Anim. Syst. Evol. Divers. Vol. 36, No. 4: 396-399, October 2020 https://doi.org/10.5635/ASED.2020.36.4.044
Short communication DNA Barcoding of Antarctic Freshwater Copepod Boeckella poppei (Crustacea: Copepoda: Calanoida: Centropagidae) Inhabiting King George Island, South Shetland Islands, Antarctica
Seunghyun Kang1,*, Euna Jo1,2
1Korea Polar Research Institute, Incheon 21990, Korea 2College of Life Sciences and Biotechnology, Korea University, Seoul 02841, Korea
ABSTRACT
The Antarctic freshwater copepod, Boeckella poppei (Mrazek, 1901), has the widest range of distribution extending from southern South America to Antarctic continent, among all Boeckella species. Boeckella poppei is the only freshwater copepod known to be inhabiting the Antarctic continent. In present study, we analyzed the DNA barcodes of the mitochondrial cytochrome c oxidase subunit I (COI) gene of B. poppei from King George Island, Antarctica. The intraspecific genetic distances varied from 0% to 13% and interspecific genetic distances ranged from 11% to 14%. The overlap of DNA barcode gap suggests careful threshold-based delimitation of species boundaries. Keywords: Boeckella poppei, Antarctica, DNA barcode, COI, copepod
INTRODUCTION sphere, the DNA barcode information of the species has been reported from two geographical regions: Argentina (Cabo Boeckella de Guerne & Richard, 1889, is a freshwater cala- Virgenes and Tierra del Fuego) (Adamowicz et al., 2007), noid copepod belonging to the family Centropagidae, which and Signy Island (Maturana et al., 2020). Here, we report the is mostly restricted to the Southern Hemisphere (Bayly, DNA barcodes of B. poppei from King George Island and 1992). A total of 14 species of Boeckalla have been reported verify the usefulness of DNA barcode for this species com- from the southern latitudes of South (Bayly et al., 2003; Ada- pared with other populations and closely related species. The mowicz et al., 2007; Walter and Boxshall, 2018; Maturana et specimens of B. poppei were collected from Barton Peninsu- al., 2019). la (62°14′24″S, 58°44′36″W) of King George Island, South Boeckella poppei (Mrázek, 1901) is the only freshwater Shetland Islands (Fig. 1) in January 2020. Genomic DNA was copepod known to be distributed in Antarctica and extending extracted from the whole body using a QIAamp DNA Micro widely from southern South America to Antarctica (Heywood, Kit (Qiagen, Hilden, Germany) according to the manufac- 1977; Bayly and Burton, 1993; Menu-Marque et al., 2000; turer’s instructions. The Boeckella specific primers (Matur- Bayly et al., 2003; Gibson and Bayly, 2007; Nedbalová et al., ana et al., 2019) were used under the following polymerase 2009; Maturana et al., 2018, 2019). chain reaction conditions: initial denaturation at 94°C for 1 A 650 base pair (bp) section of the mitochondrial cyto- min; 10 cycles of 94°C for 1 min, 40°C for 90 s and 72°C chrome c oxidase subunit I (COI) DNA barcode has been for 1 min; followed by 30 cycles of 94°C for 1 min, 46°C used to distinguish species belonging to the subphylum Crus- for 90 s and 72°C for 90 s; and a final extension at 72°C for tacea (Costa et al., 2007) including copepods (Machida and 10 min. We have retrieved GenBank sequences of B. poppei Tsuda, 2010; Blanco-Bercial et al., 2011, 2014; Cepeda et al., from Signy Island (MN87317–MN087400) and Argentina 2012; Aarbakke et al., 2014; Sharma et al., 2014). Despite (DQ356568-DQ356570, DQ356576) and Boeckella brasil the widespread distribution of B. poppei in Southern Hemi- iensis (DQ356544–DQ356546) as an outgroup. The sequence
This is an Open Access article distributed under the terms of the Creative *To whom correspondence should be addressed Commons Attribution Non-Commercial License (http://creativecommons.org/ Tel: 82-32-760-5578, Fax: 82-32-760-5575 licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, E-mail: [email protected] and reproduction in any medium, provided the original work is properly cited. eISSN 2234-8190 Copyright The Korean Society of Systematic Zoology DNA Barcode of Antarctic Freshwater Copepod 10 0.133 9 0.012 0.126 8 0.123 0.121 0.112 7 0.002 0.123 0.122 0.113 6 0.008 0.009 0.124 0.122 0.118
5 Fig. 1. Location of Boeckella poppei (Mrázek, 1901) field sam- 0.004 0.005 0.005 0.123 0.122 0.116 pling site (62°14′24″S, 58°44′36″W) at Barton Peninsula, King George Island (Google Earth V, 2020). This map from Google, populations and outgroup species Maxar Technologies. 4 0.005 0.002 0.003 0.009 0.123 0.121 0.113
Table 2. Within group mean distance based on Kimura 2-pa- 3 rameter of Boeckella poppei populations and outgroup species 0.003 0.007 0.006 0.007 0.003 0.123 0.121 0.119 Boeckella poppei Mean Species name Location distance 2 0.003 0.007 0.001 0.006 0.007 0.003 0.123 0.122 0.118 Boeckella poppei King George Barton 0 Island Signy Island Changing 0.001 1 0.003 0.003 0.003 0.003 0.002 0.003 0.005 0.126 0.123 0.110 Heywood 0.002 Light 0.003 1 2 5 4 3 8 7 6 9 Pumphouse 0.004 10 11 No. COI DNA barcodes between Sombre 0.005 8 7 3 1 3 N 20 12 14 14 14 15 Tranquil 0.003 Twisted 0.001 Argentina Cabo Vigenes - Tierra del Fuego 0.006 Boeckella brasiliensis Argentina 0.021 Changing Heywood Barton Sombre Pumphouse Light Tierra del Fuego Tierra
Cabo Vigenes Twisted Tranquil
alignment, genetic distance calculation using the Kimura 2-parameter (K2P) model and neighbor-joining (NJ) phyloge- netic tree construction based on K2P genetic distance and the bootstrap with 1000 iteration have been performed by using MEGA X (ver. 10.1.8) (Kumar et al., 2018). Signy Island Signy King George Island Argentina Argentina Location
RESULTS AND DISCUSSION
The length of three COI sequences obtained was 403 bp and these new sequences were registered to GenBank (accession No. MT590771-MT590773 and MW008651–MW008667). Boeckella poppei Boeckella brasiliensis Pairwise genetic distances based on Kimura 2-parameter of 2-parameter genetic distances based on Kimura Table 1. Pairwise individuals. of N, number Species name
Anim. Syst. Evol. Divers. 36(4), 396-399 397 Seunghyun Kang, Euna Jo
Fig. 2. Neighbor joining tree based on Kimura 2-parameter distances of COI DNA barcodes Boeckella poppei individuals and out- group species.
Within the species, the pairwise genetic distances of B. pop CONFLICTS OF INTEREST pei based on K2P model ranged from 0% to 13% and in- ter-specific genetic distances ranged from 11% to 14% (Table No potential conflict of interest relevant to this article was 1). Within population mean distance based on K2P model reported. showed the lowest genetic distance value observed from Bar- ton population (King George Island) (Table 2) and the Barton populations showed low level of pairwise genetic distance ACKNOWLEDGMENTS with Signy Island populations ranged from 2% to 5% even though 700 km geographic distance apart. The NJ phyloge- This work was supported by Post-Polar Genomics Project: netic tree successfully delimitated B. poppei from the out- Functional genomic study for securing of polar useful genes group species (Fig. 2). The previous molecular systematics (PE20040) funded by Korea Polar Research Institute. of Centropagidae in Argentina based on COI gene showed that the average K2P genetic distance of the genus Boeckella varied from 10.9% to 25% (Adamowicz et al., 2007). Conse- REFERENCES quently, we observed an overlap between intraspecific and in- terspecific variations in B. poppei. Usually, the DNA barcode Aarbakke ONS, Bucklin A, Halsband C, Norrbin, F, 2014. Com- overlap does not affect the identification of unknown species parative phylogeography and demographic history of five in a thoroughly sampled tree but does have an impact when sibling species of Pseudocalanus (Copepoda: Calanoida) in the sampled group is incomplete (Meyer and Paulay, 2005). the North Atlantic Ocean. Journal of Experimental Marine In conclusion, the results suggest a threshold-based delimita- Biology and Ecology, 461:479-488. https://doi.org/10.1016/ tion of species boundary is not promising for identification of j.jembe.2014.10.006 B. poppei and closely related species, suggesting the need for Adamowicz SJ, Menu-Marque S, Hebert PDN, Purvis A, 2007. careful consideration and large sample sizes. Molecular systematics and patterns of morphological evo- lution in the Centropagidae (Copepoda: Calanoida) of Ar- gentina. Biological Journal of the Linnean Society, 90:279- 292. https://doi.org/10.1111/j.1095-8312.2007.00723.x ORCID Bayly IAE, 1992. Fusion of the genera Boeckella and Pseu doboeckella (Copepoda) and revision of their species from Seunghyun Kang: https://orcid.org/0000-0003-0115-9526 South America and sub-Antarctic islands. Revista Chilena de Euna Jo: https://orcid.org/0000-0003-2666-6743 Historia Natural, 65:17-63.
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