Simple trichal types constitute a group of cyanobacteria with an abundance of novel, often cryptic taxa. Here, we investigated material collected from wet surface-soil in a saline environment in Petchaburi Province, central Thailand. A morphological comparison of the isolated strain with similar known species, as well as its phylogenetic and species delimitation analyses based on the combined datasets of other related organisms, especially simple trichal cyanobacteria, revealed that the material of this study represented an independent taxon. Using a multifaceted method, we propose that this material represents a new genus, Thainema gen. nov., belonging to the family Leptolyngbyaceae, with the type species Thainema salinarum sp. nov. This novel taxon shares similar ecological habitats with strains previously placed in the same lineage.

Department of Biology Faculty of Science Chiang Mai University Chiang Mai Thailand

Department of Botany Faculty of Science University of South Bohemia České Budějovice Czech Republic

Zobrazit více v PubMed

Lemes-da-Silva NM, Branco LHZ, Necchi O Júnior, Necchi O, Necchi O Júnior. Corticolous cyanobacteria from tropical forest remnants in northwestern São Paulo State, Brazil. Brazilian J Bot. 2012;35: 169–179.

Schirrmeister BE, Antonelli A, Bagheri HC. The origin of multicellularity in cyanobacteria. BMC Evol Biol. 2011;11: 45. doi: 10.1186/1471-2148-11-45 . PubMed DOI PMC

Satyanarayana T, Raghukumar C, Shivaji S. Extremophilic microbes: Diversity and perspectives. Curr Sci. 2005;89: 78–90. http://www.jstor.org/stable/24110434.

DasSarma S, Arora P. Halophiles. e LS. 2001. doi: 10.1038/npg.els.0000394 DOI

Dennis PP, Shimmin LC. Evolutionary divergence and salinity-mediated selection in halophilic archaea. Microbiol Mol Biol Rev. 1997;61: 90–104. doi: 10.1128/mmbr.61.1.90-104.1997 . PubMed DOI PMC

Miller SR, Castenholz RW. The evolution of thermotolerance in hot spring cyanobacteria of the genus Synechococcus. J Phycol. 2000;36: 48. doi: 10.1128/AEM.66.10.4222-4229.2000 . PubMed DOI PMC

Castenholz RW. Species usage, concept, and evolution in the cyanobacteria (blue-green algae). J Phycol. 1992;28: 737–745. doi: 10.1007/s10531-015-0888-6 DOI

Johansen JR, Casamatta DA. Recognizing cyanobacterial diversity through adoption of a new species paradigm. Algol Stud für Hydrobiol Suppl Vol. 2005;117: 71–93. doi: 10.1127/1864-1318/2005/0117-0071 DOI

Komárek J, Kaštovský J, Mareš J, Johansen JR. Taxonomic classification of cyanoprokaryotes (cyanobacterial genera) 2014, using a polyphasic approach. Preslia. 2014;86: 295–335.

Dvořák P, Poulíčková A, Hašler P, Belli M, Casamatta DA, Papini A. Species concepts and speciation factors in cyanobacteria, with connection to the problems of diversity and classification. Biodivers Conserv. 2015;24: 739–757. doi: 10.1007/s10531-015-0888-6 DOI

Stanier RY, Deruelles J, Rippka R, Herdman M, Waterbury JB. Generic Assignments, Strain Histories and Properties of Pure Cultures of Cyanobacteria. Microbiology. 1979;111: 1–61. doi: 10.1099/00221287-111-1-1 DOI

Castenholz RW, Wilmotte A, Herdman M, Rippka R, Waterbury JB, Iteman I, et al. Phylum BX. cyanobacteria. Bergey’s manual® of systematic bacteriology. Springer; 2001.

Becerra-Absalón I, Muñoz-Martín M, Montejano G, Mateo P. Differences in the cyanobacterial community composition of biocrusts from the drylands of Central Mexico. Are there endemic species? Front Microbiol. 2019;10: 937. doi: 10.3389/fmicb.2019.00937 . PubMed DOI PMC

Mai T, Johansen JR, Pietrasiak N, Bohunická M, Martin MP. Revision of the Synechococcales (Cyanobacteria) through recognition of four families including Oculatellaceae fam. nov. and Trichocoleaceae fam. nov. and six new genera containing 14 species. Phytotaxa. 2018;365: 1–59. doi: 10.11646/PHYTOTAXA.365.1.1 DOI

Giovannoni SJ, Turner S, Olsen GJ, Barns S, Lane DJ, Pace NR. Evolutionary relationships among cyanobacteria and green chloroplasts. J Bacteriol. 1988;170: 3584–3592. doi: 10.1128/jb.170.8.3584-3592.1988 . PubMed DOI PMC

Turner S, Pryer KM, Miao VPW, Palmer JD. Investigating deep phylogenetic relationships among cyanobacteria and plastids by small subunit rRNA sequence analysis. J Eukaryot Microbiol. 1999;46: 327–338. doi: 10.1111/j.1550-7408.1999.tb04612.x . PubMed DOI

Wilmotte A, Herdman M. Phylogenetic relationships among the cyanobacteria based on 16S rRNA sequences. Bergey’s Man Syst Bacteriol Vol One Archaea Deep Branch phototrophic Bact. 2001; 487–493. http://www.jstor.org/stable/20794265.

Palinska KA, Surosz W. Taxonomy of cyanobacteria: A contribution to consensus approach. Hydrobiologia. 2014;740: 1–11. doi: 10.1007/s10750-014-1971-9 DOI

Johansen JR, Kovacik L, Casamatta DA, Iková KF, Kastovský J. Utility of 16S-23S ITS sequence and secondary structure for recognition of intrageneric and intergeneric limits within cyanobacterial taxa: Leptolyngbya corticola sp. nov.(Pseudanabaenaceae, Cyanobacteria). Nov Hedwigia. 2011;92: 283. doi: 10.1127/0029-5035/2011/0092-0283 DOI

Boyer SL, Flechtner VR, Johansen JR. Is the 16S–23S rRNA internal transcribed spacer region a good tool for use in molecular systematics and population genetics? A case study in cyanobacteria. Mol Biol Evol. 2001;18: 1057–1069. doi: 10.1093/oxfordjournals.molbev.a003877 . PubMed DOI

Casamatta DA, Gomez SR, Johansen JR. Rexia erecta gen. et sp. nov. and Capsosira lowei sp. nov., two newly described cyanobacterial taxa from the Great Smoky Mountains National Park (USA). Hydrobiologia. 2006;561: 13–26. doi: 10.1007/s10750-005-1602-6 DOI

Komárková J, Zapomělová E, Komárek J. Chakia (cyanobacteria), a new heterocytous genus from Belizean marshes identified on the basis of the 16S rRNA gene. Fottea. 2013;13: 227–233. doi: 10.5507/fot.2013.018 DOI

Kilgore C, Johansen JR, Mai T, Hauer T, Casamata DA, Sheil C. Molecular characterization of Geitleria appalachiana sp. nov.(Nostocales, Cyanobacteria) and formation of Geitleriaceae fam. nov. Fottea, Olomouc. 2018. doi: 10.5507/fot.2018.002 DOI

Malavasi V, Škaloud P, Rindi F, Tempesta S, Paoletti M, Pasqualetti M. DNA-based taxonomy in ecologically versatile microalgae: a re-evaluation of the species concept within the coccoid green algal genus Coccomyxa (Trebouxiophyceae, Chlorophyta). PLoS One. 2016;11. doi: 10.1371/journal.pone.0151137 e0151137. PubMed DOI PMC

Košuthová A, Bergsten J, Westberg M, Wedin M. Species delimitation in the cyanolichen genus Rostania. BMC Evol Biol. 2020;20: 1–17. PubMed PMC

Wiens JJ. Species delimitation: new approaches for discovering diversity. Syst Biol. 2007;56: 875–878. doi: 10.1080/10635150701748506 . PubMed DOI

Sites JW Jr, Marshall JC. Delimiting species: a Renaissance issue in systematic biology. Trends Ecol Evol. 2003;18: 462–470. doi: 10.1016/S0169-5347(03)00184-8 DOI

Dvořák P, Casamatta DA, Hašler P, Jahodářová E, Norwich AR, Poulíčková A. Diversity of the cyanobacteria. Modern topics in the phototrophic prokaryotes. Springer; 2017. pp. 3–46. doi: 10.1007/978-3-319-46261-5 DOI

Miscoe LH, Johansen JR, Kociolek JP, Lowe RL, Vaccarino MA, Pietrasiak N, et al.. Novel cyanobacteria from caves on Kauai, Hawaii. The diatom flora and cyanobacteria from caves on Kauai, Hauwaii. Borntraeger, 2016; 2016. pp. 75–152.

Osorio-santos K, Pietrasiak N, Bohunická M, Laura H, Kováčik L, Martin MP, et al.. Seven new species of Oculatella (Pseudanabaenales, Cyanobacteria): taxonomically recognizing cryptic diversification. Eur J Phycol. 2014;49: 450–470. doi: 10.1080/09670262.2014.976843 DOI

Perkerson RB, Johansen JR, Kovácik L, Brand J, Kaštovský J, Casamatta DA, et al.. A unique pseudanabaenalean (cyanobacteria) genus Nodosilinea gen. nov. based on morphological and molecular data. J Phycol. 2011;47: 1397–1412. doi: 10.1111/j.1529-8817.2011.01077.x PubMed DOI

Zammit G, Billi D, Albertano P. The subaerophytic cyanobacterium Oculatella subterranea (Oscillatoriales, Cyanophyceae) gen. et sp. nov.: A cytomorphological and molecular description. Eur J Phycol. 2012;47: 341–354. doi: 10.1080/09670262.2012.717106 DOI

Taton A, Wilmotte A, Šmarda J, Elster J, Komarek J, Komárek J. Plectolyngbya hodgsonii: a novel filamentous cyanobacterium from Antarctic lakes. Polar Biol. 2011;34: 181–191. doi: 10.1007/s00300-010-0868-y DOI

Komárek J. Bd. 19/1: Cyanoprokaryota: teil: Chroococcales. Subwasserflora von Mitteleuropa. Gustav Fischer, Jena Stuttgart Lubeck Ulm; 1999.

Li X, Li R. Limnolyngbya circumcreta gen. & comb. nov. (Synechococcales, Cyanobacteria) with three geographical (provincial) genotypes in China. Phycologia. 2016;55: 478–491. doi: 10.2216/15-149.1 DOI

Vaz MGMV, Genuário DB, Andreote APD, Malone CFS, Sant’Anna CL, Barbiero L, et al.. Pantanalinema gen. nov. and Alkalinema gen. nov.: Novel pseudanabaenacean genera (Cyanobacteria) isolated from saline–alkaline lakes. Int J Syst Evol Microbiol. 2015;65: 298–308. doi: 10.1099/ijs.0.070110-0 . PubMed DOI

Abed RMM, Garcia-Pichel F, Hernández-Mariné M. Polyphasic characterization of benthic, moderately halophilic, moderately thermophilic cyanobacteria with very thin trichomes and the proposal of Halomicronema excentricum gen. nov., sp. nov. Arch Microbiol. 2002;177: 361–370. doi: 10.1007/s00203-001-0390-2 . PubMed DOI

Fourçans A, Solé A, Diestra E, Ranchou-Peyruse A, Esteve I, Caumette P, et al.. Vertical migration of phototrophic bacterial populations in a hypersaline microbial mat from Salins-de-Giraud (Camargue, France). FEMS Microbiol Ecol. 2006;57: 367–377. doi: 10.1111/j.1574-6941.2006.00124.x PubMed DOI

Chatchawan T, Peerapornpisal Y, Komárek J. Diversity of cyanobacteria in man-made solar saltern, Petchaburi Province, Thailand—A pilot study. Fottea. 2011;11: 203–214. doi: 10.5507/fot.2011.019 DOI

Komárek J, Anagnostidis K. Bd. 19/2: Cyanoprokaryota: teil 2: Oscillatoriales. Elsevier, München; 2005.

Spurr AR. A low-viscosity epoxy resin embedding medium for electron microscopy. J Ultrastruct Res. 1969;26: 31–43. doi: 10.1016/s0022-5320(69)90033-1 . PubMed DOI

Wilmotte A, Van der Auwera G, De Wachter R. Structure of the 16 S ribosomal RNA of the thermophilic cyanobacterium chlorogloeopsis HTF (’mastigocladus laminosus HTF’) strain PCC7518, and phylogenetic analysis. FEBS Lett. 1993;317: 96–100. doi: 10.1016/0014-5793(93)81499-p . PubMed DOI

Taton A, Grubisic S, Brambilla E, De Wit R, Wilmotte A. Cyanobacterial diversity in natural and artificial microbial mats of Lake Fryxell (McMurdo Dry Valleys, Antarctica): A morphological and molecular approach. Appl Environ Microbiol. 2003;69: 5157–5169. doi: 10.1128/AEM.69.9.5157-5169.2003 . PubMed DOI PMC

Seo P-S, Yokota A. The phylogenetic relationships of cyanobacteria inferred from 16S rRNA, gyrB, rpoC1 and rpoD1 gene sequences. J Gen Appl Microbiol. 2003;49: 191–203. doi: 10.2323/jgam.49.191 . PubMed DOI

Rudi K, Skulberg OM, Jakobsen KS. Evolution of cyanobacteria by exchange of genetic material among phyletically related strains. J Bacteriol. 1998;180: 3453–3461. doi: 10.1128/JB.180.13.3453-3461.1998 . PubMed DOI PMC

Nübel U, Garcia-Pichel F, Muyzer G. PCR primers to amplify 16S rRNA genes from cyanobacteria. Appl Environ Microbiol. 1997;63: 3327–3332. doi: 10.1128/aem.63.8.3327-3332.1997 . PubMed DOI PMC

Hall T. BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symp. Ser. 1999.

Katoh K, Rozewicki J, Yamada KD. MAFFT online service: multiple sequence alignment, interactive sequence choice and visualization. Brief Bioinform. 2019;20: 1160–1166. doi: 10.1093/bib/bbx108 . PubMed DOI PMC

Tamura K, Stecher G, Peterson D, Filipski A, Kumar S. MEGA6: molecular evolutionary genetics analysis version 6.0. Mol Biol Evol. 2013;30: 2725–2729. doi: 10.1093/molbev/mst197 . PubMed DOI PMC

Lanfear R, Frandsen PB, Wright AM, Senfeld T, Calcott B. PartitionFinder 2: new methods for selecting partitioned models of evolution for molecular and morphological phylogenetic analyses. Mol Biol Evol. 2017;34: 772–773. doi: 10.1093/molbev/msw260 . PubMed DOI

Trifinopoulos J, Nguyen L-T, von Haeseler A, Minh BQ. W-IQ-TREE: a fast online phylogenetic tool for maximum likelihood analysis. Nucleic Acids Res. 2016;44: W232–W235. doi: 10.1093/nar/gkw256 . PubMed DOI PMC

Minh BQ, Schmidt HA, Chernomor O, Schrempf D, Woodhams MD, Von Haeseler A, et al.. IQ-TREE 2: new models and efficient methods for phylogenetic inference in the genomic era. Mol Biol Evol. 2020;37: 1530–1534. doi: 10.1093/molbev/msaa015 . PubMed DOI PMC

Rambaut A, Drummond AJ, Xie D, Baele G, Suchard MA. Posterior summarization in Bayesian phylogenetics using Tracer 1.7. Syst Biol. 2018;67: 901. doi: 10.1093/sysbio/syy032 . PubMed DOI PMC

Rambaut A. FigTree Version 1.4. 4. 2020. (default version). Reference Source. http://tree.bio.ed.ac.uk/software/figtree/.

Řeháková K, Johansen JR, Bowen MB, Martin MP, Sheil CA. Variation in secondary structure of the 16s rRNA molecule in cyanobacteria with implications for phylogenetic analysis. Fottea. 2014;14: 161–178. doi: 10.5507/fot.2014.013 DOI

Drummond AJ, Suchard MA, Xie D, Rambaut A. Bayesian phylogenetics with BEAUti and the BEAST 1.7. Mol Biol Evol. 2012;29: 1969–1973. doi: 10.1093/molbev/mss075 . PubMed DOI PMC

Zimba PV, Huang I, Foley JE, Linton EW. Identification of a new-to-science cyanobacterium, Toxifilum mysidocida gen. nov. & sp. nov.(Cyanobacteria, Cyanophyceae). J Phycol. 2017;53: 188–197. doi: 10.1111/jpy.12490 . PubMed DOI

Yarza P, Yilmaz P, Pruesse E, Glöckner FO, Ludwig W, Schleifer KH, et al.. Uniting the classification of cultured and uncultured bacteria and archaea using 16S rRNA gene sequences. Nat Rev Microbiol. 2014;12: 635–645. doi: 10.1038/nrmicro3330 . PubMed DOI

González-Resendiz L, Johansen JR, León-Tejera H, Sánchez L, Segal-Kischinevzky C, Escobar-Sánchez V, et al.. A bridge too far in naming species: a total evidence approach does not support recognition of four species in Desertifilum (Cyanobacteria). J Phycol. 2019;55: 898–911. doi: 10.1111/jpy.12867 PubMed DOI

Stackebrandt E. Taxonomic parameters revisited: tarnished gold standards. Microbiol Today. 2006;33: 152–155.

Turner S. Molecular systematics of oxygenic photosynthetic bacteria. Orig Algae their Plast. 1997; 13–52. doi: 10.1007/978-3-7091-6542-3_2 DOI

Řeháková K, Johansen JR, Casamatta DA, Xuesong L, Vincent J. Morphological and molecular characterization of selected desert soil cyanobacteria: three species new to science including Mojavia pulchra gen. et sp. nov. Phycologia. 2007;46: 481–502.

Komarek J. Cyanobacterial taxonomy: current problems and prospects for the integration of traditional and molecular approaches. Algae. 2006;21: 349–375.

Gupta RS, Naushad S, Baker S. Phylogenomic analyses and molecular signatures for the class Halobacteria and its two major clades: a proposal for division of the class Halobacteria into an emended order Halobacteriales and two new orders, Haloferacales ord. nov. and Natrialbales ord. n. Int J Syst Evol Microbiol. 2015;65: 1050–1069. doi: 10.1099/ijs.0.070136-0 . PubMed DOI

Fučíková K, Lewis PO, Lewis LA. Putting incertae sedis taxa in their place: a proposal for ten new families and three new genera in Sphaeropleales (Chlorophyceae, Chlorophyta). J Phycol. 2014;50: 14–25. doi: 10.1111/jpy.12118 . PubMed DOI

Sawana A, Adeolu M, Gupta RS. Molecular signatures and phylogenomic analysis of the genus Burkholderia: proposal for division of this genus into the emended genus Burkholderia containing pathogenic organisms and a new genus Paraburkholderia gen. nov. harboring environmental species. Front Genet. 2014;5: 429. doi: 10.3389/fgene.2014.00429 . PubMed DOI PMC

Adeolu M, Gupta RS. Phylogenomics and molecular signatures for the order Neisseriales: proposal for division of the order Neisseriales into the emended family Neisseriaceae and Chromobacteriaceae fam. nov. Antonie Van Leeuwenhoek. 2013;104: 1–24. doi: 10.1007/s10482-013-9920-6 . PubMed DOI

Zammit G. Systematics and biogeography of sciophilous cyanobacteria; an ecological and molecular description of Albertania skiophila (Leptolyngbyaceae) gen. & sp. nov. Phycologia. 2018;57: 481–491. doi: 10.2216/17-125.1 DOI

Chakraborty S, Maruthanayagam V, Achari A, Pramanik A, Jaisankar P, Mukherjee J. Euryhalinema mangrovii gen. nov., sp. nov. And Leptoelongatus litoralis gen. nov., sp. nov. (Leptolyngbyaceae) isolated from an Indian mangrove forest. Phytotaxa. 2019;422: 58–74. doi: 10.11646/phytotaxa.422.1.4 DOI

Hauer T. & Komárek J. CyanoDB 2.0—On-line database of cyanobacterial genera.—World-wide electronic publication. Univ. of South Bohemia & Inst. of Botany AS CR. 2021. http://www.cyanodb.cz.

Turland NJ, Wiersema JH, Barrie FR, Greuter W, Hawksworth DL, Herendeen PS, et al.. International Code of Nomenclature for algae, fungi, and plants (Shenzhen Code) adopted by the Nineteenth International Botanical Congress Shenzhen, China, July 2017. Koeltz Botanical Books; 2018.

Garrity GM, Parker CT, Tindall BJ. International code of nomenclature of prokaryotes. Int J Syst Evol Microbiol. 2015;90. doi: 10.1099/ijsem.0.000778 . PubMed DOI

Blank CE, Hinman NW. Cyanobacterial and algal growth on chitin as a source of nitrogen; ecological, evolutionary, and biotechnological implications. Algal Res. 2016;15: 152–163. doi: 10.1016/j.algal.2016.02.014 DOI

Brito Â, Ramos V, Mota R, Lima S, Santos A, Vieira J, et al.. Description of new genera and species of marine cyanobacteria from the Portuguese Atlantic coast. Mol Phylogenet Evol. 2017;111: 18–34. doi: 10.1016/j.ympev.2017.03.006 . PubMed DOI

Caires TA, de Mattos Lyra G, Hentschke GS, de Gusmão Pedrini A, Sant’Anna CL, de Castro Nunes JM. Neolyngbya gen. nov. (Cyanobacteria, Oscillatoriaceae): A new filamentous benthic marine taxon widely distributed along the Brazilian coast. Mol Phylogenet Evol. 2018;120: 196–211. doi: 10.1016/j.ympev.2017.12.009 . PubMed DOI

Akagha SC, Johansen JR, Nwankwo DI, Yin K. Lagosinema tenuis gen. et sp. nov. (Prochlorotrichaceae, Cyanobacteria): a new brackish water genus from Tropical Africa. Fottea. 2019;19: 1–12. doi: 10.5507/fot.2018.012 DOI

Sherwood AR, Conklin KY, Liddy ZJ. What’s in the air? Preliminary analyses of Hawaiian airborne algae and land plant spores reveal a diverse and abundant flora. Phycologia. 2014;53: 579–582. doi: 10.2216/14-059.1 DOI

Bornet E, Flahault C. Revision des Nostocacées hétérocystées contenues dans les principaux herbiers de France. H. R. Engelmann; 1888.

Pietrasiak N, Osorio-Santos K, Shalygin S, Martin MP, Johansen JR. When is a lineage a species? A case study in Myxacorys gen. nov.(Synechococcales: Cyanobacteria) with the description of two new species from the Americas. J Phycol. 2019;55: 976–996. doi: 10.1111/jpy.12897 . PubMed DOI

Mishler BD. The phylogenetic species concept (sensu Mishler and Theriot): monophyly, apomorphy, and phylogenetic species concepts. Species concepts and phylogenetic theory, a debate. New York: Columbia University Press.; 2000. pp. 44–54.

Rosselló-Mora R, Amann R. The species concept for prokaryotes. FEMS Microbiol Rev. 2001;25: 39–67. PubMed

Bohunická M, Pietrasiak N, Johansen JR, Gómez EB, Hauer T, Gaysina LA, et al.. Roholtiella, gen. nov. (Nostocales, Cyanobacteria)—A tapering and branching cyanobacteria of the family Nostocaceae. Phytotaxa. 2015;197: 84–103. doi: 10.11646/PHYTOTAXA.197.2.2 DOI

Xu X, Liu F, Ono H, Chen J, Kuntner M, Li D. Targeted sampling in Ryukyus facilitates species delimitation of the primitively segmented spider genus Ryuthela (Araneae: Mesothelae: Liphistiidae). Zool J Linn Soc. 2017;181: 867–909.

Giarla TC, Voss RS, Jansa SA. Hidden diversity in the Andes: comparison of species delimitation methods in montane marsupials. Mol Phylogenet Evol. 2014;70: 137–151. doi: 10.1016/j.ympev.2013.09.019 . PubMed DOI

Derkarabetian S, Hedin M. Integrative taxonomy and species delimitation in harvestmen: a revision of the western North American genus Sclerobunus (Opiliones: Laniatores: Travunioidea). PLoS One. 2014;9: e104982. doi: 10.1371/journal.pone.0104982 . PubMed DOI PMC

Satler JD, Carstens BC, Hedin M. Multilocus species delimitation in a complex of morphologically conserved trapdoor spiders (Mygalomorphae, Antrodiaetidae, Aliatypus). Syst Biol. 2013;62: 805–823. . PubMed

Staley JT. The bacterial species dilemma and the genomic–phylogenetic species concept. Philos Trans R Soc B Biol Sci. 2006;361: 1899–1909. doi: 10.1098/rstb.2006.1914 . PubMed DOI PMC

Gevers D, Dawyndt P, Vandamme P, Willems A, Vancanneyt M, Swings J, et al.. Stepping stones towards a new prokaryotic taxonomy. Philos Trans R Soc B Biol Sci. 2006;361: 1911–1916. doi: 10.1098/rstb.2006.1915 . PubMed DOI PMC

Gevers D, Cohan FM, Lawrence JG, Spratt BG, Coenye T, Feil EJ, et al.. Re-evaluating prokaryotic species. Nat Rev Microbiol. 2005;3: 733–739. doi: 10.1038/nrmicro1236 . PubMed DOI

Erwin PM, Thacker RW. Cryptic diversity of the symbiotic cyanobacterium Synechococcus spongiarum among sponge hosts. Mol Ecol. 2008;17: 2937–2947. doi: 10.1111/j.1365-294X.2008.03808.x . PubMed DOI

Kekkonen M, Hebert PDN. DNA barcode-based delineation of putative species: efficient start for taxonomic workflows. Mol Ecol Resour. 2014;14: 706–715. doi: 10.1111/1755-0998.12233 . PubMed DOI PMC

Tang CQ, Obertegger U, Fontaneto D, Barraclough TG. Sexual species are separated by larger genetic gaps than asexual species in rotifers. Evolution (N Y). 2014;68: 2901–2916. doi: 10.1111/evo.12483 . PubMed DOI PMC

Fontaneto D, Flot J-F, Tang CQ. Guidelines for DNA taxonomy, with a focus on the meiofauna. Mar Biodivers. 2015;45: 433–451. doi: 10.1007/s12526-015-0319-7 DOI

Eckert EM, Fontaneto D, Coci M, Callieri C. Does a barcoding gap exist in prokaryotes? Evidences from species delimitation in cyanobacteria. Life. 2015;5: 50–64. doi: 10.3390/life5010050 . PubMed DOI PMC

Meyer CP, Paulay G. DNA barcoding: error rates based on comprehensive sampling. PLoS Biol. 2005;3: e422. doi: 10.1371/journal.pbio.0030422 . PubMed DOI PMC

Pons J, Barraclough TG, Gomez-Zurita J, Cardoso A, Duran DP, Hazell S, et al.. Sequence-based species delimitation for the DNA taxonomy of undescribed insects. Syst Biol. 2006;55: 595–609. . PubMed

Fujisawa T, Barraclough TG. Delimiting species using single-locus data and the Generalized Mixed Yule Coalescent approach: a revised method and evaluation on simulated data sets. Syst Biol. 2013;62: 707–724. . PubMed PMC

Zhang J, Kapli P, Pavlidis P, Stamatakis A. A general species delimitation method with applications to phylogenetic placements. Bioinformatics. 2013;29: 2869–2876. . PubMed PMC

Lorén JG, Farfán M, Fusté MC. Species delimitation, phylogenetic relationships, and temporal divergence model in the genus Aeromonas. Front Microbiol. 2018;9: 770. doi: 10.3389/fmicb.2018.00770 . PubMed DOI PMC

Dvořák P, Jahodářová E, Casamatta DA, Hašler P, Poulíčková A. Difference without distinction? Gaps in cyanobacterial systematics; when more is just too much. Fottea. 2018;18: 130–136. doi: 10.5507/fot.2017.023 DOI

Talavera G, Dincă V, Vila R. Factors affecting species delimitations with the GMYC model: insights from a butterfly survey. Methods Ecol Evol. 2013;4: 1101–1110. doi: 10.1111/2041-210X.12107 DOI

Miralles A, Vences M. New metrics for comparison of taxonomies reveal striking discrepancies among species delimitation methods in Madascincus lizards. PLoS One. 2013;8: e68242. doi: 10.1371/journal.pone.0068242 . PubMed DOI PMC

Esselstyn JA, Evans BJ, Sedlock JL, Anwarali Khan FA, Heaney LR. Single-locus species delimitation: a test of the mixed Yule–coalescent model, with an empirical application to Philippine round-leaf bats. Proc R Soc B Biol Sci. 2012;279: 3678–3686. doi: 10.1098/rspb.2012.0705 . PubMed DOI PMC

Siegesmund MA, Johansen JR, Karsten UFT, Siegesmund MA, Johansen JR, Karsten U, et al.. Coleofasciculus gen. nov.(Cyanobacteria): Morphological and Molecular Criteria for Revision of the Genus microcoleus gomont 1. J Phycol. 2008;44: 1572–1585. doi: 10.1111/j.1529-8817.2008.00604.x . PubMed DOI