bona fide orthologs. However, limited by their analysis setup, the previous study could not easily test the reverse question: which proportion of orthologs are BBH? In this follow-up study, we consider this question in theory and answer it based on conceptual arguments, simulated data, and real biological data from all three domains of life. Our analyses corroborate the findings of the previous study, but also show that because of the high rate of gene duplication in plants and animals, as much as 60% of orthologous relations are missed by the BBH criterion.