Morphometry and DNA barcoding reveal cryptic diversity in the genus Enteromius (Cypriniformes: Cyprinidae) from the Congo basin, Africa

Marjolein Van Ginneken, Eva Decru, Erik Verheyen, Jos Snoeks


One of the main challenges to adequately conserve the African fish fauna is to improve our so far unsatisfactory taxonomic knowledge of important portions of the ichthyofauna. In the present study, we attempted to unravel the taxonomic diversity of some species of Enteromius Cope, 1867, a problematic African fish genus, recently collected in the north-eastern part of the Congo basin. We used an integrative approach, combining DNA barcodes and morphological analyses. For one of the species complexes found, the E. miolepis/eutaenia species complex, we evaluated taxonomic diversity over a larger geographic scale within the Congo drainage system. Although initial literature-based species identifications allowed us to assign all examined specimens to four tentative species, DNA barcodes indicated the presence of 23 distinct mitochondrial lineages. The majority of these lineages appeared endemic to particular rivers, and in most rivers multiple lineages occur in sympatry. Subsequent exploratory morphometric analyses indicated that almost all these lineages are morphologically distinguishable and that they may therefore represent undescribed species. As only a part of the Congo basin and a subset of the species diversity within Enteromius were examined, it appears that the species richness of Enteromius in the Congo basin is severely underestimated.


‘Barbus’; COI; integrative taxonomy; species richness

Full Text:




Agnèse J.F., Berrebi P., Lévêque C. & Guégan J.F. 1990. Two lineages, diploid and tetraploid, demonstrated in African species Barbus (Osteichthyes, Cyprinidae). Aquatic Living Resources 3: 305–311.

Bamba M. 2012. The ‘Barbus’ species (Cypriniformes, Cyprinidae) from Côte d’Ivoire: A Systematic Revision of Some West African Species Complexes. PhD thesis, KU Leuven, Belgium.

Bamba M., Vreven E.J. & Snoeks J. 2011. Description of Barbus teugelsi sp. nov. (Cypriniformes: Cyprinidae) from the Little Scarcies basin in Guinea, Africa. Zootaxa 2988: 48–65.

Banyankimbona G., Vreven E. & Snoeks J. 2012. ‘Barbus’ devosi, new species from the Malagarazi River basin in Burundi and Tanzania, East Africa (Cypriniformes: Cyprinidae). Ichthyological Exploration of Freshwaters 23: 181–192.

Becker S., Hanner R.H. & Steinke D. 2011. Five years of FISH-BOL: Brief status report. Mitochondrial DNA 22: 3–9.

Berrebi P. & Tsigenopoulos C.S. 2003. Phylogenetic organization of the genus Barbus sensu stricto: A review based on data obtained using molecular markers. In: Bãnãrescu P.M. & Bogutskaya N.G. (eds) The Freshwater Fishes of Europe Volume 5/II Cyprinidae 2, Part II: Barbus: 11–22. Aula-Verlag, Wiesbaden.

Berrebi P. & Valiushok D. 1998. Genetic divergence among morphotypes of Lake Tana (Ethiopia) barbs. Biological Journal of the Linnean Society 64: 369–384.

Berrebi P., Kottelat M., Skelton P. & Rab P. 1996. Systematics of Barbus: State of the art and heuristic comments. Folia Zoologica 45: 5–12.

Berrebi P., Chenuil A., Kotlik P., Machordom A. & Tsigenopoulos C.S. 2014. Disentangling the evolutionary history of the genus Barbus sensu latu, a twenty years adventure. In: Alves M.J., Cartaxana A., Correia A.M. & Lopes L.F. (eds) Professor Carlos Almaça (1934–2010) – Estado da arte em áreas científicas do seu interesse: 29–55. Museu Nacional de História Natural e da Ciência, Lisboa.

Bookstein F.L., Chernoff B., Elder R.L., Humphries J.M., Smith G.R. & Strauss R.E. 1985. Morpho-metrics in Evolutionary Biology: the Geometry of Size and Shape Change, with Examples from Fishes. Academy of Natural Sciences of Philadelphia, Philadelphia.

Collins R.A. & Cruickshank R.H. 2012. The seven deadly sins of DNA barcoding. Molecular Ecology Resources 13: 969–975.

Decru E., Vreven E. & Snoeks J. 2012. A revision of the West African Hepsetus (Characiformes; Hepsetidae) with a description of H. akawo sp. nov. and a redescription of H. odoe (Bloch, 1794). Journal of Natural History 46: 1–23.

Decru E., Moelants T., De Gelas K., Vreven E., Verheyen E. & Snoeks J. 2016. Taxonomic challenges in freshwater fishes: a mismatch between morphology and DNA barcoding in fish of the north-eastern part of the Congo basin. Molecular Ecology Resources 16: 342–352.

DeSalle R., Egan M.G. & Siddall M. 2011. The unholy trinity, taxonomy, species delimitation and DNA barcoding. Philosophical Transactions of the Royal Society B 360: 1905–1916.

Edgar R.C. 2004. MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Research 32: 1792–1797.

Fernández M.H. & Vrba E.S. 2005. A complete estimate of the phylogenetic relationships in Ruminantia: a dated species-level supertree of the extant ruminants. Biological Reviews 80: 269–302.

Fouquet A., Gilles A., Vences M., Marty C., Blanc M. & Gemmell N.J. 2007. Underestimation of species richness in Neotropical frogs revealed by mtDNA analyses. PLoS ONE 2: e1109.

Froese R. & Pauly D. 2017. FishBase. Available from [accessed 30 Mar. 2017].

Golubtsov A.S. & Krysanov E.Y. 1993. Karyological study of some cyprinid species from Ethiopia. The ploidy differences between large and small Barbus of Africa. Journal of Fish Biology 42: 445–455.

Goodier S.A.M., Cotterill F.P.D., O’Ryan C., Skelton P.H. & de Wit M.J. 2011. Cryptic diversity of African Tigerfish (Genus Hydrocynus) reveals palaeogeographic signatures of linked Neogene geotectonic events. PLoS ONE 6: e28875.

Hammer Ø., Harper D.A.T. & Ryan P.D. 2001. PAST: Paleontological statistics software package for education and data analysis. Palaeontologia Electronica 4: 1–9.

Hebert P.D.N., Cywinska A., Ball S.L. & de Waard J.R. 2003. Biological identifications through DNA barcodes. Proceedings of the Royal Society B 270: 313–321.

Ivanova N.V., Zemlak T.S., Hanner R.H. & Hebert P.D. 2007. Universal primer cocktails for fish DNA barcoding. Molecular Ecology Notes 7: 544–548.

Koblmüller S., Egger B., Sturmbauer C. & Sefc K.M. 2010. Rapid radiation, ancient incomplete lineage sorting and ancient hybridization in the endemic Lake Tanganyika cichlid tribe Tropheini. Molecular Phylogenetics and Evolution 55: 318–334.

Lavoué S. & Sullivan J.P. 2014. Petrocephalus boboto and Petrocephalus arnegardi, two new species of African electric fish (Osteoglossomorpha, Mormyridae) from the Congo River basin. ZooKeys 400: 43–65.

Lévêque C. & Daget J. 1984. Cyprinidae. In: Daget J., Gosse J.P. & Van den Audenaerde T.D. (eds) Check-list of the Freshwater Fishes of Africa (CLOFFA1): 217–342. ORSTOM & MRAC, Paris & Tervuren.

Lowenstein J.H., Osmundson T.Z., Becker S., Hanner R. & Stiassny M.L.J. 2011. Incorporating DNA barcodes into a multi-year inventory of the fishes of the hyperdiverse Lower Congo River, with a multi-gene performance assessment of the genus Labeo as a case study. Mitochondrial DNA 21: 1–19.

Messing J. 1983. New M13 vectors for cloning. Methods in Enzymology 101: 20–78.

Oellermann L.K. & Skelton P.H. 1990. Hexaploidy in yellowfish species (Barbus, Pisces, Cyprinidae) from southern Africa. Journal of Fish Biology 37: 105–115.

Olayemi A., Nicolas V., Hulselmans J.A.N., Missoup A.D., Fichet-Calvet E., Amundala D., Dudu A., Dierckx T., Wendelen W., Leirs H. & Verheyen E. 2012. Taxonomy of the African giant pouched rats (Nesomyidae: Cricetomys): molecular and craniometric evidence support an unexpected high species diversity. Zoological Journal of the Linnean Society 165: 700–719.

Pereira L.H.G., Hanner R., Foresti F. & Oliveira C. 2013. Can DNA barcoding accurately discriminate megadiverse Neotropical freshwater fish fauna? BMC Genetics 14: 20.

Pethiyagoda R., Meegaskumbura M. & Maduwage K. 2012. A synopsis of the South Asian fishes referred to Puntius (Pisces: Cyprinidae). Ichthyological Exploration of Freshwaters 23: 69–95.

Poll M. 1976. Exploration du Parc National de l’Upemba Mission G.F. de Witte, 73: Poissons. Fondation pour favoriser les recherches scientifiques en Afrique, Bruxelles.

Rab P., Machordom A., Perdices A. & Guegan J.F. 1995. Karyotypes of three «small» Barbus species (Cyprinidae) from Republic of Guinea (Western Africa) with a review on karyology of African small Barbus. Caryologia 48: 299–307.

Ren Q. & Mayden R.L. 2016. Molecular phylogeny and biogeography of African diploid barbs, ‘Barbus’, and allies in Africa and Asia (Teleostei: Cypriniformes). Zoologica Scripta 45 (6): 642–649.

Runge J. 2007. The Congo River, Central Africa. In: Gupta A. (ed.) Large Rivers: Geomorphology and Management: 293–309. John Wiley & Sons Ltd, West Sussex.

Schmidt R.C. & Bart Jr. H.L. 2015. Nomenclatural changes should not be based on equivocally supported phylogenies: Reply to Yang et al. 2015. Molecular Phylogenetics and Evolution 90: 193–194.

Snoeks J. 2004. The Cichlid Diversity of Lake Malawi/Nyassa/Niassa: Identification, Distribution and Taxonomy. Cichlid Press, Texas.

Snoeks J., Harrison I.J. & Stiassny M.L.J. 2011. The status and distribution of freshwater fishes. In: Darwall W., Smith K., Allen D., Holland R., Harrison I. & Brooks E. (eds) The Diversity of Life in African Freshwaters: Underwater, under Threat. An Analysis of the Status and Distribution of Freshwater Species throughout Mainland Africa: 42–91. IUCN, Gland & Cambridge.

Steinke D. & Hanner R. 2011. The FISH-BOL collaborators’ protocol. Mitochondrial DNA 22: 10–14.

Stiassny M.L., Denton J.S. & Monsebula Iyaba R.J.C. 2013. A new ectoparasitic distichodontid of the genus Eugnathichthys (Characiformes: Citharinoidei) from the Congo basin of central Africa, with a molecular phylogeny for the genus. Zootaxa 3693: 479–490.

Swartz E.R., Skelton P.H. & Bloomer P. 2009. Phylogeny and biogeography of the genus Pseudobarbus (Cyprinidae): shedding light on the drainage history of rivers associated with the Cape Floristic Region. Molecular Phylogenetics and Evolution 51: 75–84.

Tamura K., Peterson D., Peterson N., Stecher G., Nei M. & Kumar S. 2011. MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Molecular Biology and Evolution 28: 2731–2739.

Tsigenopoulos C.S., Kasapidis P. & Berrebi P. 2010. Phylogenetic relationships of hexaploid large-sized barbs (genus Labeobarbus, Cyprinidae) based on mtDNA data. Molecular Phylogenetics and Evolution 56: 851–856.

Tweddle D. & Skelton P.H. 2008. New species of ‘Barbus’ and Labeobarbus (Teleostei: Cyprinidae) from the South Rukuru River, Malawi, Africa. Smithiana Publications in Aquatic Biodiversity, Bulletin 8: 25–39.

Yang L., Sado T., Hirt M.V., Pasco-Viel E., Arunachalam M., Li J., Wang X., Freyhof J., Saitoh K., Simons A.M., Miya M., He S. & Mayden R.L. 2015. Phylogeny and polyploidy: Resolving the classification of cyprinine fishes (Teleostei: Cypriniformes). Molecular Phylogenetics and Evolution 85: 97–116.


  • There are currently no refbacks.

License URL: