Background: Strains of the Beijing sublineage of Mycobacterium tuberculosis have caused large outbreaks of tuberculosis, often involving multidrug resistance strains and this genetically highly conserved family of strains predominates in some geographic areas. For most of the countries of Latin America, no country-wide studies about the prevalence of the Beijing lineage are available. Methods: In this study, we determine the prevalence of the Beijing sublineage in Ecuador, using a large nation-wide sample of 991 isolates from the years 2014-2016 and with the strains, in case-related-proportional representation, emerging from most of the provinces of the country. The isolates were genotyped with asinglenucleotidespecific polymorphism (SNP) polymerase chain reaction for the Beijing sublineage. SNPpositive strains were confirmed as belonging to this lineage with 24 mycobacterial interspersed repetitive unitvariable number of tandem repeat and DNA sequencing. Results: We identified only four Beijing isolates in this collection of 991 strains and calculated a prevalence rate of 0.43%. Conclusions: Our study shows a limited dissemination of the Beijing strains in the Ecuadorian population. This in contrast with the neighbor countries of Peru and Colombia were locally a prevalence of up to 16% has been reported.