A phylogeny was reconstructed for four species belonging to the Neotropical Anopheles (Nyssorhynchus) albitarsis complex using partial sequences from the mitochondrial cytochrome oxidase I (COI) and NADH dehydrogenase 4 (ND4) genes and the ribosomal DNA ITS2 and D2 expansion region of the 28S subunit. The basis for initial characterization of each member of the complex was by correlated random amplification of polymorphic DNA-polymerase chain reaction (RAPD-PCR) markers. Analyses were carried out with and without an outgroup (An. (Nys.) argyritarsis Robineau-Desvoidy) by using maximum parsimony, maximum likelihood, and Bayesian methods. A total evidence approach without the outgroup, using separate models for “fast” (COI and ND4 position 3) and “slow” (rDNA ITS2 and D2, and COI and ND4 position 1) partitions, gave the best supported topology, showing close relationships of An. albitarsis Lynch-Arribálzaga to An. albitarsis B and An. marajoara Galvão & Damasceno to An. deaneorum Rosa-Freitas. Analyses with the outgroup included showed poorer support, possibly because of a long branch attraction effect caused by a divergent outgroup, which caused one of the An. marajoara specimens to cluster with An. deaneorum in some analyses. The relationship of the above-mentioned result to a separately proposed hypothesis suggesting a fifth species in the complex is discussed.