Culture-free genomics: a shift toward genome-wide applications in chagas disease and leishmaniasis

INTRODUCTION: Chagas disease and leishmaniasis remain major neglected tropical diseases, with diagnosis and surveillance constrained by low parasite burden, multiclonal infections, and complex parasite biology. Traditional culture-dependent and targeted molecular approaches fail to capture the full genomic diversity of Trypanosoma cruzi and Leishmania spp. limiting clinical and epidemiological utility. High-throughput sequencing has enabled a shift toward culture-free genomic approaches, allowing direct parasite DNA analysis from host-derived samples. AREAS COVERED: We review the evolution from early sequencing to second- and third-generation platforms, highlighting culture-free detection and genomic surveillance. We discuss enrichment strategies (selective whole-genome amplification (SWGA) and capture-enrichment sequencing (CES)) addressing low parasite DNA abundance in complex samples, alongside metagenomics and portable sequencing for field-based surveillance and diagnostics. Biological and technical challenges are examined, including genomic complexity, structural variation, life-cycle-driven plasticity, and fragmented reference genomes across T. cruzi discrete typing units (DTUs) and Leishmania species. We further explore how direct-from-host data can improve diagnostics, enhance transmission surveillance, support treatment monitoring, and guide control strategies. EXPERT OPINION: Culture-free genomic approaches represent a transformative advance in kinetoplastid research, providing resolution that culture-dependent methods cannot deliver. Their diagnostic contribution is at present largely indirect, operating through the identification of improved molecular and serological targets rather than through sequencing as the assay itself. Persistent barriers of cost, infrastructure, standardization, and bioinformatics capacity, together with the absence of formal clinical validation, currently confine these methods to research and surveillance settings. Successful translation will require methodological innovation, expanded genomic resources, and investment to bridge the gap between research and public health.