The Atlantidae (shelled heteropods) is a family of microscopic aragonite shelled holoplanktonic gastropods with a wide biogeographical distribution in tropical, sub-tropical and temperate waters. The aragonite shell and surface ocean habitat of the atlantids makes them particularly susceptible to ocean acidification and ocean warming, and atlantids are likely to be useful indicators of these changes. However, we still lack fundamental information on their taxonomy and biogeography, which is essential for monitoring the effects of a changing ocean. Integrated morphological and molecular approaches to taxonomy have been employed to improve the assessment of species boundaries, which give a more accurate picture of species distributions. Here a new species of atlantid heteropod is described based on shell morphology, DNA barcoding of the Cytochrome Oxidase I gene, and biogeography. All specimens of Atlanta ariejansseni sp. n. were collected from the Southern Subtropical Convergence Zone of the Atlantic and Indo-Pacific oceans suggesting that this species has a very narrow latitudinal distribution (37–48°S). Atlanta ariejansseni sp. n. was found to be relatively abundant (up to 2.3 specimens per 1000 m³ water) within this narrow latitudinal range, implying that this species has adapted to the specific conditions of the Southern Subtropical Convergence Zone and has a high tolerance to the varying ocean parameters in this region.

Atlantidae , biogeography, DNA barcoding, shelled heteropod, southern subtropical convergence zone

The Southern Ocean Sub-Tropical Front (STF) is the boundary between the colder, fresher Sub-Antarctic Zone (SAZ) and the warmer, more saline subtropical waters to the north (Orsi et al. 1995). The Southern Subtropical Convergence Zone (SSTC) is a narrow region along the STF with highly variable physical parameters experiencing strong currents and large gradients of salinity and temperature (Longhurst 1998, Graham and Boer 2013). The STF acts as a dispersal barrier for many zooplankton taxa, resulting in changes in genetic population structure and biomass across this front (Labat et al. 2001, Hiral et al. 2015, Burridge et al. in review a, b). This region is also at a high risk from ocean changes, particularly ocean acidification, because of the high solubility of CO₂ in cold water (Roberts et al. 2014).

The shelled atlantid heteropods are likely to be particularly susceptible to ocean acidification. Although, to date, there have been no studies into the effects of ocean changes upon atlantids, we can expect that they will react in a similar way to the shelled pteropods (Thecosomata). While not closely related, atlantids share many of the characteristic features that make shelled pteropods vulnerable to ocean acidification. These include living in the upper layers of the ocean, one of the areas most affected, and producing a very small (up to ~10 mm), thin shell of aragonite, which is particularly vulnerable to dissolution in waters undersaturated with carbonate (Fabry et al. 2008). In pteropods, synergistic effects of decreasing carbonate saturation and increasing temperature has been shown to reduce the ability to produce aragonite shells (e.g. Lischka and Riebesell 2012). These effects have already been recorded in natural populations living at high latitudes (Bednaršek et al. 2012), which are predicted to be affected first (Steinacher et al. 2009). However, improvements in taxonomy are extremely important to understanding the effects of these changes on holoplanktonic gastropods. Roberts et al. (2014) found that different forms of the pteropod species Limacina helicina (Phipps, 1774), living in the same area of the Southern Ocean, showed opposing trends in shell weight over a long-term study. This demonstrates the importance of assessing species boundaries in order to fully understand the effects of a changing ocean.

Here an integrated morphological and molecular approach is used to present a new species of atlantid heteropod, Atlanta ariejansseni, that is restricted to a narrow transitional zone of only 11° of latitude within the SSTC, but has a circumpolar longitudinal range. In common with other sub-polar planktonic gastropod species, A. ariejansseni reaches relatively high abundances compared to other atlantids and is the dominant atlantid species living in this area. Most atlantid species are thought to be restricted to warmer tropical and sub-tropical waters, with only one other species, Atlanta californiensis Seapy & Richter, 1993, showing a preference for cold water regions in the California Current. Atlanta ariejansseni is the only atlantid species specific to sub-polar waters and that appears to be tolerant of such a variable environment.

All specimens examined and included in this study were recorded within the SSTC, between 37°S and 48°S (Fig. 1). A total of 184 specimens of A. ariejansseni were examined from a number of sources (Table 1). From the Atlantic Ocean, 164 specimens for combined molecular and morphological analysis were collected during the Atlantic Meridional Transects AMT20 and AMT24 (Burridge et al. in review a). On both cruises, specimens were caught using a WP2 bongo net with an aperture diameter of 0.71 m and a mesh of 200 μm. Specimens from AMT24 were fixed and preserved in 96% ethanol and stored at -20 °C prior to DNA barcoding. Specimens from AMT20 were fixed and stored in 96% ethanol and stored at room temperature. Storage at room temperature is not optimal for the preservation of DNA; therefore, specimens from AMT20 were not used for DNA barcoding. From the Pacific Ocean, two further specimens, collected by Erica Goetze during the DRFT cruise of the RV Revelle in 2001, were used for molecular analysis (Table 1). Finally, 18 Indo-Pacific specimens were examined from sediment trap samples, collected from south of Tasmania between 1997–2006 by the Antarctic Climate and Ecosystems Cooperative Research Centre (Bray et al. 2000, Roberts et al. 2011). Upon removal from the sediment traps, specimens were washed in buffered peroxide to remove organic matter and dried.

Figure 1. 

The biogeography of A. ariejansseni based on known specimens. Dashed lines show the latitudinal limits of distribution at 37°S and 48°S.

Two published records of atlantids are also available for this region and both are considered here to include misidentified specimens of A. ariejansseni sp. n. Howard et al. (2011) recorded 14 specimens of Atlanta gaudichaudi Gray, 1850 in net hauls and a sediment trap positioned south of Tasmania. However, specimens from the same sediment traps (Roberts et al. 2011) that were re-examined for this study were also originally misidentified as A. gaudichaudi. A single image of a specimen caught by Howard et al. (2011) is morphologically consistent with A. ariejansseni, but is too small to identify with certainty.

Pilkington (1970) described a single species of atlantid, provisionally identified as Atlanta helicinoidea Gray, 1850, off-shore of Taiaroa Head, New Zealand. Pilkington (1970) found it difficult to identify specimens to species level, noting that the morphology did not agree perfectly with any of the atlantid species that had already been described. The detailed descriptions and figures presented by Pilkington (1970) unquestionably resemble the shell morphology of A. ariejansseni. Moreover, descriptions of the juvenile stages made by Pilkington (1970) match the juvenile specimens that were examined for this study. Therefore, the Atlanta specimens described by Pilkington (1970) are considered to be A. ariejansseni.

DNA barcoding

A total of 17 undamaged adult (N = 9) and juvenile (N = 8) specimens of A. ariejansseni were selected from samples collected during AMT24 and DRFT research cruises. DNA barcoding was also carried out for the morphologically similar species Atlanta selvagensis de Vera & Seapy, 2006 from the Atlantic Ocean. Five specimens of adult (N = 2) and juvenile (N = 3) A. selvagensis were selected from AMT24 sites (St. 5, 34.75°N, 26.62°W; St. 6, 31.30°N, 27.73°W and St. 14, 3.8°N, 25.78°W). All specimens were imaged prior to analysis using a Zeiss automated z-stage light microscope. DNA was extracted from whole specimens, using the NucleoMag 96 Tissue kit by Macherey-Nagel on a Thermo Scientific KingFisher Flex magnetic bead extraction robot, with a final elution volume of 75 μl. A standard Cytochrome Oxidase I (COI) barcoding fragment (Hebert et al. 2003) was amplified using primers jgLCO1490 and jgHCO2198 (Geller et al. 2013). Primers were tailed with M13F and M13R for sequencing (Messing 1983). PCR reactions contained 17.75 μl mQ, 2.5 μl 10x PCR buffer CL, 0.5 μl 25mM MgCl₂, 0.5 μl 100mM BSA, 1.0 μl 10 mM of each primer, 0.5 μl 2.5 mM dNTPs and 0.25 μl 5U Qiagen Taq, with 1.0 μl of template DNA, which was diluted 10 or 100 times for some samples. PCR was performed using an initial denaturation step of 180 s at 94 °C, followed by 40 cycles of 15 s at 94 °C, 30 s at 50 °C and 40 s at 72 °C, and finishing with a final extension of 300 s at 72 °C and pause at 12 °C. Sequencing was carried out by Macrogen, Europe.

All sequences were aligned and edited using the ClustalW algorithm in MEGA 6 (Tamura et al. 2013) and submitted to GenBank (Fig. 2, Table 2). Previously published COI sequences from GenBank (Jennings et al. 2010, Wall-Palmer et al. in press), identified as Atlanta inclinata Gray, 1850, Oxygyrus inflatus Benson, 1835, Firoloida desmarestia Lesueur, 1817, Pterotrachea hippocampus Philippi, 1836, Pterotrachea coronata Forsskål in Niebuhr, 1775 and Protatlanta souleyeti (Smith, 1888), were added to represent the families and genera most closely related to A. ariejansseni. Based on these data, a maximum-likelihood tree was constructed in MEGA6 using nucleotide sequences in a General Time Reversible model with gamma distribution and invariant sites (GTR+G+I) and 1000 bootstraps. Kimura-2-parameter (K2P) genetic distances were calculated between and within species belonging to the family Atlantidae using MEGA 6 (Tamura et al. 2013).

Figure 2. 

Maximum-likelihood tree showing the relationship of A. ariejansseni to different species of Atlanta, different Atlantidae genera, and different Pterotracheoidea families, based on Cytochrome Oxidase I DNA sequences. Branch lengths are proportional to the amount of inferred change, indicated by the scale bar. Only bootstrap support (1000 replicates) above 70% are displayed. GenBank sequence numbers are presented in Table 2. Sequences from Jennings et al. 2010 begin with FJ.

Table 2.

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XLSX

Original specimen codes and GenBank accession numbers for all specimens included in the phylogenetic analysis (Fig. 2).

Species	Specimen code or reference	GenBank accession number
Atlanta ariejansseni	Aari_AMT24_26_01	KX343177
	Aari_AMT24_26_02	KX343178
	Aari_AMT24_27_01	KX343179
	Aari_AMT24_27_02	KX343180
	Aari_AMT24_27_03	KX343181
	Aari_AMT24_27_04	KX343182
	Aari_AMT24_27_05	KX343183
	Aari_AMT24_28_01	KX343184
	Aari_AMT24_29_01	KX343185
	Aari_AMT24_29_02	KX343186
	Aari_AMT24_29_03	KX343187
	Aari_AMT24_29_04	KX343188
	Aari_AMT24_29_05	KX343189
	Aari_AMT24_29_06	KX343190
	Aari_AMT24_29_07	KX343191
	Aari_DRFT_14_01	KX343192
	Aari_DRFT_14_02	KX343193
Atlanta selvagensis	Asel_AMT24_05_03	KX343194
	Asel_AMT24_06_01	KX343195
	Asel_AMT24_06_02	KX343196
	Asel_AMT24_06_04	KX343197
	Asel_AMT24_14_02	KX343198
Atlanta gaudichaudi	Jennings et al. 2010	FJ876837
		FJ876839
Oxygyrus inflatus		FJ876848.1
		FJ876849.1
Protatlanta souleyeti	Wall-Palmer et al. in press	KU841501
		KU841495
		KU841506
		KU841502
		KU841497
		KU841494
		KU841496
		KU841493
Pterotrachea coronata	Jennings et al. 2010	FJ876852.1
		FJ876853.1
Pterotrachea hippocampus		FJ876854.1
		FJ876855.1
Firoloida desmarestia		FJ876850.1
		FJ876851.1

Combined molecular, morphological, and biogeographical information has allowed the introduction of a new species of the genus Atlanta that can be easily identified by means of its shell ornamentation using light microscopy. Atlanta ariejansseni is the only atlantid species that has been found living at high latitudes, restricted to a narrow circumpolar region. It is, therefore, an extremely important species in the current race to understand the effects of a changing ocean. It can be assumed that this species is able to tolerate a variable environment, which suggests that it may also be able to adapt to a changing ocean. This resilience and adaptability may be demonstrated by the successful rearing of veliger A. ariejansseni through to adults under laboratory conditions by Pilkinton (1970), which has never since been accomplished with other atlantid species.

Large sampling efforts have been made for holoplanktonic gastropods in the Southern Ocean; however, A. ariejansseni has never been recognised as an undescribed species in these studies. This is undoubtedly due to our incomplete understanding of atlantid taxonomy, particularly for the Atlantic Ocean. We hope that this study will increase awareness of A. ariejansseni and encourage others to record this circumpolar species when observed to build up a more complete biogeography. It is only with more biogeographical and ecological data that we will be able to determine the ecology and effects of a changing ocean upon this species.

We are grateful to Donna Roberts (University of Tasmania) for providing specimens from sediment traps from off-shore of Tasmania. The Australian Antarctic Division supports this ongoing sediment trap program (AAS #1156). We would like to thank Elaine Fileman and Rachel Harmer (Plymouth Marine Laboratory) for providing specimens from AMT20. We are grateful to Aline Nieman, Kevin Beentjes and Frank Stokvis (Naturalis Biodiversity Center) for help with DNA barcoding of specimens and Erica Goetze and Rachel Harmer for plankton collection on cruises AMT20, AMT24 and DRFT. We would like to acknowledge the Plymouth Electron Microscopy Centre and Glenn Harper for help with SEM imaging, the scientists and crew who took part in cruises AMT20, AMT24 and DRFT, and the Atlantic Meridional Transect (AMT) programme. This study is a contribution to the international IMBER project and was supported by the UK Natural Environment Research Council National Capability funding to Plymouth Marine Laboratory and the National Oceanography Centre, Southampton. This is contribution number 302 of the AMT programme. We acknowledge Diamond Light Source for time on Beamline/Lab I13-2 under Proposal MT12300-1 and Christophe Rau and Andrew Bodey for help with x-ray tomography. We are extremely grateful to María Moreno-Alcántara and Nathalie Yonow for reviewing our manuscript and for their constructive comments. DW-P was funded by the Leverhulme Trust (RPG-2013-363, 2014-2017, PA Christopher Smart, Plymouth University, Co-A Richard Kirby, Marine Biological Association, Plymouth) and a Martin-Fellowship from the Naturalis Biodiversity Center, Leiden (2015).