BACKGROUND: Plasmodium vivax malaria (Pv-malaria) is still considered a neglected disease despite an alarming number of individuals being infected annually. Malaria pathogenesis occurs with the onset of the vector-parasite-host interaction through the binding of pathogen-associated molecular patterns (PAMPs) and receptors of innate immunity, such as toll-like receptors (TLRs). The triggering of the signaling cascade produces an elevated inflammatory response. Genetic polymorphisms in TLRs are involved in susceptibility or resistance to infection, and the identification of genes involved with Pv-malaria response is important to elucidate the pathogenesis of the disease and may contribute to the formulation of control and elimination tools. METHODOLOGY/PRINCIPAL FINDINGS: A retrospective case-control study was conducted in an intense transmission area of Pv-malaria in the state of Amazonas, Brazil. Genetic polymorphisms (SNPs) in different TLRs, TIRAP, and CD14 were genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis in 325 patients infected with P. vivax and 274 healthy individuals without malaria history in the prior 12 months from the same endemic area. Parasite load was determined by qPCR. Simple and multiple logistic/linear regressions were performed to investigate association between the polymorphisms and the occurrence of Pv-malaria and parasitemia. The C/T (TLR5 R392StopCodon) and T/T (TLR9 -1486C/T) genotypes appear to be risk factors for infection by P. vivax (TLR5: C/C vs. C/T [OR: 2.116, 95% CI: 1.054-4.452, p = 0.031]; TLR9: C/C vs. T/T [OR: 1.919, 95% CI: 1.159-3.177, p = 0.010]; respectively). Fever (COEF = 7599.46, 95% CI = 3063.80-12135.12, p = 0.001) and the C/C genotype of TLR9 -1237C/T (COEF = 17006.63, 95% CI = 3472.83-30540.44, p = 0.014) were independently associated with increased parasitemia in patients with Pv-malaria. CONCLUSIONS: Variants of TLRs may predispose individuals to infection by P. vivax. The TLR5 R392StopCodon and TLR9 -1486C/T variants are associated with susceptibility to Pv-malaria. Furthermore, the TLR9 variant -1237C/C correlates with high parasitemia.