Multiple-marker (COI-5P, UPA, psb A and rbc L-3P) and two algorithmic approaches [automatic barcode gap discovery (ABGD) and Poisson tree process (PTP)] were used for species delimitation of Lithophylloideae in Brazil. The integrative approach was mostly congruent between markers and algorithmic methods of species delimitation, suggesting the occurrence of 24 species. Based on morphology and molecular data, Amphiroa rigida , Amphiroa vanbosseae , Lithophyllum atlanticum , Lithophyllum kaiseri , Lithophyllum margaritae , Titanoderma pustulatum , Titanoderma prototypum and Paulsilvella huveorum , which were previously reported for Brazil, are confirmed in this work. Six new species are distinguished by both molecular and morphological traits, and they are provisionally named as Amphiroa sp. 1, Amphiroa sp. 2, Amphiroa sp. 3, Lithophyllum sp. 1, Lithophyllum sp. 2 and Lithophyllum sp. 3. Another 10 species are cryptic and cannot be distinguished based on traditionally used morphological traits. These include Amphiroa sp. 4, Lithophyllum sp. 4, three species that are morphologically named under Amphiroa beauvoisii , and six that share the morphology described for Amphiroa fragilissima . All four markers used were useful for species delimitation. However, a combination of practical aspects and levels of intra- and interspecific divergence values led us to propose the use of rbc L-3P as a standard DNA barcode marker for the Corallinales.