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  • Research article
  • Open Access

Phylogenetic relationships of cone snails endemic to Cabo Verde based on mitochondrial genomes

BMC Evolutionary BiologyBMC series – open, inclusive and trusted201717:231

https://doi.org/10.1186/s12862-017-1069-x

  • Received: 15 June 2017
  • Accepted: 6 November 2017
  • Published:

Abstract

Background

Due to their great species and ecological diversity as well as their capacity to produce hundreds of different toxins, cone snails are of interest to evolutionary biologists, pharmacologists and amateur naturalists alike. Taxonomic identification of cone snails still relies mostly on the shape, color, and banding patterns of the shell. However, these phenotypic traits are prone to homoplasy. Therefore, the consistent use of genetic data for species delimitation and phylogenetic inference in this apparently hyperdiverse group is largely wanting. Here, we reconstruct the phylogeny of the cones endemic to Cabo Verde archipelago, a well-known radiation of the group, using mitochondrial (mt) genomes.

Results

The reconstructed phylogeny grouped the analyzed species into two main clades, one including Kalloconus from West Africa sister to Trovaoconus from Cabo Verde and the other with a paraphyletic Lautoconus due to the sister group relationship of Africonus from Cabo Verde and Lautoconus ventricosus from Mediterranean Sea and neighboring Atlantic Ocean to the exclusion of Lautoconus endemic to Senegal (plus Lautoconus guanche from Mauritania, Morocco, and Canary Islands). Within Trovaoconus, up to three main lineages could be distinguished. The clade of Africonus included four main lineages (named I to IV), each further subdivided into two monophyletic groups. The reconstructed phylogeny allowed inferring the evolution of the radula in the studied lineages as well as biogeographic patterns. The number of cone species endemic to Cabo Verde was revised under the light of sequence divergence data and the inferred phylogenetic relationships.

Conclusions

The sequence divergence between continental members of the genus Kalloconus and island endemics ascribed to the genus Trovaoconus is low, prompting for synonymization of the latter. The genus Lautoconus is paraphyletic. Lautoconus ventricosus is the closest living sister group of genus Africonus. Diversification of Africonus was in allopatry due to the direct development nature of their larvae and mainly triggered by eustatic sea level changes during the Miocene-Pliocene. Our study confirms the diversity of cone endemic to Cabo Verde but significantly reduces the number of valid species. Applying a sequence divergence threshold, the number of valid species within the sampled Africonus is reduced to half.

Keywords

  • Mitochondrial genomes
  • Africonus
  • Trovaoconus
  • Kalloconus

Background

The cone snails (Conidae, Gastropoda) endemic to the archipelago of Cabo Verde in West Africa represent one of the few textbook examples of a well-documented insular species radiation involving marine organisms [13]. Cone snails, which are found in tropical and subtropical marine waters throughout the world, show a hotspot of species diversity in the Cabo Verde archipelago with up to 95 endemic species (roughly 10% of cone species diversity worldwide) narrowly confined to about 4000 km2 [4]. As in other parts of the world, cone snails endemic to Cabo Verde constitute a key component of the intertidal and subtidal ecosystems associated to rocky shores, coral reefs, and sandy bottoms. All cones endemic to Cabo Verde feed on marine annelid worms [1] and use a sophisticated venom apparatus (including a venom gland that produces conotoxins and a specialized harpoon-like radular tooth) to capture their preys [5]. Another interesting biological feature common to all these endemic species is that they have direct development. Their larvae lack a pelagic stage, and thus show a considerably reduced dispersal capacity [1]. Survival rate is higher for this type of larvae since they are less likely to be eaten by predators and are not dependent on plankton for feeding (i.e, non-planktotrophic).

The origin and evolutionary history of cones endemic to Cabo Verde has been the subject of several recent phylogenetic studies [1, 2, 6, 7]. Molecular phylogenies demonstrated that two different ancestors reached the archipelago independently and subsequently diversified following recurrent biogeographic patterns [1, 2, 7]. The existence of two clades led to the classification of cone species endemic to Cabo Verde into two genera, Africonus and Trovaoconus [8]. The question of which species are the closest living sister groups to Africonus and Trovaoconus remains open [1, 2]. According to a previous study, the ancestor of Africonus colonized the archipelago in the Miocene, about 16.5 million years ago (mya; [1]), and spread to all islands (except Fogo, the youngest, with steep slopes in the coast and ongoing volcanic activity). Most (95%) of the currently described species endemic to Cabo Verde belong to Africonus, and are normally referred to as restricted to a single island and in some cases even to single bays within an island [3]. The ancestor of Trovaoconus arrived at Cabo Verde archipelago in the Pliocene, about 4.6 mya, and diversified only in four islands (Sal, Boa Vista, Maio, and possibly Santiago), which are the closest to the continent [1]. These cones are significantly larger in size than those belonging to Africonus and show wider distributions extending in some cases to more than one island. It has been hypothesized that diversification within each genus was in allopatry and followed recurrent eustatic sea level changes during the Neogene that intermittently connected and disconnected the islands [1, 7]. However, sea level fluctuations alone do not fully explain the extraordinary diversity of cones in Cape Verde since nearby archipelagos in the Macaronesia biogeographic region such as the Canary Islands subjected to similar trends since the Miocene do not have endemic cone species [6]. A larger distance to the mainland, which enhances isolation and restricts gene flow combined with a higher mean sea surface temperature and the presence of more suitable habitats may have promoted a significant increase in diversification rates in the Cabo Verde archipelago [6].

The rate of description of new cone species endemic to Cabo Verde has accelerated more than expected during the last years (Fig. 1). After the early descriptions in the eighteenth and nineteenth centuries based on samples brought to Europe by naturalists [9], the main contribution to the cataloguing of cone species endemic to Cabo Verde was due to the work of Emilio Rolán [10], who drew attention to this singular radiation. Hence, around year 2000, there were about 50 species recognized [11] and remarkably this number has almost doubled in the last 2-3 years [1219]. However, it is important to note that many of the recent species diagnoses in cones are mainly based on the shape, color, and banding patterns of the shell. These phenotypic characters are highly variable at the population level and prone to local adaptation and convergence, making species assignment problematic and sometimes, misleading [7]. In many cases, distinguishing whether different shell morphotypes of cone snails represent valid species or ecotypes of the same species is challenging [20]. Therefore, determination of genetic variation and inference of phylogenetic relationships based on DNA sequence data are timely as part of a multidisciplinary approach [21] to identify and delimit species and to understand evolutionary processes underlying diversification within cones, in general, and within those endemic to Cabo Verde, in particular.
Fig. 1
Fig. 1

Number (blue) and accumulated number (red) of cone species described for the Cabo Verde archipelago per year

Here, we used nearly complete mitochondrial (mt) genomes, which have proven to successfully reconstruct robust phylogenies of Conidae [22] and of particular groups such as the cones endemic to Senegal [23]. In this study, we sequenced the nearly complete mt genomes of 88 individuals representing different populations and species of Africonus and Trovaoconus endemic to Cabo Verde. We aimed to: (1) reconstruct a highly resolved phylogeny of cones endemic to Cabo Verde; (2) determine the closest living sister groups of Africonus and Trovaoconus; (3) date major cladogenetic events and analyze biogeographical patterns; (4) study radular tooth evolution within the two genera; and (5) provide a first genetic hypothesis of species delimitation in the radiation of Cabo Verde endemic cones.

Results

Sequencing, assembly, and genome organization

The nucleotide sequences of the near-complete mt genomes of 75 specimens of Africonus, 13 specimens of Trovaoconus, and one specimen of Lautoconus ventricosus were determined (Table 1). These mt genomes lacked the trnF gene, the control region, and the start of the cox3 gene because the corresponding fragment was not PCR amplified. The number of reads, mean coverage, and length of each mt genome are provided in Table 1. The mt genomes of Africonus boavistensis and Africonus denizi received the minimum (42,021) and maximum (906,765) number of reads, respectively. The same samples received the minimum (412×) and maximum (8,885×) mean coverage, respectively (Table 1). All sequenced mt genomes encode for 13 protein-coding, 2 rRNA and 21 tRNA genes (but note that the trnF gene could not be determined; see above). They all share the same genome organization: the major strand encodes all genes, except those forming the cluster MYCWQGE (trnM, trnY, trnC, trnW, trnQ, trnG, trnE) and the trnT gene.

Table 1

Mitochondrial (mt) genomes analyzed in this study

New mt genomes

ID CV

Initial species identification

Location

Coordinates

Coverage

Length (bp)

GenBank Acc. No

Voucher DNA (MNCN/ADN)

Voucher shell (MNCN 15.05/)

New species proposeda

n° reads

mean depth

1020

Africonus antoniaensis

Água Doce, Boa Vista, Cabo Verde

16°12'29"N, 22°44'7"W

151104

1476.8

15332

MF491587

95072

79889

0885

Africonus antoniomonteiroi

Pedra Lume, Sal, Cabo Verde

16°45'44"N, 22°53'2"W

232069

2273.4

15328

MF491578

95063

79794

0927

Africonus bernardinoi

Pedra Lume, Sal, Cabo Verde

16°45'44"N, 22°53'2"W

59799

583.3

15328

MF491582

95067

79835

Africonus cuneolus

0520

Africonus boavistensis

Baía do Ervatão (North), Boa Vista, Cabo Verde

16°12'3"N, 22°54'43"W

42021

412.8

15217

MF491563

95045

80413

1135

Africonus cabraloi

Estancinha, Boa Vista, Cabo Verde

16°13'12"N, 22°55'9"W

74446

730.4

15329

MF491598

95083

80004

Africonus crotchii

0895

Africonus cagarralensis

Pedra Lume, Sal, Cabo Verde

16°45'44"N, 22°53'2"W

161290

1367.2

15320

MF491579

95064

79804

Africonus longilineus

0173

Africonus calhetae

Praia da Soca, Maio, Cabo Verde

15°15'8"N, 23°13'4"W

55433

544.7

15242

MF491534

95016

78798

0920

Africonus cf. anthonyi

Ilhéus do Chano, Sal, Cabo Verde

16°41'37"N, 22°52'47"W

172336

1678.6

15315

MF491581

95066

79828

Africonus cuneolus

0162

Africonus cf. claudiae

Praia da Soca, Maio, Cabo Verde

15°15'8"N, 23°13'4"W

87407

858.6

15326

MF491533

95015

78787

Africonus calhetae

0465

Africonus cf. delanoyae

Ponta Antónia, Boa Vista, Cabo Verde

16°13'24"N, 22°46'59"W

382817

3736.1

15335

MF491559

95041

80409

Africonus fuscoflavus

0207

Africonus cf. galeao

Ponta Pipa, Maio, Cabo Verde

15°19'30"N, 23° 9'48"W

81447

797.3

15325

MF491536

95018

78832

Africonus galeao

0135

Africonus cf. gonsaloi

Praia Gonçalo, Maio, Cabo Verde

15°16'13"N, 23°6'15"W

148032

1455.5

15250

MF491529

95011

78760

Africonus gonsaloi

0380

Africonus cf. miguelfiaderoi

Jorrita, Baía da Gata, Boa Vista, Cabo Verde

16°12'9"N, 22°42'22"W

358342

3507.9

15328

MF491548

95030

80398

Africonus vulcanus

1400

Africonus cf. miruchae

Calhau, São Vicente, Cabo Verde

16°51'7"N, 24°51'59"W

523002

5104.9

15321

MF491601

95088

78562

Africonus sp. nov. 1

0223

Africonus claudiae

Ponta Pipa, Maio, Cabo Verde

15°19'30"N, 23° 9'48"W

148508

1434.5

15337

MF491537

95019

78848

Africonus galeao

0303

Africonus condei

Baía Grande, Derrubado, Boa Vista, Cabo Verde

16°13'31"N, 22°47'17"W

253863

2472.2

15248

MF491542

95024

80392

Africonus crotchii

0045

Africonus crioulus

Praia Santana, Maio, Cabo Verde

15°18'13"N, 23°11'49"W

255019

2502

15247

MF491521

95003

78670

Africonus maioensis

1075

Africonus crotchii

Morro de Areia, Boa Vista, Cabo Verde

16°5'24"N, 22°57'7"W

332385

3237.6

15329

MF491591

95076

79944

0803

Africonus cuneolus

Calheta Funda, Sal, Cabo Verde

16°39'6"N, 22°56'53"W

184181

1791.6

15329

MF491569

95053

79712

0936

Africonus cuneolus

Santa Maria, Sal, Cabo Verde

16°35'38"N, 22°53'36"W

80472

787.4

15328

MF491583

95068

79844

1420

Africonus curralensis

Praia de Palmo Tostão, Santa Luzia, Cabo Verde

16°45'19"N, 24°45'24"W

857123

8358.6

15329

MF491602

95089

78581

1017

Africonus damioi

Água Doce, Boa Vista, Cabo Verde

16°12'29"N, 22°44'7"W

76477

745.5

15326

MF491586

95071

79886

Africonus roeckeli

0405

Africonus damottai

Baía da Gata (center), Boa Vista, Cabo Verde

16°11'50"N, 22°42'32"W

315488

2914.3

15358

MF491551

95033

80401

1428

Africonus decoratus

Curral, Santa Luzia, Cabo Verde

16°46'23"N, 24°47'13"W

566822

5540.2

15326

MF491603

95090

78589

0370

Africonus delanoyae

Jorrita, Baía da Gata, Boa Vista, Cabo Verde

16°12'9"N, 22°42'22"W

158489

1543.7

15323

MF491547

95029

80397

1471

Africonus denizi

Praia Grande, São Vicente, Cabo Verde

16°51'40"N, 24°52'30"W

906765

8885.2

15326

MF491605

95092

78621

0315

Africonus derrubado

Baía Grande, Derrubado, Boa Vista, Cabo Verde

16°13'31"N, 22°47'17"W

214173

2089.2

15243

MF491543

95025

80393

Africonus damottai

0565

Africonus diminutus

Ilhéu de Sal Rei, Boa Vista, Cabo Verde

16°9'50"N, 22°55'31"W

840424

8204.1

15330

MF491566

95049

80416

1025

Africonus docensis

Água Doce, Boa Vista, Cabo Verde

16°12'29"N, 22°44'7"W

47313

464.8

15329

MF491588

95073

79894

Africonus crotchii

0385

Africonus evorai

Zebraca (near Ilhéu do Galeão), Boa Vista, Cabo Verde

16°12'6"N, 22°42'40"W

226416

2218

15243

MF491549

95031

80399

Africonus crotchii

0070

Africonus fantasmalis

Porto Cais, Maio, Cabo Verde

15°19'15"N, 23°11'10"W

97527

954.7

15330

MF491524

95006

78695

Africonus fuscoflavus

0835

Africonus felitae

Rabo de Junco, Sal, Cabo Verde

16°41'44"N, 22°58'35"W

344190

3343.9

15404

MF491573

95057

79744

1437

Africonus fernandesi

Porto Novo, Santo Antão, Cabo Verde

17°1'4"N, 25°3'22"W

742414

7244.2

15324

MF491604

95091

78598

0332

Africonus fiadeiroi

Derrubado (bay West), Boa Vista, Cabo Verde

16°13'22"N, 22°47'41"W

205910

2016.5

15243

MF491545

95027

80395

Africonus crotchii

0855

Africonus fontonae

Baía da Fontona, Sal, Cabo Verde

16°44'22"N, 22°58'46"W

156259

1523.9

15328

MF491575

95059

79764

Africonus cuneolus

0945

Africonus fontonae

Regona, Sal, Cabo Verde

16°48'5"N, 22°59'33"W

56310

549.8

15327

MF491584

95069

79853

Africonus regonae

0450

Africonus fuscoflavus

Derrubado (bay East), Boa Vista, Cabo Verde

16°13'331"N, 22°47'3"W

151904

1478.6

15331

MF491557

95039

80407

0052

Africonus galeao

Navio Quebrado, Terras Salgadas, Maio, Cabo Verde

15°18'54"N, 23°11'2"W

117940

1139.2

15326

MF491522

95004

78677

0134

Africonus gonsaloi

Praia Gonçalo, Maio, Cabo Verde

15°16'13"N, 23°6'15"W

188174

1835.9

15339

MF491528

95010

78759

1390

Africonus grahami

Calhau, São Vicente, Cabo Verde

16°51'7"N, 24°51'59"W

464704

4536.1

15325

MF491599

95086

78552

0140

Africonus irregularis

Porto Cais (North), Maio, Cabo Verde

15°19'45"N, 23°10'57"W

202254

1937.9

15321

MF491530

95012

78765

Africonus maioensis

0317

Africonus irregularis

Baía Grande, Derrubado, Boa Vista, Cabo Verde

16°13'31"N, 22°47'17"W

170523

1668.2

15331

MF491544

95026

80394

Africonus maioensis

0392

Africonus irregularis

Baía da Gata, Boa Vista, Cabo Verde

16°11'50"N, 22°42'32"W

252126

2454.3

15324

MF491550

95032

80400

Africonus crotchii

1084

Africonus irregularis

Morro de Areia, Boa Vista, Cabo Verde

16°5'24"N, 22°57'7"W

125264

1225.1

15330

MF491593

95078

79953

Africonus crotchii

1128

Africonus irregularis

Estancinha, Ponta do Sol, Boa Vista, Cabo Verde

16°13'12"N, 22°55'9"W

469101

4597.1

15313

MF491597

95082

79997

Africonus crotchii

0225

Africonus isabelarum

Ponta do Pau Seco, Maio, Cabo Verde

15°15'26"N, 23°13'16"W

247567

2431.7

15244

MF491538

95020

78850

0085

Africonus josephinae

Lage Branca, Maio, Cabo Verde

15°18'32"N, 23°8'17"W

224495

2204.6

15239

MF491525

95007

78710

Africonus sp. nov. 2

0555

Africonus josephinae

Ilhéu de Sal Rei, Boa Vista, Cabo Verde

16°9'50"N, 22°55'31"W

169723

1611.8

15330

MF491565

95048

80415

0830

Africonus longilineus

Serra Negra, Sal, Cabo Verde

16°38'17"N, 22°53'56"W

148726

1453

15316

MF491572

95056

79739

0847

Africonus longilineus

Rabo de Junco, Sal, Cabo Verde

16°41'44"N, 22°58'35"W

308057

3000.9

15333

MF491574

95058

79756

Africonus miruchae

0410

Africonus luquei

Praia Canto, Boa Vista, Cabo Verde

16°11'10"N, 22°42'28"W

83198

815.1

15244

MF491552

95034

80402

Africonus delanoyae

0064

Africonus maioensis

Porto Cais, Maio, Cabo Verde

15°19'15"N, 23°11'10"W

143797

1402.9

15327

MF491523

95005

78689

0510

Africonus marckeppensi

Ervatao Norte, Boa Vista, Cabo Verde

16°12'3"N, 22°54'43"W

254030

2480.3

15330

MF491562

95044

80412

Africonus josephinae

0102

Africonus marcocastellazzii

Lage Branca, Maio, Cabo Verde

15°18'32"N, 23°8'17"W

80254

783.1

15326

MF491527

95009

78727

Africonus maioensis

0870

Africonus melissae

Baía da Parda, Sal, Cabo Verde

16°45'7"N, 22°53'56"W

195531

1907.9

15328

MF491577

95061

79779

Africonus longilineus

0455

Africonus messiasi

Derrubado (bay East), Boa Vista, Cabo Verde

16°13'33"N, 22°47'3"W

250171

2447.7

15260

MF491558

95040

80408

Africonus fuscoflavus

0426

Africonus miguelfiaderoi

Praia Canto, Boa Vista, Cabo Verde

16°11'10"N, 22°42'28"W

130100

1270.5

15328

MF491554

95036

80404

Africonus vulcanus

0905

Africonus mordeirae

Baía do Roucamento, Sal, Cabo Verde

16°41'20"N, 22°56'24"W

99337

971.4

15241

MF491580

95065

79814

Africonus cuneolus

1091

Africonus morroensis

Morro de Areia, Boa Vista, Cabo Verde

16°5'24"N, 22°57'7"W

172039

1515.1

15337

MF491594

95079

79960

Africonus diminutus

1395

Africonus navarroi

Calhau, São Vicente, Cabo Verde

16°51'7"N, 24°51'59"W

665250

6509.2

15331

MF491600

95087

78557

0250

Africonus nelsontiagoi

Tarrafal, Santiago, Cabo Verde

15°16'50"N, 23°45'15"W

173873

1687.2

15339

MF491541

95023

78875

Africonus verdensis

0820

Africonus pseudocuneolus

Serra Negra, Sal, Cabo Verde

16°38'17"N, 22°53'56"W

131838

1288.7

15337

MF491571

95055

79729

Africonus cuneolus

0036

Africonus raulsilvai

Praia da Soca, Maio, Cabo Verde

15°15'8"N, 23°13'4"W

345872

1699.6

15534

MF491520

95002

78661

0865

Africonus regonae

Baía da Fontona, Sal, Cabo Verde

16°44'22"N, 22°58'46"W

246041

2411

15328

MF491576

95060

79774

0950

Africonus regonae

Regona, Sal, Cabo Verde

16°48'5"N, 22°59'33"W

88673

864.1

15337

MF491585

95070

79858

0586

Africonus roeckeli

Praia Canto, Boa Vista, Cabo Verde

16°11'10"N, 22°42'28"W

141600

1385.7

15320

MF491567

95050

80417

0549

Africonus salreiensis

Ilhéu de Sal Rei, Boa Vista, Cabo Verde

16°9'50"N, 22°55'31"W

349070

3402.9

15331

MF491564

95047

80414

Africonus crotchii

0810

Africonus serranegrae

Serra Negra, Sal, Cabo Verde

16°38'17"N, 22°53'56"W

182124

1777.6

15335

MF491570

95054

79719

Africonus cuneolus

1078

Africonus silviae

Morro de Areia, Boa Vista, Cabo Verde

16°5'24"N, 22°57'7"W

293568

2876.1

15336

MF491592

95077

79947

Africonus fuscoflavus

0445

Africonus swinneni

Porto Ferreira, Boa Vista, Cabo Verde

16°7'45"N, 22°40'17"W

221672

2169.1

15244

MF491556

95038

80406

Africonus delanoyae

1125

Africonus teodorae

Estancinha, Ponta do Sol, Boa Vista, Cabo Verde

16°13'12"N, 22°55'9"W

326654

3185.5

15334

MF491596

95081

79994

Africonus crotchii

1035

Africonus umbelinae

Espingueira, Boa Vista, Cabo Verde

16°12'55"N, 22°47'49"W

110526

1085.5

15333

MF491589

95074

79904

Africonus damottai

0240

Africonus verdensis

Tarrafal, Santiago, Cabo Verde

15°16'50"N, 23°45'15"W

53400

519.8

15339

MF491540

95022

78865

0435

Africonus vulcanus

Porto Ferreira, Boa Vista, Cabo Verde

16°7'45"N, 22°40'17"W

284766

2787.4

15242

MF491555

95037

80405

1110

Africonus zinhoi

Curral Velho, Boa Vista, Cabo Verde

15°58'4"N, 22°47'42"W

388828

3785.5

15331

MF491595

95080

79979

Africonus maioensis

7036

Trovaoconus atlanticoselvagem

Baixo João Valente, Cabo Verde

15°44′27″N, 23°5′26″W

95264

933.8

15352

MF491606

7036

Kalloconus trochulus

0616

Trovaoconus cf. ateralbus

Serra Negra, Sal, Cabo Verde

16°38'17"N, 22°53'56"W

87550

853.7

15344

MF491568

95052

79664

Kalloconus sp. nov. 1

0010

Trovaoconus pseudonivifer

Ponta do Pau Seco, Maio, Cabo Verde

15°15'26"N, 23°13'16"W

56486

555.5

15351

MF491519

95000

78635

Kalloconus trochulus

0094

Trovaoconus pseudonivifer

Lage Branca, Maio, Cabo Verde

15°18'32"N, 23°8'17"W

454415

4429.8

15351

MF491526

95008

78719

Kalloconus trochulus

0154

Trovaoconus pseudonivifer

Porto Cais (north), Maio, Cabo Verde

15°19'45"N, 23°10'57"W

199736

1954.2

15352

MF491532

95014

78779

Kalloconus trochulus

0420

Trovaoconus pseudonivifer

Praia Canto, Boa Vista, Cabo Verde

16°11'10"N, 22°42'28"W

111182

1085

15347

MF491553

95035

80403

Kalloconus pseudonivifer

0500

Trovaoconus trochulus

Baía do Ervatão (North), Boa Vista, Cabo Verde

16°12'3"N, 22°54'43"W

223915

2177.2

15351

MF491561

95043

80411

Kalloconus trochulus

0149

Trovaoconus venulatus

Lage Branca, Maio, Cabo Verde

15°18'32"N, 23°8'17"W

105732

1014.9

15276

MF491531

95013

78774

Kalloconus venulatus

0187

Trovaoconus venulatus

Praia Real, Maio, Cabo Verde

15°19'45"N, 23°10'40"W

144651

1415

15330

MF491535

95017

78812

Kalloconus venulatus

0234

Trovaoconus venulatus

Ponta do Pau Seco, Maio, Cabo Verde

15°15'26"N, 23°13'17"W

67867

661.5

15320

MF491539

95021

78859

Kalloconus venulatus

0347

Trovaoconus venulatus

Derrubado (bay West), Boa Vista, Cabo Verde

16°13'22"N, 22°47'41"W

48636

475.9

15326

MF491546

95028

80396

Kalloconus venulatus

0475

Trovaoconus venulatus

Ponta Antónia, Boa Vista, Cabo Verde

16°13'24"N, 22°46'59"W

409041

3966.1

15340

MF491560

95042

80410

Kalloconus venulatus

1038

Trovaoconus venulatus

Praia Canto, Boa Vista, Cabo Verde

16°11'10"N, 22°42'28"W

143644

2403.8

15336

MF491590

95075

79907

Kalloconus venulatus

IB001

Lautoconus ventricosus

Estani des Peix, Formentera, Balearic Islands, Spain

38°43'49"N, 1°24'42"E

86290

842.5

15341

MF491607

95094

80426

Lautoconus sp. nov 1

GenBank mt genomes

ID

Species

Location

Coordinates

Reference

Length (bp)

GenBank Acc. No

Voucher (MNCN/ADN)

Voucher shell (MNCN 15.05/)

New species proposeda

6990

Africonus borgesi

Porto Ferreira, Boa Vista, Cabo Verde

16°7'45"N, 22°40'170"W

Cunha et al., (2009)

15536

NC_013243

6990

0025

Africonus infinitus

Ponta do Pau Seco, Maio, Cabo Verde

15°15'26"N, 23°13'17"W

Abalde et al., (in prep.)

15522

KY864967

95001

78650

0875

Africonus miruchae

Terrinha Fina, Palhona, Sal, Cabo Verde

16°49'12"N, 22°59'12"W

Abalde et al., (in prep.)

15336

KY864971

95062

79784

0534

Trovaoconus pseudonivifer

Estancinha, Ponta do Sol, Boa Vista, Cabo Verde

16°13'12"N, 22°55'9"W

Abalde et al., (in prep.)

15351

KY864969

95046

80418

Kalloconus trochulus

0550

Trovaoconus venulatus

Ilhéu de Sal Rei, Boa Vista, Cabo Verde

16°9'56"N, 22°55'23"W

Uribe et al., (2017)

15524

KX263250

86741

80419

Kalloconus venulatus

0601

Trovaoconus ateralbus

Calheta Funda, Sal, Cabo Verde

16°39'6"N, 22°56'53"W

Abalde et al., (in prep.)

15327

KY864970

95051

79649

Kalloconus ateralbus

1375

Kalloconus cf. byssinus

North Senegal

unknown

Abalde et al., (in prep.)

15348

KY864973

95085

78536

Kalloconus pulcher

1253

Kalloconus pulcher

Les Almadies, Dakar, Senegal

14°44'40"N, 17°31'442"W

Abalde et al., (in prep.)

15332

KY864972

95084

78414

Kalloconus pulcher

1343

Lautoconus belairensis

Terrou-Bi. Dakar, Senegal

14°40'29"N, 17°28'12"W

Abalde et al., (2017)

15321

KY801849

91293

78504

Gen. nov. belairensis

1338

Lautoconus bruguieresi

Île de Gorée, Dakar, Senegal

14°40'16"N, 17°23'58"W

Abalde et al., (2017)

15340

KY801851

91291

78499

Gen. nov. bruguieresi

1296

Lautoconus cloveri

Ndayane, Senegal

14°33'45"N, 17°7'34"W

Abalde et al., (2017)

15323

KY801859

91283

78457

Gen. nov. cloveri

CG13

Lautoconus guanche

Lanzarote, Canary Islands, Spain

28°57'16"N, 13°34'22"W

Abalde et al., (2017)

15506

KY801847

91295

Gen. nov. guanche

1266

Lautoconus hybridus

NGor, Dakar, Senegal

14°45'67"N, 17°30'36.33"W

Abalde et al., (2017)

15507

KY801863

91279

78427

Gen. nov. hybridus

1278

Lautoconus mercator

NGor, Dakar, Senegal

14°45'6"N, 17°30'36"W

Abalde et al., (2017)

15329

KY801862

91280

78439

Gen. nov. mercator

CV13

Lautoconus ventricosus

Ria Formosa, Faro, Portugal

36°58'0"N, 7°53'2"W

Uribe et al., (2017)

15534

KX263251

86742

CVERM1

Chelyconus ermineus

Praia Gonçalo, Maio, Cabo Verde

15°16'13"N, 23°6'15"W

Abalde et al., (in prep.)

15365

KY864977

95095

78876

ahyphen indicates that original species name is maintained and considered valid

Phylogenetic relationships and sequence divergences between clades

Phylogenetic relationships of cones endemic to Cabo Verde were reconstructed based on the nucleotide sequences of the concatenated 13 mt protein-coding and two rRNA genes using probabilistic methods and Chelyconus ermineus as outgroup. The final matrix was 13,572 positions in length. According to the AIC, the best partition scheme for the protein-coding genes was the one combining all these genes but analyzing each codon position separately. The best substitution model for each of the three codon positions was GTR + I + G. For the rRNA genes, the best scheme had both genes combined under the GTR + I + G model. Both, ML (−lnL= 75,600.18) and BI (−lnL = 76,002.71 for run 1; −lnL = 76,288.44 for run 2) arrived at almost identical topology (Figs. 2 and 3). Most nodes received high statistical support and differences in topology were restricted exclusively to three relatively shallow nodes that had low support in ML and were unresolved in BI. Two of these nodes involved almost identical sequences and corresponded to Africonus bernardinoi/ Africonus pseudocuneolus and Africonus teodorae/ Africonus fiadeiroi, respectively. The third unresolved node corresponded to a trichotomy involving Africonus felitae, Africonus regonae and Africonus longilineus/ Africonus cagarralensis/ Africonus melissae.
Fig. 2
Fig. 2

Closest sister groups of cone snails endemic to Cabo Verde based on mt genomes (concatenated protein coding plus rRNA genes analyzed at the nucleotide level). The reconstructed ML tree using C. ermineus as outgroup is shown. Number of specimen, initial species assignment, and a ventral picture of the shell are provided. Numbers at nodes are statistical support values for ML (bootstrap proportions)/BI (posterior probabilities). Scale bar indicates substitutions/site. Two major clades are recovered: the first one includes Kalloconus from Western Africa (purple) and Trovaoconus from Cabo Verde (green) whereas the second one includes Lautoconus from the Mediterranean Sea (and neighboring Atlantic Ocean1; orange), Lautoconus from Senegal (and L. guanche from Canary Islands2; red), and Africonus (blue) from Cabo Verde. Lineages within Africonus are expanded in Fig. 3. The “robust” type of radular tooth is shown as the ancestral character state for the second clade (scale bar equals 0.1 mm; see diversity of radular teeth of the first clade in Additional file 1). The two transitions from planktonic to non-planktonic larvae are indicated by a sac of eggs

Fig. 3
Fig. 3

Phylogeny of Africonus based on mt genomes (concatenated protein coding plus rRNA genes analyzed at the nucleotide level). Number of specimen, initial species assignment, island, and a ventral picture of the shell are provided. Numbers at nodes are statistical support values for ML (bootstrap proportions)/BI (posterior probabilities). Hyphen indicates a bootstrap value below 50%. Scale bar indicates substitutions/site. Four major lineages (I-IV) are recovered and indicated with different colors. All Africonus have the “robust” type of radular tooth except when indicated (scale bar equals 0.1 mm). 1All taxa endemic to Maio except A. irregularis endemic to Boa Vista. 2All taxa endemic to Boa Vista except A. fantamalis endemic to Maio

The reconstructed phylogeny (Fig. 2) grouped the analyzed species into two main clades, one including Kalloconus from mainland West Africa sister to Trovaoconus from Cabo Verde and the other having paraphyletic Lautoconus due to the sister group relationship of Africonus from Cabo Verde and Lautoconus ventricosus from Mediterranean Sea and neighboring Atlantic Ocean to the exclusion of Lautoconus endemic to Senegal (plus Lautoconus guanche from Mauritania, Morocco, and Canary Islands). Within Trovaoconus, up to three main lineages could be distinguished (Fig. 2). The first one included two specimens from Sal initially identified as Trovaoconus ateralbus, which were sister to a clade including one lineage with specimens from Maio and Boa Vista identified as Trovaoconus venulatus and another lineage having mostly specimens of Trovaoconus pseudonivifer from Maio and Boa Vista but also one specimen of Trovaoconus trochulus from Boa Vista and one of Trovaoconus atlanticoselvagem from Baixo João Valente (Fig. 2).

The clade of Africonus from Cabo Verde included four main lineages (named I to IV), each further subdivided into two monophyletic groups (Figs. 2 and 3). Lineage I was the sister group of the remaining Africonus and its two lineages had each species from Maio sister to species from Boa Vista (Fig. 3a). Lineage II included species from Santiago and Maio sister to species endemic to the westernmost islands (Santo Antão, São Vicente and Santa Luzia). These latter species could be grouped into three main lineages, one containing species endemic to São Vicente, another containing species distributed both in Santa Luzia and São Vicente, and the third one including species from the three islands (Fig. 3a). Lineage III included species from Maio sister to species from Boa Vista (Fig. 3a). Lineage IV contained specimens representing most of the described species of Africonus. One monophyletic group included species endemic to Sal whereas the other clade included Africonus isabelarum from Maio as sister to four lineages, two containing exclusively species from Boa Vista, one having species from Maio sister to Africonus irregularis from Boa Vista, and one having species from Boa Vista and Africonus fantasmalis from Maio (Fig. 3b).

Pairwise uncorrected sequence divergences were estimated based on the alignment including the nucleotide sequences of the 13 mt protein-coding and two rRNA genes. Pairwise uncorrected sequence divergences between C. ermineus and ingroup taxa averaged 18%. The average pairwise uncorrected sequence divergence between the two main ingroup clades (genera Kalloconus + Trovaoconus versus genera Lautoconus + Africonus) was 16%. Pairwise uncorrected sequence divergences between Lautoconus endemic to Senegal (plus L. guanche) and L. ventricosus plus Africonus averaged 11%. The average pairwise uncorrected sequence divergence between the sister groups L. ventricosus and Africonus was 10% whereas between Kalloconus and Trovaoconus, it was 5%. The pairwise uncorrected sequence divergences between the four main lineages within Africonus averaged 6%. The corresponding values for the pairwise divergences between the two major clades defined within each of the lineages I-IV were 4%, 6%, 3%, and 3%, respectively. Pairwise uncorrected sequence divergence comparisons between sister species level were distributed into two different ranges, one closer to 1% (0.5-1.5%) and the other closer to 0% (0-0.5%). The latter divergences were particularly abundant among sister species comparisons within Maio, Boa Vista and Sal. Several mt genomes of different species were almost identical (<0.05%) in sequence including (1) Africonus delanoyae and Africonus luquei, (2) Africonus fuscoflavus, Africonus cf. delanoyae, and Africonus messiasi, (3) Africonus irregularis (#1128) and Africonus cabraloi, (4) Africonus verdensis and Africonus nelsontiagoi, and (5) Africonus gonsaloi and Africonus cf. gonsaloi.

Evolution of radular types

The different lineages within Africonus exhibit distinct radular types (Fig. 3). Most lineages and species showed the “robust” type, which is of medium relative size, with a short, pointed barb and a basal spur (see Additional file 1). The anterior section of the tooth is equal or slightly shorter than the posterior section, and the blade covers most of the anterior section (80% – 85%). There are usually 19 to 30 denticles in the serration, arranged in one row (occasionally two). Several species within lineage IV (Africonus delanoyae, Africonus luquei, Africonus swinneni, Africonus fuscoflavus, Africonus messiasi, Africonus silviae and Africonus cf. delanoyae from Boa Vista island, and Africonus fantasmalis from Maio island) exhibited radular teeth of the “elongated” type, similar to the “robust” type but characterized by an anterior section which is longer than the posterior section, a blade covering 40 to 50% of the anterior section, and more numerous denticles in the serration (usually more than 30) often arranged in two rows. Several species (Africonus borgesi, Africonus josephinae and Africonus marckeppensi in lineage I, Africonus navarroi in lineage II), all species in lineage III, plus Africonus vulcanus, Africonus miguelfiadeiroi and Africonus cf. miguelfiadeiroi in lineage IV) displayed radular tooth of the “broad” type, which is characterized by a medium-sized (Shell Length/Tooth Length = 32-45) and very broad radular tooth (Shell Length/Anterior section Width = 7-12), with an anterior section which is shorter than the posterior section (Tooth Length/Anterior section Length = 2.1-2.9), a blade covering most of the anterior section, and with a variable number of denticles (8 to 30) in the serration arranged in two or more rows. The radular morphology of Africonus felitae may represent a special case with a small relative size (Shell Length/Tooth Length = 63-67), narrow (Tooth Length/Anterior section Width = 20-23), the anterior section shorter than the posterior section (Tooth Length/Anterior section Length = 2.2-2.4), and characterized by the total absence of denticles in the serration. The base of this tooth is relatively large and broad.

The species of Kalloconus and Trovaoconus exhibit essentially two kinds of radular morphologies (Additional file 1). The teeth in K. pulcher, and also in Trovaoconus trochulus, T. pseudonivifer and Trovaoconus atlanticoselvagem are narrow and elongated; the blade is moderately short being about one third to almost one-half the length of the anterior section of the tooth, which is distinctly longer than the posterior section of the tooth. There are many denticles (25 to 45 or more) in the long serration, arranged usually in multiple rows with a major row flanked by numerous smaller serrations. In the case of T. venulatus, Trovaoconus ateralbus, and Trovaoconus cf. ateralbus the teeth are broader, and the anterior and posterior sections are almost equal in length. There are 16 to 33 denticles in the serration, often coarse and hook-shaped in the middle portion, arranged initially in one row becoming two rows below.

Dating of major cladogenetic events

Major cladogenetic events within the reconstructed phylogeny were dated using an uncorrelated relaxed molecular clock model, which was calibrated using the age of Sal (28 mya; the oldest island of the archipelago) for the node separating Africonus from its sister group, L. ventricosus, and the age of the origins of São Vicente, Santo Antão, and Santa Luzia (7.5 mya) for the node splitting the lineage including the endemics to these islands from its sister group lineage including endemics to Maio and Santiago islands [24]. The first divergence event involving Kalloconus + Trovaoconus versus (paraphyletic) Lautoconus + Africonus was dated at 34 mya (Fig. 4; note that genera and species labels in the chronogram take into account proposed synonymizations, see Discussion). The divergence between the clade containing cones endemic to Senegal (+ L. guanche) and the clade including L. ventricosus plus Africonus was dated at 26 mya. The split between the latter two lineages was dated at 23 mya. The diversification of the crown group of Africonus into its four main lineages (I-IV) was estimated to have occurred between 9.4 - 6.9 mya (Fig. 4). The separation of Kalloconus and Trovaoconus was dated 9 mya and the diversification of the crown group of Trovaoconus was established at 4 mya (Fig. 4).
Fig. 4
Fig. 4

Chronogram based on mt genomes (concatenated protein coding plus rRNA genes analyzed at the nucleotide level) and using the fixed topology of the ML tree shown in Figs. 2 and 3. Lineages are colored as in Fig. 2. Synonymizations proposed (see text) are taken into account. A Bayesian uncorrelated relaxed lognormal clock with geographic-based calibration priors (denoted by asterisks) was used in BEAST. Horizontal bars represent 95% credible intervals for time estimates; dates are in millions of years. Geological periods are indicated. Brown and orange bands indicate main divergence periods around the Oligocene-Miocene and Miocene-Pliocene transitions, respectively. A red line indicates the threshold for species delimitation. The bathymetry of Cabo Verde archipelago is shown in an inset

Diversification rates through time

Variations in the diversification rates through time were estimated for Africonus and the hypothesis of a radiation during the evolutionary history of the clade was tested. The gamma-statistic, which measures departures from a constant rate of diversification, had values of 7.19 (p < 0.05) and 3.12 (p < 0.05) when considering the currently named (based on phenotypic traits) or only the here-proposed (considering genetic evidence) species for the genus, respectively. In both cases, the hypothesis of a radiation is accepted. According to the lineage through time plots (Fig. 5), the initial rate of increase in the number of species slowed down between six and one and a half million years ago regardless of the species delimitation hypothesis tested. Afterwards, the diversification rate accelerated considerably, and the increase in number of species either continued or abandoned a normal Yule process of speciation when considering the species delimitation hypothesis here proposed or the currently number of named species, respectively.
Fig. 5
Fig. 5

Logarithmic lineage through time (LTT) plots of described (a) and proposed valid (b) species. The red bar indicates the pure Yule process of speciation, the grey shadow shows 1000 simulated trees and the black lines represent 100 trees randomly chosen among the trees generated by BEAST

Discussion

Cone snails are marine gastropods well known to evolutionary biologists due to their extraordinary species and ecological diversity [25], but also to molecular biologists and pharmacologists due to their sophisticated venom cocktails [26], as well as to amateur naturalists due to their brightly colored and highly appreciated shells [27]. Therefore, they are the subject of intensive research across disciplines and additionally have received wide attention from the general public. There are more than 800 described species and this number increases steadily every year [28]. Thus far, species description and identification of cones heavily relies on shell form, color and banding patterns, which may show great variety at local scale leading to important levels of synonymy within the family Conidae [7, 20]. In this regard, species delimitation could greatly improve with the aid of robust molecular phylogenies, which could be used in addition as framework to uncover the evolutionary patterns and processes underlying the diversification of the group. While reconstructing a robust phylogeny for all described cone species worldwide is cumbersome and at present unrealistic within the framework of a single study, it is possible, however, to accomplish a proof-of-concept study in a particular region [23].

We have here reconstructed a molecular phylogeny of cones endemic to Cabo Verde and allied species in the Macaronesian region, continental West Africa, and the Mediterranean region. These cones are particularly interesting from an evolutionary perspective as they have radiated in an oceanic archipelago and constitute a natural experiment to gain insights onto the processes governing diversification and adaptation [29]. Phylogenetic analyses were based on nearly complete mt genomes (only missing the control region and neighboring sequences) and included 105 specimens comprising most of the cone species diversity of the analyzed regions. Probabilistic methods of phylogenetic inference arrived at a robust and highly resolved phylogeny (virtually all nodes received high statistical support, which in most cases was maximal). To our knowledge, this is the first wide application of mt genomes to the resolution of a phylogeny within mollusks (but see [30, 31] for comparable examples in fish and insects, respectively). Previous studies in gastropods were restricted in the number of taxa analyzed (e.g., [22] for the family Conidae) but here we were able to achieve a lineage representation of the reference group (in this case, Cabo Verde cones) only previously attained by studies using few concatenated partial gene sequences (see e.g., [25] for the family Conidae or [1] for the cones of Cabo Verde). Previous phylogenetic studies using complete mt genomes have demonstrated that the level of resolution of these molecular markers is compromised above the superfamily level due to saturation, base composition biases, and among-lineage rate heterogeneity [32, 33]. Here, we show that phylogenetic performance of mt genomes achieves best results when analyzing closely related genera (and their corresponding species). Moreover, results were particularly promising taking into account that Africonus diversity in Cabo Verde was originated through radiation processes, which normally lead to relatively short tree nodes (often difficult to disentangle).

A thorough sampling of closely-related outgroup taxa allowed us to tackle key questions on the origin of the cones endemic to Cabo Verde and on their closest living sister groups. As previously reported, there are two independent origins of Cabo Verde cones, leading to the genera Africonus and Trovaoconus, respectively [1]. The closest living sister group of Africonus is L. ventricosus from the Mediterranean Sea and neighboring Atlantic Ocean. Therefore, the origin of this clade is clearly Macaronesian/Mediterranean and these cones are only distantly related to the geographically closer cones endemic to Senegal. These latter cones were ascribed to the genus Lautoconus (as was the case of L. guanche from Canary islands, deeply nested within the clade of Senegal cones; [23]). However, the closest sister group relationship of Africonus and L. ventricosus requires formal description of a different genus for Senegal cones (plus L. guanche), which will be done elsewhere. We could not include any representative of cones endemic to Angola (genus Varioconus) but a recent phylogeny based on partial cox1 gene sequences recovered all these cones (including Varioconus jourdani from Saint Helena Island) as a monophyletic group sister to Senegal cones [34]. Alternatively, all previously mentioned genera could be merged into genus Lautoconus [35]. However, the relatively high levels of sequence divergence (using the sequence divergence of genus Chelyconus as reference) and the restricted (endemic) distribution of the clades fit better with the former taxonomic proposal. The closest living sister group of Trovaoconus is genus Kalloconus from West Africa. Therefore, the origin of this clade is clearly related to neighboring regions of the continent. Actually, the sequence divergence between Trovaoconus and Kalloconus is much lower than that estimated between Lautoconus and Africonus. This observation argues against maintaining the generic status of Trovaoconus, and supports the inclusion of their species within genus Kalloconus, as some authors have proposed [35]. Hence, Kalloconus would be a genus that is present throughout the coast of West Africa from Morocco to Angola as well as in Canary Islands and Cabo Verde.

Altogether, the two main clades in the reconstructed phylogeny show very distinct patterns of distribution. One clade includes a single genus with widespread distribution in Macaronesia and West Africa whereas the other clade, which occupies the same geographical regions, is divided into several valid genera (Africonus, Lautoconus, Varioconus, Gen. nov. for Senegal endemics). These distinct patterns could be explained partly taking into account differences in larval dispersal capabilities between the two clades [1]. According to the phylogeny, the ancestor of the Kalloconus clade was inferred to have planktotrophic larvae, capable of long dispersals whereas the ancestor of the other clade would have non-planktotrophic larvae, and thus a limited dispersal capability leading to restricted gene flow and higher rates of diversification [36, 37]. Interestingly, the ancestor of Kalloconus species endemic to Cabo Verde (former Trovaoconus species) lost planktotrophy, which is a common evolutionary pattern in insular species [38].

According to the reconstructed phylogeny, cones belonging to genus Africonus are divided into four main lineages (I-IV; with each further subdivided into two distinct clades). Species endemic to Maio and Boa Vista are found in all four lineages whereas species endemic to Sal form a clade within lineage IV, species from the westernmost islands (Santo Antão, São Vicente and Santa Luzia) form a clade within lineage II, and the single species from Santiago is recovered within lineage II. Unfortunately, we could not sample specimens of Africonus furnae from Brava and Africonus kersteni from São Nicolau, and cannot determine whether they could be ascribed to any of the above-mentioned four lineages or form their own independent lineages. The single origin of cones endemic to Sal, Santiago, and westernmost islands could be explained by the deep slopes separating these islands whereas the multiple origins of the cones found in Maio and Boa Vista could be associated to the relatively shallow seamount (Baixo João Valente) connecting both islands [1]. These differences in bathymetry in connection with past eustatic sea level changes could be determinant in preventing or promoting dispersal in Africonus species, whose larvae are all non-planktotrophic.

Diversification events among main lineages were concentrated in three major periods. The first one, around the Oligocene-Miocene boundary (23 mya), includes the divergence of cones endemic to Senegal (and Angola) from their sister clade, and the posterior separation within this sister clade of cones endemic to Cabo Verde and those endemic to the Mediterranean Sea and neighboring Atlantic Ocean. During Oligocene-Miocene transition, there was a global cooling event [39, 40], the ice sheet of Antarctica greatly expanded, and a sea level drop of ~50 m occurred [41]. The second period corresponds to a sustained global cooling in the Late Miocene starting 12 mya [42] that produced an eustatic sea level drop between −10 and −30 m from 6.26 to 5.50 Mya [43] and culminated with the Messinian Salinity crisis and the desiccation of the Mediterranean Sea at the end of the Miocene from 5.96 to 5.33 Mya [44]. During this period, the divergence of the main lineages within Africonus (I-IV), the cones endemic to Senegal, and Kalloconus occurred. Finally, a burst of speciation events is inferred during the Pleistocene when another cooling period characterized by extreme climate oscillations and drastic eustatic sea level changes concurring with glacial-interglacial periods [45]. Global cooling has been recently proposed to be a driver of diversification of marine species [46] in agreement with our results. The reconstructed phylogeny, the chronogram, and the current geographical distribution of the species altogether support that allopatry is the main mode of speciation for cone snails with non-planktotrophic larvae, as previously suggested [1]. The complex geology of the island of Boa Vista with several eruptions at >16, 15-12.5, and 9.5-4.5 mya [47], involving different parts of the island may have also contributed to creating additional niches along the coast and could explain that this island harbors the highest number of endemic cones.

The reconstructed phylogeny also allows inferring the evolution of the radula in the studied lineages [8, 48]. All analyzed ingroup taxa are vermivorous [8]. Studies documenting potential specialization of the vermivore radular type to prey on specific worm species are scarce and restricted thus far to cone species preying on amphinomids [49]. Here, we show that most Africonus species show a “robust” radular type, which is shared also with L. ventricosus and a lineage of Senegal cones represented by Lautoconus cloveri and Lautoconus bruguieresi in the phylogeny [23]. Therefore, the common ancestor of cones endemic to Senegal (plus L. guanche), Africonus, and L. ventricosus was inferred to have a “robust” type radula. The “elongated” type of radular tooth, which was found in several species within lineage IV of Africonus, also appears in a lineage of Senegal cones that is represented by Lautoconus hybridus and L. guanche in the phylogeny [23]. The radular tooth of A. felitae resembles the “small” type observed in a lineage of Senegal cones represented by Lautoconus reticulatus and Lautoconus belairensis in the phylogeny [23]. The “broad” type of radular tooth that appeared independently in several lineages of Africonus has not been observed in any cone from Senegal. While shifts in radular type could be correlated with early cladogenesis in cones endemic to Senegal [23], the evolution of different types of radular tooth within Africonus was restricted to few specific cases. Thus, future studies are needed to determine whether in such cases there has been a dietary shift to prey on specific worms. The radula teeth identified in Kalloconus resemble the types “elongated” and “robust” observed within Lautoconus and Africonus, although are clearly distinct. This might indicate instances of convergence, and that only a discrete number of different main types of radula could be found in a given clade.

During the last few years, the number of new cone species described from Cabo Verde has increased at an astonishing rate (e.g., [12]). These new species are identified based on differences (often subtle) in shell shape and color, and their status needs to be contrasted with genetic data to uncover cases of local phenotypic variation within species due to either genetic polymorphism or phenotypic plasticity that may be producing overestimations of the number of species in the group [21]. In addition, genetic data could help identify cases of phenotypic convergence due to adaptation of genetically distinct populations (ecotypes) or species (sibling or cryptic) to similar environments [5052], also affecting the total number of valid species. Comparative analyses of pairwise uncorrected sequence divergences taking into account the reconstructed phylogeny showed that some described species shared almost identical mt genomes with levels of sequence divergence normally considered to be associated to genetic variation at the population level. Clades comprising these sets of closely related sequences indicate that an uncorrected sequence divergence threshold around 1% could be associated to the species status. This threshold lies well within the so-called grey zone of speciation between 0.5-2% [53]. Of course, these results need to be further confirmed with genomic nuclear data that discard potential events of incomplete lineage sorting and hybridization [54]. In addition, the present study could be further improved in the future by increasing the number of individuals analyzed per original species. Importantly, the comparative analyses on variation of diversification rates through time support the here proposed hypothesis of species delimitation as it concurs with a Yule process of speciation whereas the number of currently named species clearly exceed expectations and would imply an extraordinary recent acceleration of speciation rates.

Our study confirms the diversity of cone endemic to Cabo Verde but significantly reduces the number of valid species. Applying the threshold in a conservative manner (i.e., maintaining described species as valid in case of doubt due to closeness to the threshold) to cones endemic to Cabo Verde would reduce the number of valid species within the sampled Africonus from 65 to 32 (see Table 1 and Fig. 4). The proposed nomenclatural changes follow standard ICZN recommendations maintaining the most senior (oldest) name. Among the species not sampled, two correspond to São Nicolau and Brava islands, four of them are from the islands of São Vicente and Santa Luzía, and most likely represent valid species (Africonus bellulus, Africonus lugubris, Africonus saragasae, Africonus santaluziensis, A. kersteni and A. furnae) given the relative high sequence divergences found among species endemic to these islands. The 19 remaining ones were recently described, mostly from Boa Vista, and are expected to fall in most cases into some of the clades already discussed in the present work, and therefore may correspond to morphs of other described species. A direct consequence of synonymization is that some previously described species of rather restricted distribution are merged as populations into the new species, which considerably increase their range of distribution (Additional file 1). For instance, A. crotchii, which was reported as endemic from Southwest Boa Vista, would be now distributed also in the whole north half of the island. This increase in range of distribution of several species has important effects on their IUCN conservation status [3]. In the case of Kalloconus, some morphotypes attributed to Kalloconus pseudonivifer are now assigned to Kalloconus trochulus, and Kalloconus atlanticoselvagem is synonymized with K. trochulus. Our specimen of Kalloconus cf. byssinus is from North Senegal and has little sequence divergence compared to Kalloconus pulcher. In this case, it would be important to study K. byssinus from Mauritania or Morocco before considering synonymization. In the opposite direction, there are three clear instances of morphological convergence and thus, of the existence of cryptic species. Those are the cases of Africonus josephinae from Maio, Africonus cf. miruchae from São Vicente, and Kalloconus cf. ateralbus from Sal, which will be described as new species in due course.

Conclusions

We reconstructed a robust phylogeny based on mitochondrial genomes of cone snails endemic to Cabo Verde, which provides the necessary framework for future evolutionary studies focused on this radiation. The double origin of Cabo Verde endemic cones was supported. The ancestor of Africonus separated from L. ventricosus during the Oligocene-Miocene boundary (about 23 mya) and diversified into four main lineages (I to IV) in the Late Miocene (about 9.4-6.9 mya). The divergence of the ancestor of Kalloconus endemic to Cabo Verde from those inhabiting mainland occurred also in the Late Miocene whereas its diversification into three main lineages was dated in the Pliocene (4 mya). Main cladogenetic events within cones endemic to Cabo Verde coincide with global cooling periods, which were characterized by radical climate oscillations and eustatic sea level changes. Recurrent cycles of island connection/ disconnection likely favored speciation in allopatry in these cones, which lack a pelagic larval stage, and thus have limited dispersal capacity. Direct development evolved in the ancestor of Kalloconus endemic to Cabo Verde, likely associated to the colonization of the archipelago by a cone with a planktotrophic larval stage. However, in the case of Africonus, the ancestor that arrived to Cabo Verde was already non-planktotrophic as the corresponding independent evolutionary shift to direct development predated the separation of cones endemic to Senegal (and Canary Islands) from L. ventricosus plus Africonus. Radular types were modified during the diversification of Africonus from an ancestral “robust” type, although correlation with diet specializations await better knowledge of the specific worm species preyed by the different species of cones. Sequence divergence comparisons and reconstructed phylogenies supported the diversity of cone species endemic to Cabo Verde but significantly reduced its number, which was likely overestimated in the past due to important homoplasy in shell morphology, the, thus far, main discriminant character used for species description and identification.

Methods

Samples and DNA extraction

The complete list of specimens analyzed in this study corresponding to different populations and species of Africonus and Trovaoconus from Cabo Verde is shown in Table 1, as well as details on the respective sampling localities and museum vouchers. As outgroup taxa, we also sampled and analyzed one specimen of L. ventricosus from Formentera Island (Spain). Specimens were collected by snorkel at 1-3 m depth, or picked by hand at low tide. All samples were stored in 100% ethanol. The initial species identification (see corresponding column in Table 1) was based on comparison with type material (mostly deposited in the MNCN) or consulting the original publications. Total DNA was isolated from 5 to 10 mg of foot tissue following a standard phenol-chloroform extraction [55].

Radular tooth preparation

The radular sac was dissected from the main body and soft parts were digested in concentrated aqueous potassium hydroxide for 24 h. The resulting mixture was then placed in a petri dish and examined with a binocular microscope. The entire radula was removed with fine tweezers and rinsed with distilled water, then mounted on a slide using Aquatex (Merck, Germany) mounting medium, and observed under a compound microscope. Photographs were taken with a charge-coupled device (CCD) camera attached to the microscope. Terminology for radular morphology follows [8], with abbreviations following [48]. Names of radular types follow [23].

PCR amplification and sequencing

Near-complete (without the control region) mt genomes were amplified through a combination of standard and long PCRs using the primers and following the protocols of [22]. Standard-PCR products were sequenced using Sanger technology. Long-PCR products were subjected to next-generation sequencing. Briefly, PCR amplified fragments from the same mt genome were pooled together in equimolar concentrations. For each cone mt genome a separate indexed library was constructed using the NEXTERA XT DNA library prep kit (Illumina, San Diego, CA, USA). The average size of the Nextera libraries varied between 307 and 345 bp. Libraries were pooled and run in an Illumina MiSeq platform (v.2 chemistry; 2 × 150 paired-end) at Sistemas Genómicos (Valencia, Spain).

Genome assembly and annotation

The reads corresponding to each mt genome were sorted using the corresponding library indices, and read assembly was performed in the TRUFA webserver [56]. Briefly, adapters were removed using SeqPrep [57], quality of the reads was checked using FastQC v.0.10.1 [58], and raw sequences were trimmed and filtered out according to their quality scores using PRINSEQ v.0.20.3 [59]. Filtered reads were used for de novo assembly of each mt genome using default settings (minimum contig length: 200; sequence identity threshold: 0.95) of Trinity r2012-06-08 [60] in TRUFA, and only retaining contigs with a minimum length of 3 kb. These contigs were used as starting point to assemble the mt genomes using Geneious® 8.0.3. First, the (raw) reads with a minimum identity of 99% were mapped against the contigs to correct possible sequence errors. Then, successive mapping iterations using a 100% identity as threshold were performed to elongate the contigs.

The mt genomes were annotated with the option “Annotate from Database” in Geneious® 8.0.3, using published mt genomes of Conidae as references. Annotations of the 13mt protein-coding genes were refined manually identifying the corresponding open reading frames using the invertebrate mitochondrial code. The transfer RNA (tRNA) genes were further identified with tRNAscan-SE 1.21 [61], which infer cloverleaf secondary structures (with a few exceptions that were determined manually). The ribosomal RNA (rRNA) genes were identified by sequence comparison with other Conidae mt genomes [22], and assumed to extend to the boundaries of adjacent genes [62]. GenBank accession numbers of each mt genome are provided in Table 1.

Sequence alignment and phylogenetic analyses

The newly sequenced mt genomes were aligned with the mt genomes of A. borgesi, Africonus infinitus, Africonus miruchae, T. ateralbus, T. pseudonivifer, T. venulatus, and C. ermineus from Cabo Verde, L. hybridus, L. mercator, L. belairensis, L. cloveri, L. bruguieresi, K. pulcher, and K. cf. byssinus from Senegal, L. guanche from Canary Islands, and L. ventricosus from Portugal, which were downloaded from GenBank (Table 1). A sequence data set was constructed concatenating the nucleotide sequences of the 13 mt protein-coding and two rRNA genes. The deduced amino acid sequences of the 13 mt protein-coding genes were aligned separately and used to guide the alignment of the corresponding nucleotide sequences with Translator X [63]. Nucleotide sequences of the mt rRNA genes were aligned separately using MAFFT v7 [64] with default parameters. Ambiguously aligned positions were removed using Gblocks, v.0.91b [65] with the following settings: minimum sequence for flanking positions: 85%; maximum contiguous non-conserved positions: 8; minimum block length: 10; gaps in final blocks: no. Finally, the different single alignments were concatenated using Geneious® 8.0.3. Sequences where format converted for further analyses using the ALTER webserver [66]. The concatenated alignment is available at http://purl.org/phylo/treebase/phylows/study/TB2:S21557.

Phylogenetic relationships were inferred using maximum likelihood (ML, [67]) and Bayesian inference (BI, [68]). For ML, we used RAxML v8.1.16 [69] with the rapid hill-climbing algorithm and 10,000 bootstrap pseudoreplicates (BP). BI analyses were conducted with MrBayes v3.1.2 [70], running four simultaneous Markov chains for 10 million generation, sampling every 1000 generations, and discarding the first 25% generations as burn-in (as judged by plots of ML scores and low SD of split frequencies) to prevent sampling before reaching stationarity. Two independent Bayesian inference runs were performed to increase the chance of adequate mixing of the Markov chains and to increase the chance of detecting failure to converge, as determined using Tracer v1.6 [71]. The effective sample size (ESS) of all parameters was checked to be above 200. Node support was assessed based on Bayesian Posterior Probabilities (BPP). A node was considered highly supported with BP and BPP values above 70% and 0.95, respectively. The ML and BI phylogenetic trees are available at http://purl.org/phylo/treebase/phylows/study/TB2:S21557.

The best partition schemes and best-fit models of substitution for the data set were identified using PartitionFinder2 [72] with the Akaike information criterion [73]. For the protein-coding genes, the partitions tested were: all genes grouped; all genes separated (except atp6-atp8 and nad4-nad4L); and genes grouped by subunits (atp, cob, cox, and nad). In addition, these three partitions schemes were tested taking into account separately the three codon positions. The rRNA genes were tested with two different schemes, genes separated or combined.

Estimation of divergence times

The program BEAST v.1.8.0 [74] was used to perform a Bayesian estimation of divergence times. An uncorrelated relaxed molecular clock was used to infer branch lengths and nodal ages. The tree topology was fixed using the one recovered by the ML analysis. For the clock model, the lognormal relaxed-clock model was selected, which allows rates to vary among branches without any a priori assumption of autocorrelation between adjacent branches. For the tree prior, a Yule process of speciation was employed. Concatenated protein coding plus rRNA genes were analyzed at the nucleotide level. The partitions and models selected by PartitionFinder2 were applied (see results). The final Markov chain was run twice for 100 million generations, sampling every 10,000 generations, and the first 1000 trees were discarded as part of the burn-in process, according to the convergence of chains checked with Tracer v.1.5. [71]. The ESS of all parameters was above 200.

Despite the fact that there are many fossils of Conidae, it is difficult in many instances to be certain about species identifications given the important levels of homoplasy in shell shape [75]. Hence, although there are fossils attributed to L. ventricosus [76] and L. mercator [77], which could be applied to the reconstructed phylogeny, we opted to calibrate the clock using biogeographical events (i.e., the age of the islands of Cabo Verde). We run a preliminary analysis in which the posterior distribution of the estimated divergence times was obtained by specifying one calibration point as prior for the divergence time of the split between L. ventricosus and the genus Africonus. This genus is endemic to Cabo Verde, and we used the age of formation of the oldest island, Sal (28 Mya; [24]), as biogeographical calibration point. We applied a log-normal distribution as the prior model for the calibration and enforced the median divergence time to equal 25 (s.d. = 0.05, offset = 0.7). According to the results of the preliminary analysis, we found that only in the case of São Vicente, Santo Antão, and Santa Luzia, the early divergence of living cone endemic lineages followed the origin of the corresponding island, and therefore, we used a second calibration point corresponding to the origin of these islands about 7.5 mya [24]. We applied a log-normal distribution as the prior model for the calibration and enforced the median divergence time to equal 7.5 (s.d. = 0.03, offset = 0). The BEAST tree is available at http://purl.org/phylo/treebase/phylows/study/TB2:S21557.

Diversification rate through time

The chronogram was used to determine diversification rate through time of genus Africonus under alternative (phenotypic versus genetic) species delimitation hypotheses. A lineage through time (LTT) plot analysis was conducted using the APE 4.1 R package [78]. A random sample of 100 trees was selected and mapped over a simulation of 1000 trees following a Yule process of speciation (net diversification rate = 0.4). The phytools R package [79] was used to calculate the Gamma-Statistic [80].

Declarations

Acknowledgements

We are indebted to Cabo Verde biology students Paulo Vasconcelos and Stiven Pires, as well as to our colleagues Sara Rocha, David Posada and Julio Rozas for their valuable help during sampling in Cabo Verde. We thank Dr. Rui Freitas from the Universidade de Cabo Verde (UniCV) for his continuous support and for granting us access to the facilities of the UniCV in Mindelo, São Vicente. We also thank Dr. Iderlindo Silva dos Santos and Dra. Sonia Monteiro de Pina Araujo from the Direcçao Nacional do Medio Ambiente of the Ministério do Ambiente, Habitação e Ordenamento do Território (MAHOT) of the Republic of Cabo Verde for their help with collecting permits (Autorizações 07/2013, 26/2013, 01/2104, 04/2015, and 03/2016). The sample of L. ventricosus from Formentera Island was obtained by Paula C. Rodríguez Flores. We are grateful to Jesús Marco and Aida Palacios, who provided access to the supercomputer Altamira at the Institute of Physics of Cantabria (IFCA-CSIC), member of the Spanish Supercomputing Network, for performing assembling and phylogenetic analyses.

Funding

This work was supported by the Spanish Ministry of Science and Innovation (CGL2013-45211-C2-2-P and CGL2016-75255-C2-1-P (AEI/FEDER, UE)) to RZ; BES-2011-051469 to JEU; BES-2014-069575 to SA; Doctorado Nacional-567, Colciencias-Universidad Nacional de Colombia to AME.

Availability of data and materials

Sequence data is available in GenBank.

Authors’ contributions

MJT, CMLA, and RZ collected the material. MJT prepared the radula. SA, JEU, and AME generated the molecular data. SA analyzed the data. RZ wrote the first draft of the manuscript and all authors contributed to writing the final version. All authors read and approved the final manuscript.

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

Publisher’s Note

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Authors’ Affiliations

(1)
Museo Nacional de Ciencias Naturales (MNCN-CSIC), José Gutiérrez Abascal 2, 28006 Madrid, Spain
(2)
Departamento CMIM y Q. Inorgánica-INBIO, Facultad de Ciencias, Universidad de Cádiz, 11510 Puerto Real, Cádiz, Spain
(3)
Centre of Marine Sciences (CCMAR), Universidade do Algarve, Campus de Gambelas, 8005 - 139 Faro, Portugal

References

  1. Cunha RL, Castilho R, Rüber L, Zardoya R. Patterns of cladogenesis in the venomous marine gastropod genus Conus from the Cape Verde Islands. Syst Biol. 2005;54(4):634–50.PubMedView ArticleGoogle Scholar
  2. Duda TF, Rolán E. Explosive radiation of Cape Verde Conus, a marine species flock. Mol Ecol. 2005;14(1):267–72.PubMedView ArticleGoogle Scholar
  3. Peters H, O’Leary BC, Hawkins JP, Roberts CM. The cone snails of Cape Verde: marine endemism at a terrestrial scale. Global Ecology and Conservation. 2016;7:201–13.View ArticleGoogle Scholar
  4. Tucker JK, Tenorio MJ. Illustrated catalog of the living cone shells. Wellington: MDM Publishing; 2013.Google Scholar
  5. Olivera BM, Watkins M, Bandyopadhyay P, Imperial JS, de la Cotera EPH, Aguilar MB, Vera EL, Concepcion GP, Lluisma A. Adaptive radiation of venomous marine snail lineages and the accelerated evolution of venom peptide genes. Ann N Y Acad Sci. 2012;1267(1):61–70.PubMedPubMed CentralView ArticleGoogle Scholar
  6. Cunha RL, Lima FP, Tenorio MJ, Ramos AA, Castilho R, Williams ST. Evolution at a different pace: distinctive phylogenetic patterns of cone snails from two ancient oceanic archipelagos. Syst Biol. 2014;63(6):971–87.PubMedView ArticleGoogle Scholar
  7. Cunha RL, Tenorio MJ, Afonso C, Castilho R, Zardoya R. Replaying the tape: recurring biogeographical patterns in Cape Verde Conus after 12 million years. Mol Ecol. 2008;17(3):885–901.PubMedView ArticleGoogle Scholar
  8. Tucker JK, Tenorio MJ. Systematic classification of recent and fossil conoidean gastropods: with keys to the genera of cone shells. Hackenheim: Conchbooks; 2009.Google Scholar
  9. Röckel D, Rolán E, Monteiro M. Cone shells from Cape Verde Islands - a difficult puzzle. I. Feito: Vigo, Spain; 1980.Google Scholar
  10. Rolán E: Descripción de nuevas especies y subespecies del género Conus (Mollusca, Neogastropoda) para el archipiélago de Cabo Verde. Iberus 1990, Suppl. 2:5 - 70.Google Scholar
  11. Monteiro A, Tenorio MJ, Poppe GT. The family Conidae. The west African and Mediterranean species of Conus. A Conchological Iconography. Hackenheim: ConchBooks; 2004.Google Scholar
  12. Cossignani T. Dieci nuovi coni da Capo Verde. Malacol Mostra Mond. 2014;82:18–29.Google Scholar
  13. Cossignani T, Fiadeiro R. Cinque nuovi coni da Capo Verde. Malacol Mostra Mond. 2014;84:21–7.Google Scholar
  14. Tenorio MJ, Afonso CML, Cunha RL, Rolán E. New species of Africonus (Gastropoda, Conidae) from boa vista in the Cape Verde archipelago: molecular and morphological characterization. Xenophora Taxonomy. 2014;2:5–21.Google Scholar
  15. Afonso CML, Tenorio MJ. Recent findings from the islands of Maio and boa vista in the Cape Verde archipelago, West Africa: description of three new Africonus species (Gastropoda: Conidae). Xenophora Taxonomy. 2014;3:47–60.Google Scholar
  16. Cossignani T, Fiadeiro R. Quattro nuovi coni da Capo Verde. Malacol Mostra Mond. 2014;83:14–9.Google Scholar
  17. Cossignani T, Fiadeiro R. Otto nuovi coni da Capo Verde. Malacol Mostra Mond. 2017;94:26–36.Google Scholar
  18. Cossignani T, Fiadeiro R. Tre nuovi coni da Capo Verde. Malacol Mostra Mond. 2015;86:17–21.Google Scholar
  19. Cossignani T, Fiadeiro R. Due nuovi coni da Capo Verde. Malacol Mostra Mond. 2015;87:3–5.Google Scholar
  20. Duda TF, Palumbi SR. Developmental shifts and species selection in gastropods. Proc Natl Acad Sci U S A. 1999;96(18):10272–7.PubMedPubMed CentralView ArticleGoogle Scholar
  21. Dayrat B. Towards integrative taxonomy. Biol J Linn Soc. 2005;85(3):407–15.View ArticleGoogle Scholar
  22. Uribe JE, Puillandre N, Zardoya R. Beyond Conus: phylogenetic relationships of Conidae based on complete mitochondrial genomes. Mol Phylogenet Evol. 2017;107:142–51.PubMedView ArticleGoogle Scholar
  23. Abalde S, Tenorio MJ, CML A, Zardoya R. Mitogenomic phylogeny of cone snails endemic to Senegal. Mol Phylogenet Evol. 2017;112:79–87.PubMedView ArticleGoogle Scholar
  24. Holm PM, Grandvuinet T, Friis J, Wilson JR, Barker AK, Plesner S. An 40Ar-39Ar study of the Cape Verde hot spot: temporal evolution in a semistationary plate environment. J. Geophys. Res. Solid Earth. 2008;113(B8):n/a-n/a.View ArticleGoogle Scholar
  25. Puillandre N, Bouchet P, Duda TF Jr, Kauferstein S, Kohn AJ, Olivera BM, Watkins M, Meyer C. Molecular phylogeny and evolution of the cone snails (Gastropoda, Conoidea). Mol Phylogenet Evol. 2014;78:290–303.PubMedPubMed CentralView ArticleGoogle Scholar
  26. Olivera BM. Conus peptides: biodiversity-based discovery and exogenomics. J Biol Chem. 2006;281(42):31173–7.PubMedView ArticleGoogle Scholar
  27. Filmer RM. A catalogue of nomenclature and taxonomy in the living Conidae 1758 – 1998. Leiden: Backhuys Publishers; 2001.Google Scholar
  28. Bouchet P, Gofas S. Conidae Fleming, 1822. In: MolluscaBase (2017). 2010. Accessed through: World Register of Marine Species at https://marinespeciesorg/aphiaphp?p=taxdetails&id=14107. Accessed 11 Nov 2017.
  29. Warren BH, Simberloff D, Ricklefs RE, Aguilée R, Condamine FL, Gravel D, Morlon H, Mouquet N, Rosindell J, Casquet J, et al. Islands as model systems in ecology and evolution: prospects fifty years after MacArthur-Wilson. Ecol Lett. 2015;18(2):200–17.PubMedView ArticleGoogle Scholar
  30. Crampton-Platt A, Timmermans MJTN, Gimmel ML, Kutty SN, Cockerill TD, Vun Khen C, Vogler AP. Soup to tree: the phylogeny of beetles inferred by mitochondrial metagenomics of a Bornean rainforest sample. Mol Biol Evol. 2015;32(9):2302–16.PubMedPubMed CentralView ArticleGoogle Scholar
  31. Saitoh K, Sado T, Mayden RL, Hanzawa N, Nakamura K, Nishida M, Miya M. Mitogenomic evolution and interrelationships of the Cypriniformes (Actinopterygii: Ostariophysi): the first evidence toward resolution of higher-level relationships of the world’s largest freshwater fish clade based on 59 whole mitogenome sequences. J Mol Evol. 2006;63(6):826–41.PubMedView ArticleGoogle Scholar
  32. Stöger I, Schrödl M. Mitogenomics does not resolve deep molluscan relationships (yet?). Mol Phylogenet Evol. 2013;69(2):376–92.PubMedView ArticleGoogle Scholar
  33. Osca D, Irisarri I, Todt C, Grande C, Zardoya R. The complete mitochondrial genome of Scutopus ventrolineatus (Mollusca: Chaetodermomorpha) supports the Aculifera hypothesis. BMC Evol Biol. 2014;14(1):1–10.View ArticleGoogle Scholar
  34. Tenorio MJ, Lorenz F, Dominguez M. New insights into Conus jourdani da Motta, 1984 (Gastropoda, Conidae), an endemic species from Saint Helena Island. Xenophora Taxonomy. 2016;11:32–42.Google Scholar
  35. Puillandre N, Duda TF, Meyer C, Olivera BM, Bouchet P. One, four or 100 genera? A new classification of the cone snails. J Molluscan Stud. 2015;81:1–23.PubMedView ArticleGoogle Scholar
  36. Jablonski D. Larval ecology and macroevolution in marine invertebrates. Bull Mar Sci. 1986;39(2):565–87.Google Scholar
  37. Scheltema RS. Planktonic and non-planktonic development among prosobranch gastropods and its relationship to the geographic range of species. In: Ryland JS, Tyler RA, editors. Reproduction, genetics and distribution of marine organisms. Fredensborg: Olsen & Olsen; 1989. p. 183–8.Google Scholar
  38. Swearer SE, Shima JS, Hellberg ME, Thorrold SR, Jones GP, Robertson DR, Morgan SG, Selkoe KA, Ruiz GM, Warner RR. Evidence of self-recruitment in demersal marine populations. Bull Mar Sci. 2002;70(1):251–71.Google Scholar
  39. Miller KG, Kominz MA, Browning JV, Wright JD, Mountain GS, Katz ME, Sugarman PJ, Cramer BS, Christie-Blick N, Pekar SF. The Phanerozoic record of global sea-level change. Science. 2005;310(5752):1293.PubMedView ArticleGoogle Scholar
  40. Zachos JC, Flower BP, Paul H. Orbitally paced climate oscillations across the Oligocene/Miocene boundary. Nature. 1997;388(6642):567–70.View ArticleGoogle Scholar
  41. Beddow HM, Liebrand D, Sluijs A, Wade BS, Lourens LJ. Global change across the Oligocene-Miocene transition: high-resolution stable isotope records from IODP site U1334 (equatorial Pacific Ocean). Paleoceanography. 2016;31(1):81–97.View ArticleGoogle Scholar
  42. Herbert TD, Lawrence KT, Tzanova A, Peterson LC, Caballero-Gill R, Kelly CS. Late Miocene global cooling and the rise of modern ecosystems. Nat Geosci. 2016;9(11):843–7.View ArticleGoogle Scholar
  43. Hodell DA, Curtis JH, Sierro FJ, Raymo ME. Correlation of late Miocene to early Pliocene sequences between the Mediterranean and North Atlantic. Paleoceanography. 2001;16(2):164–78.View ArticleGoogle Scholar
  44. Krijgsman W, Hilgen FJ, Raffi I, Sierro FJ, Wilson DS. Chronology, causes and progression of the Messinian salinity crisis. Nature. 1999;400(6745):652–5.View ArticleGoogle Scholar
  45. Lisiecki LE, Raymo ME. A Pliocene-Pleistocene stack of 57 globally distributed benthic δ18O records. Paleoceanography. 2005;20:PA1003.Google Scholar
  46. Davis KE, Hill J, Astrop TI, Wills MA. Global cooling as a driver of diversification in a major marine clade. Nat Commun. 2016;7:13003.PubMedPubMed CentralView ArticleGoogle Scholar
  47. Dyhr CT, Holm PM. A volcanological and geochemical investigation of boa vista, Cape Verde Islands; 40Ar/39Ar geochronology and field constraints. J Volcanol Geotherm Res. 2010;189(1–2):19–32.View ArticleGoogle Scholar
  48. Kohn AJ, Nishi M, Pernet B. Snail spears and scimitars: a character analysis of Conus radular teeth. J Molluscan Stud. 1999;65:461–81.View ArticleGoogle Scholar
  49. Duda TF, Kohn AJ, Palumbi SR. Origins of diverse feeding ecologies within Conus, a genus of venomous marine gastropods. Biol J Linn Soc. 2001;73(4):391–409.View ArticleGoogle Scholar
  50. Dowle EJ, Morgan-Richards M, Brescia F, Trewick SA. Correlation between shell phenotype and local environment suggests a role for natural selection in the evolution of Placostylus snails. Mol Ecol. 2015;24(16):4205–21.PubMedView ArticleGoogle Scholar
  51. Hollander J, Butlin RK. The adaptive value of phenotypic plasticity in two ecotypes of a marine gastropod. BMC Evol Biol. 2010;10(1):333.PubMedPubMed CentralView ArticleGoogle Scholar
  52. Knowlton N. Sibling species in the sea. Annu Rev Ecol Syst. 1993;24(1):189–216.View ArticleGoogle Scholar
  53. Roux C, Fraïsse C, Romiguier J, Anciaux Y, Galtier N, Bierne N. Shedding light on the Grey zone of speciation along a continuum of genomic divergence. PLoS Biol. 2016;14(12):e2000234.PubMedPubMed CentralView ArticleGoogle Scholar
  54. Gerard D, Gibbs HL, Kubatko L. Estimating hybridization in the presence of coalescence using phylogenetic intraspecific sampling. BMC Evol Biol. 2011;11(1):291.PubMedPubMed CentralView ArticleGoogle Scholar
  55. Sambrook J, Fritsch EF, Maniatis T. Molecular cloning: a laboratory manual. 2nd ed. New York: Cold Spring Harbor Laboratory Press; 1989.Google Scholar
  56. Kornobis E, Cabellos L, Aguilar F, Frías-López C, Rozas J, Marco J, Zardoya R. TRUFA: a user-friendly web server for de novo RNA-seq analysis using cluster computing. Evol Bioinforma. 2015;11(Supplementary Material 23873):97–104.Google Scholar
  57. StJohn J. SeqPrep. 2011. https://githubcom/jstjohn/SeqPrep.
  58. Andrews S. FastQC. 2010. http://wwwbioinformaticsbabrahamacuk/projects/fastqc/.
  59. Schmieder R, Edwards R. Quality control and preprocessing of metagenomic datasets. Bioinformatics. 2011;27(6):863–4.PubMedPubMed CentralView ArticleGoogle Scholar
  60. Grabherr MG, Haas BJ, Yassour M, Levin JZ, Thompson DA, Amit I, Adiconis X, Fan L, Raychowdhury R, Zeng Q, et al. Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nat Biotech. 2011;29(7):644–52.View ArticleGoogle Scholar
  61. Schattner P, Brooks AN, Lowe TM. The tRNAscan-SE, snoscan and snoGPS web servers for the detection of tRNAs and snoRNAs. Nucleic Acids Res. 2005;33(suppl 2):W686–9.PubMedPubMed CentralView ArticleGoogle Scholar
  62. Boore JL, Macey JR, Medina M. Sequencing and comparing whole mitochondrial genomes of animals. Methods Enzymol. 2005;395:311–48.PubMedView ArticleGoogle Scholar
  63. Abascal F, Zardoya R, Telford MJ. TranslatorX: multiple alignment of nucleotide sequences guided by amino acid translations. Nucleic Acids Res. 2010;38(Web Server issue):W7–13.PubMedPubMed CentralView ArticleGoogle Scholar
  64. Katoh K, Standley DM. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol. 2013;30(4):772–80.PubMedPubMed CentralView ArticleGoogle Scholar
  65. Castresana J. Selection of conserved blocks from multiple alignments for their use in phylogenetic analysis. Mol Biol Evol. 2000;17(4):540–52.PubMedView ArticleGoogle Scholar
  66. Glez-Peña D, Gómez-Blanco D, Reboiro-Jato M, Fdez-Riverola F, Posada D. ALTER: program-oriented conversion of DNA and protein alignments. Nucleic Acids Res. 2010;38(suppl 2):W14–8.PubMedPubMed CentralView ArticleGoogle Scholar
  67. Felsenstein J. Evolutionary trees from DNA sequences: a maximum likelihood approach. J Mol Evol. 1981;17(6):368–76.PubMedView ArticleGoogle Scholar
  68. Huelsenbeck J, Ronquist F. MrBayes: Bayesian inference of phylogenetic trees. Bioinformatics. 2001;17:754–5.PubMedView ArticleGoogle Scholar
  69. Stamatakis A. RAxML-VI-HPC: maximum likelihood-based phylogenetic analyses with thousands of taxa and mixed models. Bioinformatics. 2006;22(21):2688–90.PubMedView ArticleGoogle Scholar
  70. Ronquist F, Huelsenbeck JP. MrBayes 3: Bayesian phylogenetic inference under mixed models. Bioinformatics. 2003;19(12):1572–4.PubMedView ArticleGoogle Scholar
  71. Rambaut A, Drummond AJ. Tracer v1.4. 2007. Available from http://beast.bio.ed.ac.uk/Tracer.
  72. 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(3):772–3.PubMedGoogle Scholar
  73. Akaike H. Information theory and an extension of the maximum likelihood principle. In: Petrov BN, Csaki F, editors. 2nd international symposium on information theory. Budapest: Akademiai Kiado; 1973. p. 267–81.Google Scholar
  74. Drummond A, Rambaut A. BEAST: Bayesian evolutionary analysis by sampling trees. BMC Evol Biol. 2007;7:214.PubMedPubMed CentralView ArticleGoogle Scholar
  75. Duda TF, Bolin MB, Meyer CP, Kohn AJ. Hidden diversity in a hyperdiverse gastropod genus: discovery of previously unidentified members of a Conus species complex. Mol Phylogenet Evol. 2008;49(3):867–76.PubMedView ArticleGoogle Scholar
  76. Sacco F: I molluschi dei terreni terziarii del Piemonte e della Liguria. Conidae e Conorbidae, vol. XIII. Torino. Italy: Stamperia Reale; 1893.Google Scholar
  77. Glibert M. Les Conacea fossiles du Cénozoïque étranger des collections de l’Institut Royal des Sciences Naturelles de Belgique. Institut Royal des Sciences Naturelles de Belgique Mémoire, série 2. 1960;64:1–132.Google Scholar
  78. Paradis E, Claude J, Strimmer K. APE: analyses of phylogenetics and evolution in R language. Bioinformatics. 2004;20(2):289–90.PubMedView ArticleGoogle Scholar
  79. Revell LJ. Phytools: an R package for phylogenetic comparative biology (and other things). Methods Ecol Evol. 2012;3(2):217–23.View ArticleGoogle Scholar
  80. Pybus OG, Harvey PH. Testing macro–evolutionary models using incomplete molecular phylogenies. Proc R Soc Lond B Biol Sci. 2000;267(1459):2267.View ArticleGoogle Scholar

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