Malaria particularly burdens people in poor and neglected settings across the tropics of Africa. Meanwhile, a large proportion of the Togo population have poor understanding of malaria epidemiology and parasites. This study carried out a molecular survey of malaria cases in southern Togo during 2017–2019. We estimated Plasmodium species infection rates and microscopic examination compliance with nested PCR results. Sensitivity and specificity analyses were performed in conjunction with predictive values. Also, phylogenetic characterization of species of malaria parasites was assessed. Plasmodium genus-specific nested PCR identified 565 positive cases including 536/611 (87.8%) confirmed cases from the microscopy-positive group and 29/199 (14.6%) diagnosed malaria cases from the microscopy-negative group. Our findings revealed a disease prevalence (69.8%) higher than that reported (25.5–55.1%) for the country. The diagnostic test had 94.9% sensitivity and 69.4% specificity, i.e., it missed 120 of the people who had malaria and about one-third of the people tested positive for the disease, which they did not have, respectively. In conjunction, the test showed 87.7% positive predictive value and 85.4% negative predictive value, which, from a clinical perspective, indicates the chance that a person with a positive diagnostic test truly has the disease and the probability that a person with a negative test does not have the disease, respectively. Further species-specific nested PCR followed by analysis of gene sequences confirmed species of malaria parasites and indicated infection rates for Plasmodium falciparum (Pf), 95.5% (540/565); P. ovale (Po), 0.5% (3/565); and P. malariae (Pm), 0.4% (2/565). In addition, 20 cases were coinfection cases of Pf-Po (15/565) and Pf-Pm (5/565). This study publicly reports, for the first time, a molecular survey of malaria cases in Togo and reveals the presence of other malaria parasites (Po and Pm) other than Pf. These findings might provide answers to some basic questions on the malaria scenario and, knowledge gained could help with intervention deployment for effective malaria control in Togo.