Molecular phylogenetic evidence refuting the hypothesis of Batoidea (rays and skates) as derived sharks

Christophe J. Douady, Mine Dosay, Mahnmood S. Shivji, and Michael J. Stanhope

Molecular Phylogenetics and Evolution 26 (2003) 215-221

Abstract

Early morphological study:Elasmobranch (Sharks, Batoids)Sharks : monophyleticBatoids (skates and rays): monophyletic

Modern cladistic study: Hypnosqualea hypothesis: Batoids are derived sharks (sawsharks, angelsharks)

sawshark angelshark

To address this issue: 2.4kb mtDNA (12S rRNA-tRNA-16S rRNA locus)-Two order of batoides-All order of sharks(8)-Outgroup: holocephali ( sister group to elasmobranchs)

Results:Sharks: monophylyRejection of the Hypnosqualea hypothesis

Introduction

Elasmobranch fishes / Compagno(1973,1977)

Superorder Orders Common name

1.Galeomorphii 1.Orectolobiformes Carpet sharks

2.Heterodontiformes Bullhead sharks

3.Carcharhiniformes Ground sharks

4.Lamniformes Mackerel sharks

2.Squatinomorphii 5.Squatiniformes Angel sharks

3.Squalomorphii 6.Hexanchiformes Frilled and cow sharks

7.Squaliformes Dogfish shark

8.Pristiophoriformes Saw sharks

4.Batoidea 9.Myliobatiformes Stingrays

10.Pristiformes Sawfishes

11.Torpediniformes Electric rays

12.Rajiformes Skates

13.Rhinobatiformes Guitarfishes

The relative relationships between these orders and superorders remains uncertain

History

1. Early morphological work:

-A fundmental split between batoids and sharks (Bigelow and Schroeder 1948, 1953)

2. Modern morphological cladistic study

- Batoids are derived sharks, closely related to sawsharks and angelsharks (Compagno, 1977~ Shirai, 1996)

Hypnosqualea hypothesis: (Squatinomorphii (Pristiophoriformes,Batoidea))Þ Superorder Hypnosqualea (Carvalho and Maisey, 1996)

Study of Interordinal relationships of sharks: very few

3 relevant studies: Dunn and Morrisey(1995), Kitamura et al. (1996), Arnason(2001)

1. Kitamura : Cytb(732bp), 8 of 13 elasmobranch orders, - No support values, rooted with a very distant outgroup not Holocephali- No chimaeras were not used as an outgroup -> difficult to interpret- Support a possible distinction of batoids from hypnosqualea- Not good marker(Cytb)

2. Dune and Morrissey: 12S rRNA gene(303bp), 5 orders without any taxa related with hypnosqualea hypothesis (no batoids)

3. Arnason : complete mitogenome, 3 order of sharks, a skate and a ratfish- Rejection of the concept of Squalea(shirai, 1992): including skate.- No squaliform representative on same branch as the skate

=> Poorly represented elasmobranch taxonomic diversity

Purpose:- To provide a molecular phylogenetic perspective “whether sharks are paraphyletic, due to a de-rived position for Batoidea,

or whether sharks are monophyletic, to the ex-clusion of Batoidea.

- An examination of several additional questions pertaining to elasmobranch interordinal affinities

2. Materials and methods2.1. Taxa and phylogenetic loci

* mitochondrial 12S– 16S and intervening tRNA valine, : - powerful phylogenetic marker for other groups of vertebrates (Springer et al., 2001) -preliminary sequence data indicated there was little or no transition saturation, even in comparisons involving the chimaera sequence(s) (data not shown).

The sequence alignments: without ambiguous positions /software SOAP:the effect of removing all the positions that are sensitive to variationin alignment position. - Rooting phylogenetic trees: Outgroups increase the noise in the se-quence data but are required to polarise the trees. - single outgroups : more subjective to the affects of long branchattraction than multiple paraphyletic taxa. ÞDifferent outgroup permutations. The more outgroups utilized, the fewer positions were included in the results.Þ This permutation of outgroups provided an additional perspective on the batoid/shark diphyletic hypothesis

Batoidea

Galeo

Squalo

Squatino

(2010)

(1963)

440437420

2.2. Phylogenetic reconstruction

* Modeltest software (Posada and Crandall, 1998) ;to objectively determine the best suited model of sequence evolution and the accompanying parameter values for these data

- Maximum likelihood (ML): TN+G+ I : used for the 420, 437, 440 bp data sets The 1880, 1963, and 2010 data sets all used GTR+G+ I - Maximum parsimony (MP) -Minimum evolution (ME) / using both ML and LogDet distances. / using PAUP* 4b8 (Swofford, 1998).- Bayesian phylogenetic analyses in Mr.Bayes 2.01- GTR model allowing a gamma shape of among-site rate variation

- Statistical significance, under a likelihood model, of several phylogenetic hypotheses, was assessed using the KH test

3. Results and discussion

Order:MLMPME mlME LogDetBayesian

- all trees support amuch more an-cient split be-tween the two groups, withbatoids as the sis-ter group to a clade consisting of allshark orders

-Shark monophyly

-Hypnosqualea hypothesis were significantly re-jected for all data sets in the KH test

Despite of lacks representatives of the putative deepest branching batoid or-ders (e.g.,Torpediniformes), no long branch attraction has generated the basal position of Batoidea.

1, all tree reconstruction methods recover this position, not only maximum parsimony(known to be sensitive to this reconstruction artefact)

2, the batoid-shark branch is far from being the longest, with the branch leading to nurse shark and the Hexanchiformes branch

3.unroot the tree, the branching arrangement remains unchanged * shark monophyly, to support batoid monophyly: these data represent the first time a skate and ray are included in the same molecular phylogenetic analysisw.

-The reduced data sets of 440, 437, and 420 bp also supported distinction be-tween batoids and sharks, for shark monophyly, for the monophyly of Holocephali.(If additional sequence: Chimaeridae would be paraphyletic), * All elasmobranch, chondrichthyan,batoid monophyly

•Several superordinal associations within sharks were strongly supported.

-Squaliformes (Piked dogfish and gulper shark) with an angelshark/sawshark clade. =>the earlier morphological hypotheses by Shirai (1992b) and de Carvalho (1996), :Squaliformes as the sister group to Hypnosqualea. With-out Batoidea from Hypnosqualea, would agree with this arrangement

-Shirai (1992b, 1996) and de Carvalho (1996) suggested the concept of Squalea, which has the six and seven gill sharks as the sister group to Squaliformes/Hypnosqualea.

Mode of reproduction for Squalea (i.e., the association of Hexanchi-formes, Squaliformes, Pristiophoriformes, and Squatiniformes) is yolksac viviparous (Compagno, 1990).Batoid : uterine to oviparous

Compagno(1990)Extended oviparous -> retained oviparous => yolksac viviparous => canni-bal, placental or uterine viviparous.

Compagno (1973, 1977) /Shirai (1996)/ de Carvalho (1996) is Galeomorphii: (((Lamniformes/Carcharhiniformes) Orectolobiformes) Heterodontiformes )))

The only portion of this hypothesis that is clearly evidentfrom our analyses is the lamniform/carcharhiniform association. (It’s ok)The rest: Low bootstrap value / high p valueCertainly our data are insufficient to either accept or reject Shirai and de CarvalhosGaleomorphii arrangement (2110 data set)

Our study provides the first convincing molecular case for rejection of the Hypnosqualea hypothesis and suggests that the seven or more putative morphological synapomorphiesused to define this superorder are either symplesiomorphic or the consequence of convergent evolution in the ancestor to sawsharks/angelsharks and batoids (depending on the resolution of Galeomorphii).

Our data strongly suggest that an evolutionary scenario for the diversifi-cation of sharks and batoids is an early fundamental split into two respec-tively monophyletic lineages.

A basal vs. derived position for Batoidea will have significant implica-tions inthe polarization of morphological and life history characteristics for elas-mobranchs and means that much of the current thinking in this regard needs to be re-evaluated.