0.1

Chamelea gallinaVenus verrucosaVenus casina

Clausinella punctigeraVenus rosalinaClausinella fasciata

Globivenus toreumaGlobivenus rugatina

Ventricoloidea foveolataGlobivenus effosaGlobivenus isocardia

Dosina zelandicaAmeghinomya antiqua

Eurhomalea lenticularisTawera spissa

Timoclea ovataTimoclea subnodulosa

Timoclea levukensisAntigona lamellaris

Lirophora mariaeLirophora paphiaAnomalocardia brasiliana

Mercenaria campechiensisMercenaria mercenaria

Humilaria kennerleyiPuberella intapurpurea

Ameghinomya sp2Chione cancellata

Chione subimbricataChionista fluctifraga

Protothaca mcgyntyiCallithaca tenerrima

Protothaca stamineaRuditapes decussata GB

Ruditapes decussataPaphia euglypta

Paphia vernicosaPaphia dura

Katelysia sp 1Katelysia sp 2

Katelysia rhytiphoraKatelysia scalarina

Ruditapes bruguieriRuditapes philippinarum G B

Pectunculus exoletaDosinia sp S hark Bay

Dosinia victoriaeDosinia sp E sperance

Placamen berryiPlacamen flindersi

Periglypta listeriPeriglypta puerpera

Macrocallista squalidaPitar rudis

Meretrix lyrataCallista chione

5452 96

965454

7172

8076

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100100

9999

99100

8585

6769

100100100100

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100

100100

595699

99

99999894

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98

6565

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Chioninae s.s.

Venerinae s.novo

Dosiniinae

Tapetinae

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Kappner & Bieler, Mol. Phylogen. Evol. in press.

Phylogenetic studies of the marine bivalve subfamily Venerinae (Bivalvia: Veneridae)Isabella Kappner, Rüdiger BielerDepartment of Zoology (Invertebrates), Field Museum of Natural History, Chicago, IL, ikappner@fieldmuseum.org

Goal of present studiesHow did morphological diversity evolve over time in the marine bivalve subfamily Venerinae?

• Investigate evolutionary relationships using mitochondrial and nuclear gene sequences

• Provide a robust phylogenetic framework for understanding venerine morphological character evolution

• Test traditional systematic hypotheses

Taxonomic controversyVenerine shells are morphologically very similar to those of the subfamily Chioninae and can only be distinguished from each other by the presence or absence of an anterior lateral tooth. Taxonomists have been arguing for over 150 years about the significance of this minute anterior lateral tooth and the taxonomy of this group remains controversial.

Fischer-Piette (1975) carried out the last major revision of the Venerinae. He separated the two subfamilies and revised the Venerinae as a one-genus subfamily (Venus). In the most recent study dealing with this subject, Chioninae and Venerinae were synonymized based on morphological features Coan and Scott (1997). Venerinae s.l. currently comprises 41 extant genera (10 former Venerinae and 31 former Chioninae) with over 180 species.

Results & DiscussionThe phylogeny of the combined data (16S, COI and

H3) strongly suggests that Chioninae s.s. and Venerinae s.novo are discrete clades.

Several genera (Chamelea, Clausinella, Tawera, Timoclea) were probably mis-classified in the past due to homoplasy in morphological features

Monophyly of Venerinae s.l. or Venerinae s.s. (Keen, 1969) was not present in the resulting 50% majority rule consensus tree.

The two tested traditional systematic hypotheses are rejected with p<0.001 in the constraint analysis and outside the 99.9% confidence set in ELW test.

A combination of morphological characters can be used to distinguish the two subfamilies (Fig. 3):

Venerinae have separated siphons and most taxa in this group have an anterior lateral tooth

Chioninae have fused siphons and lack an anterior lateral tooth.

Within the Venerinae no monophyletic generic groupings have been identified

Problems with sequencing COI14 of the 56 sampled taxa did not deliver any

results for the partial COI gene

Museum material often in unknown fixatives can cause difficulties in amplifying longer sequences

“Barcoding” will likely be successful with fresh material, but could be very labor and time intensive with older museum material.

MethodsWe performed a phylogenetic analysis of our 3-gene dataset based on partial sequences of the mitochondrial 16S gene (602 bp), COI gene (569 bp) and nuclear Histone 3 gene (328 bp) including 55 species of 37 genera of Venerinae s.l. as well as 18 outgroup taxa of other venerid subfamilies.

Alignments were analyzed by a mixed model Bayesian approach with Markov Chain Monte Carlo (B/MCMC) methods using MrBayes 3.1.2 and maximum parsimony methods using PAUP*. MrBayeswas set to produce 3,000,000 generations and to run four replicas of four chains simultaneously. Trees were sampled every 100 generations for a total of 30,000 trees. The software Tracer 1.2 was used to determine the “burn in”. Maximum parsimony analysis was carried out with 1000 random sequence additions, and TBR branch swapping. Branch support was examined by undertaking 300 jackknife and 2000 bootstrap replicates with a minimum of 10 random sequence additions per replicate.

Taxon samplingFrom the traditional Venerinae we sampled seven of ten genera and 15 of 31 chionine genera. Outgrouptaxa were chosen from a concurrent study of our working group (Mikkelsen et al. (in press)).

Hypothesis testingBased on our results two traditional systematic hypotheses (Fig. 1) were tested. This was performed by examining suboptimal trees present in the B/MCMC sample and by calculating expected likelihood weights (ELW) with TreePuzzle 5.2.

Sponsored by NSF-PEET DEB-9978119

Morphological Trait MappingAltogether 75 conchological characters as well as six characters of internal anatomy were investigated (data not presented here). The morphological characters were mapped onto the molecular topology of the concatenated analysis and three informative characters were found:

(1) Presence or absence of crenulations in interior shell margin (Fig. 3B)

(2) Presence (Fig. 3C1) or absence (Fig. 3C2) of anterior lateral tooth in left valve

(3) Degree of siphon fusion; partially to completely separated (Fig. 3D1) vs. completely fused (Fig. 3D2)

Literature citedCoan, E.V., Scott, P.H., 1997. Checklist of the marine bivalves of

the northeastern Pacific Ocean. Sta. Barbara Mus. Nat. Hist. Contrib.Science 1, 1-28.

Fischer-Piette, É., 1975. Révision des Venerinae s.s. (Mollusques Lamellibranches). Mem. Mus. Natn. Hist. Nat. A Zool., Paris 93, 1- 64.

Kappner, I., Bieler, R. in press. Phylogeny of Venus clams (Bivalvia: Venerinae) as inferred from nuclear and mitochondrial gene sequences. Mol. Phylogen. Evol.

Keen, A.M., 1969. Superfamily Veneracea. In: Cox, L.R. et al., Part N [Bivalvia], Mollusca 6, volume 2: ii + pp. N491- N952. In: Moore, R.C. (Ed.) Treatise on Invertebrate Paleontology. Lawrence, Kansas: Geological Society of America and University of Kansas, N670-N690.

Mikkelsen, P.M., Bieler, R., Kappner, I., Rawlings, T.A., in press. Phylogeny of Veneroidea (Mollusca: Bivalvia) based on morphology and molecules. Zool. J. Linn. Soc.

AcknowledgementsThanks to the curators and staff of following museums for loan of specimens and tissue: MNHN, Paris; IRSNB, Brussels; NMNZ, Wellington, New Zealand; FLMNH, Gainesville; LACM, Los Angeles; BMNH, London.

For assistance during field collections we thank: Katja Defren-Janson (Berlin, Germany), Sonia Merino (Instituto Nacional de Desenvolvimento das Pescas - Mindelo, Cabo Verde), ElisioDelgado and crew (Mindelo, Cape Verde Islands), Eurico Barrosand crew (Sal, Cabo Verde), Patrice Petit De Voize (FédérationFrançaise d'Etudes et de Sports Sous-Marins - Commission nationale de Biologie, France), Haidar el Ali (Oceanium, Dakar, Senegal), Lisa Kirkendale (FLMNH), Emily Glover and John Taylor (BMNH), Fred Wells (Western Australian Museum, Perth), and Rachel Collin (Smithsonian Tropical Research Institute, Panama).

We are grateful to the following people for help in obtaining additional specimens: Claude Berthoult (Centre ORSTOM de Nouméa, New Caledonia), Alan G. Beu (Institute of Geological and Nuclear Sciences, Lower Hutt, New Zealand), Rachel Collin (Smithsonian Tropical Research Institute, Panama), Louise Crowley (American Museum of Natural History, New York), Brian Dyer (Universidad del Mar, Chile), Leonore Kappner (Bochum, Germany), Alan J. Kohn (University of Washington, Seattle), Taeko Kimura (Mie University, Mie, Japan), and Melita Peharda(University of Zagreb, Croatia).

Additional funds were provided by UIC’s Provost Award for Graduate Research, FMNH Zoology Department’s Marshall Field Fund, and the Conchologists of America.

Fig. 1 Different traditional systematic hypotheses (a) after Keen (1969) and (b) after Coan and Scott (1997) (author’s interpretation).

Figure 3. Morphological features that were found to be useful for classification and that were utilized for mapping on the molecular tree. A. Overview of internal shell, B. Crenulations of the interior shell margin, C. Presence of an anterior lateral tooth of Type I (C1) and absence of anterior lateral tooth (C2) in the left valve. D. Overview of internal morphology, D1. Separated siphons, D2. Fused siphons. A, B, C1, and D1. Venus verrucosa Linnaeus, 1758, C2, and D2. Lirophora paphia (Linnaeus, 1767).

B

D1 D2

Figure 2. Molecular phylogeny of Veneridae: 50% majority rule consensus tree based on a Bayesian analysis of the concatenated dataset (16S, COI and H3). Posterior probability values > 0.95 are indicated by bold lines. Support values > 70 from jackknife replicates are indicated below the branches, and from bootstrap replicates above branches. Morphological trait mapping onto the molecular phylogeny of venerid bivalves: Character 1 (crenulations in internal margin), black boxes = presence, white boxes = absence; Character 2 (anterior lateral tooth of Type I), black boxes = presence, white boxes = absence; Character 3 (siphons), black boxes = separated, white boxes = fused.

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