An integrative taxonomic analysis combining mitochondrial COI sequences, morphology, colour pattern, and two species delimitation approaches revealed existence of five lineages within the Natal mountain catfish, Amphilius natalensis, in southern Africa. These lineages are separated by substantial genetic divergences (1.6 - 9.46%), and they can be consistently distinguished from one another based on a combination of morphology and colour pattern differences. Additionally, the lineages are allopatrically distributed and confined to isolated river systems draining discrete mountain ranges, which makes gene flow among them unlikely. One of these lineages is A. natalensis s.s. which is confined to the uMngeni and Tukela river systems in KwaZulu Natal (KZN) Province in South Africa. The other four lineages represent new species to science which are described as Amphilius zuluorum sp. nov., endemic to the uMkhomazi River system in KZN; Amphilius engelbrechti sp. nov. endemic to the Inkomati River system in Mpumalanga Province in South Africa; Amphilius marshalli sp. nov. endemic to the Pungwe and Lower Zambezi river systems in Zimbabwe and Mozambique; and Amphilius leopardus sp. nov. endemic to the Ruo River in Malawi. The results showed that A. laticaudatus Skelton 1984, which is endemic to the Buzi River system in Zimbabwe and Mozambique belongs to the A. natalensis s.l. complex. A redescription of A. laticaudatus is presented and an updated identification key for the mountain catfishes of southern Africa is provided. This article is protected by copyright. All rights reserved.