Novel zoonotic cases of Plasmodium simium from São Paulo with a reference genome of the Brazilian strain.

Zoonotic malaria, caused by parasites like Plasmodium simium, poses challenges to control and elimination efforts due to undetected reservoirs and altered transmission dynamics. Recent outbreaks in Brazil underscore the importance of P. simium spillovers from non-human primates. Genomic studies are crucial for identifying molecular markers of host adaptation and improving diagnostic tools; however, a high-quality reference genome is needed. To address this, we used Oxford Nanopore and Illumina sequencing platforms to generate a high-quality P. simium reference (Ps_SãoPaulo, 21.8 Mb). Comparative genomic analyses confirmed known differences with P. vivax, including mitochondrial mutations, deletions in reticulocyte-invasion genes (RBP2a, DBP), and variants in blood-stage antigen genes (msp3.1, msp3.2). Genome-wide analyses of 38 P. simium samples from Brazil revealed expected divergence from P. vivax and potential unique signatures of selection and host adaptation. This study provides the most comprehensive P. simium reference genome, advancing our understanding of its genomic diversity, with insights for malaria control in Brazil.