Biology ofA nomuraII (A .Asa kura，e d. )，C rustac ean Resea rch，S pec ial N umber6: 15 1-166，2006 Phylogenetic relationships among western Atlantic Porcellanidae (Decapoda: Anomura)，based on partial sequences of the mitochondrial16S rRNA gene，with comments on morphology Irene Teresa Rodriguez，Gonzalo Hernandez and Darryl L. Felder Abstract.-We obtained 47 partial sequences as porcelain crabs. The family is considered of the mitochondrial16S rRNA gene for 20 west- awell-defined taxon within the superfamily ern Atlantic，three amphi-American，and two Galatheoidea Samouelle，1819 (Haig，1960) eastern Pacific species of porcellanids，grouped Other members ofthe superfamily are Aeglidae into eight genera.Neighbor Joining，Maximum Dana，1852 ，Chirost ylidae Ortmann，1892 ，and Parsimony，Maximum Li kelihood，and Bayesian Galatheidae Samouelle，1819 (Maロin & Davis， Analysis were the methods used to infer phy- 2001). logenetic relationships between and within the Porcelain crabs occur worldwide，primar- genera. The family is divided into two main ily in intertidal and sublittoral zones of tropi- clades，one with POかonyx and Porcellana，and cal and subtropical regions where they occupy the other with the remaining six genera treated here.Larval ，sperm ，and spermatophore mor- habitats such as depressions under stones， phology largely support this obvious dichotomy. spaces in worm tubes，cavities of sponges，and Species of Clastotoechus were grouped closely excavations in coral reefs (Haig，1960; Gore， together，which raises questions regarding the 1972，1982 ;Harvey ，1999; Hernandez，1999; validity of the monotypic genus Madarateuchus Werding etal. ，2003). Most species are free- recently proposed for Clastotoechus vanderhorsti. living ，but afew live commensally (Haig，1960 ; Intrageneric divergence in Megalobrachium Werding，1977 ，1983b; Baeza & Thiel ，2000 ; suggests the possible need for resurrection of Baeza eta l. ，2001). Porcellanids are primarily Porcellanopsis. Megalobrachium soriatum ap- filter feeders and use their highly specialized pears to contain acryptic species. Neopisosoma is setose third maxillipeds to extract minute food supported as agenus distinct from Pachycheles， particles of plankton and detritus 企omthe wa- which contained two independent lineages. ter.They can also take advantage of accumu- Petrolisthes is resolved into two clades，as sup- lated detritus by direct deposit feeding (Kropp， ported by previous morphological and molecular 1981). Their feeding habits allow them to evidence. In one clade Petrolisthes violaceus is ， occupy awide variety of habitats .Within the closely related to P. tridentatus，P. quadratus，P. family，Parap etroli sth estortug ensis (Glassell ， lewisi，and P. magdalenensis. The second clade contains three lineages: one with Petrolisthes ar- 1945) is unique in the shape of the fingers matus，P. marginatus，and P. politus as asubclade; and the third maxillipeds.Fingers are deeply another with P. rosariensis as an independent grooved along cutting edges，spooned and trun- lineage; and lastly the Petrolisthes galathinus- cate at tips (Haig，1962) .The filter setae of the complex，with P. caribensis，P. galathinus (with third maxillipeds are short and scarce ，with a cryptic species) ，and Parapetrolisthes tortugensis. brush-like setation on the dactylus that seems an adaptation to scraping material from the sub- strate (Kraus etal. ，2001). These features are INTRODUCTION shared with the galatheids and could represent Porcellanidae Haworth，1825 ，is afamily homologous，ancestral characters (Kraus etal. ， of anomuran crustaceans commonly known 2001) or convergent adaptations to their feed- ρ 7 T P H i し n o u ρV 152 - - U J u L G u u b I l r ing habits. The genus P etrolist hes Stimpson，1858 ，is Most porcellanids hatch as aprezoea and the largest in the family，containing over 100 molt through only two subsequent zoeal stages species. Based on aqualitative examination and one megalopal stage (postlarva or decapo- of morphological characters，several natural dite) .The reduced number of zoeal stages is groups，including two large groups，have been an abbreviated scheme of development，usu- distinguished. One large group consists of spe- ally considered as phylogenetically advanced， cies that have teeth or spines on the anterior in which the larval history is foreshortened in margin of the carpus of chelipeds，one or two comparison to related taxa，both in number of posterodistal spines on the merus of walk- instars and the duration of each (Gore，1985) . ing leg 1，and epibranchial ，supraorbital ，or This can result in areduced period of plank- other spines on the carapace.The other large tonic life in which larval dispersal is potentially group consists of species in which the ca叩us very limited . of the chelipeds is not armed with teeth or Porcellanids number nearly 280 species spines and spines are lacking on the carapace partitioned among 30 genera. Near1 yhalf of and walking legs (Haig，1960). The genus the species in this family have been recorded Pachycheles Stimpson，1858 ，is the second in the Americas (Gore，1982; Werding et al. ， most speciose of the family and includes over 2003). Seven species are reported to oc- 40 species (Rodriguez et al. ，2004). This ge- cur on both East and West American coasts: nus is aheterogeneous assemblage of species， Pachycheles chacei Haig，1956 ，P .monil 件r some of them previously assigned to Pisosoma (Dana，1852) ，Petrolisthes armatus (Gibbes， Stimpson，1858 ，which is currently considered 1850)，P. galathinus (Bosc，1802) ，P .robsonae asynonym of Pachycheles (Haig，1960). Glassell ，1945 ，P. tonsorius Haig，1960 ，and P. Two previous molecular-based studies of tridentatus Stimpson，1858 (Haig，1956 ，1960; Porcellanidae have been published. A mo・ Gore & Abele，1973; Scelzo，1982 ;Abele & lecular phylogeny of eastern Pacific genera Kim，1989; Rodriguez etal. ，2005). Forty- Petrolisthes and Pachycheles was based on mi- eight valid species and two undescribed species tochondrial 16S rRNA gene (Stillman & Reeb， grouped into eleven genera occur in the westem 2001) and included afew sequences of Atlantic Atlantic (Rodriguez et al .，2005). specimens for amphi-American species of Phylogenetic relationships of porcellanids P etrolis thes. In this study，the subdivision of are poorly understood. Porcellanid genera Petrolisthes into two main clades supported ear- have long warranted revision on aworldwide lier division based on morphological characters basis にhace，1942; Haig，1956 ，1960; Werding (Haig，1960). One of these clades containted et α1.，2001; Rodriguez et al .，2005) . ln the only species possesing regular serrate spines ， last 50 years，the number of described porcel- and the other clade contained species that are lanid species in the western Atlantic increased smooth or irregularly decorated (“ spiny"and from 31 to 48 with the addition of newly col- “smooth" species，respectively ，se nsu Stillman lected and described cryptic species (Werding and Reeb，2001). A second molecular study & Hiller，2002; Rodriguez etal. ，2005). The used partial sequences of the mitochondrial family includes three widely distributed and COI gene to infer phylogenetic relationships morphologically variable species complexes， between two species of the genus Neopisosoma Petrolisthes armatus，P. galathinus ，and P. Haig，1960 (Werding etal. ，2001). It was sug- tridentatus; and their taxonomic status remains gested that Neopisosoma should not be treated unclear (Werding et al. ，2003; Rodriguez et al. ， as separate from Pachycheles ，because it was 2005). found to be paraphyletic. However，a limited Morphological and molecular studies can number of taxa have been studied to date，and facilitate understanding of systematic and phy- the status of Neopisosoma remains unclear logenetic relationships within the Porcellanidae. (Rodriguez etal. ，2004 ，2005). Phylogeny of westem Atlantic Porcellanidae 153 In the present study，partial sequences of obtain apelle t. The pellet was washed in 20μ1 the mitochondrial 16S rRNA gene were used to 70% ethanol ，centrifuged at 14，000 rpm for 1 elucidate phylogenetic relationships within the min，and Iyophilized in aONAIIO SpeedVac Porcellanidae in the westem Atlantic in order (TherrnoSavant) at high drying rate for 30 min. to provide insights into classification ，detect Genomic ONAwas re-suspended in 10 -50μl cryptic species，and gain knowledge of phylo- of 10X TE Buffer (1 mMEOTA ，10 mMTris ， geographic pa口ems in this family. pH 7.5) or ddHpand stored at -20 oC. DNAamplification. - Amplification of a- 545 basepair product from the mitochon- MATERIALS AND ETHODS 九' 1 drial 16S rRNA gene was carried out by a Sample coIl ection and depositories polymerase・chain-reaction (PCR) using two Most porcelain crabs were collected in forward primers，16Sar (5'-CGCCTGTTTAT coastal Florida (J uly 2000，July -August 2001)， CAAAAACA下3') (Palumbi etal. ，1991) and along the coasts of Louisiana (June 2001)，and 16L2 (5'-TGCCTGTTTATCAAAAACAT・3') Texas (September 2001)，U.S.A.; from Veracruz (Schubart etal. ，2002) ，and the reverse primer to Quintana Roo，Mexico (J uly 2002); in Belize 1472 (デー AGATAGAAACCAACCTGG-3') (October 2002); in the vicinity of La Tortuga， (Crandall & Fitzpatrick，1996). The PCR mix Margarita，and Cubagua Islands，Venezuela for each sample contained 6.7μ1 ddHP，3.0μl (March-June 1996，April -June，2001); and on 10 X PCR buffer，3.0μ1 MgCI2(25 mM)，5.0 the Paci白c coast ofNicaragua (September 2000， μ1 Betaine (5 M)，4.0μ1 dNTP's (1.25 mM)， November 2001). For each species，one speci- 0.5μ1 of each forward primer (20μM)，I.Oμl men per locality was used in the analyses，as of reverse primer (20μM)，0.5μ1 AmpliTaq listed in Table 1.AII specimens were preserved Gold骨 (5 U /μ1 ) ，and 1. 0μ1 ONAtemplate (-20 in 70-95% ethano l. Genetic vouchers were de- ng/μ1 ) .The PCR profile was 42 cycles (de- posited in the Zoological Collection，University naturing at 940 C I1 min，annealing at 480 C I1 of Louisiana at Lafayette，Louisiana ，U .S.A. min，and extension at 72 0 C I1 :30 min). Four (ULLZ). Some previously archived mater卜 μ1 of PCR products underwent electrophoresis als from ULLZ and the Coleccion Nacional de on a 1.5 % agarose gel and were visualized Crustaceos，Instituto de Biologia，Universidad with ethidium bromide staining.