The family Pseudopimelodidae is widely distributed in South America and includes 51 described species organized in seven genera. Only two of four species of the genus Pseudopimelodus have been recorded for the trans-Andean basins of Colombia, Pseudopimelodus bufonius and P. schultzi , whose similarity in external morphology make their identification difficult. We performed a phylogenetic analysis using a fragment of the Cytochrome C Oxidase subunit 1 gene (COI), and analyzed osteological and traditional morphometric characters to study Pseudopimelodus from the Colombian trans-Andean region. Results provided strong support for two clades phylogenetically related to Pseudopimelodus, that showed clear-cut molecular, osteological, and morphometric differences from previously described bumblebee catfishes. Based on these results, we describe two Pseudopimelodus species from the Magdalena-Cauca River Basin : P. magnus sp. nov. with 43–44 vertebrae, dorsal-fin spine with serrations on its anterior margin; lateral margin of transverse process of the fourth vertebra of the Weberian complex forming an acute angle in ventral view and P. atricaudus sp. nov. with 39 vertebrae, dorsal-fin spine smooth on its anterior margin and a dark, vertical band covering 3/4 of the caudal fin with base of rays and tip of caudal-fin lobes hyaline.