Phylogenetic Relationships of the Enigmatic Land Snail Genus

Biological Journal of the Linnean Society, 2009, 96, 203–221. With 10 ﬁgures

Phylogenetic relationships of the enigmatic land snail genus Prestonella: the missing African element in the Gondwanan superfamily Orthalicoidea (Mollusca: Stylommatophora) Downloaded from https://academic.oup.com/biolinnean/article/96/1/203/2448044 by guest on 27 September 2021

DAVID G. HERBERT1,2* and ANDREW MITCHELL3

1Natal Museum, Private Bag 9070, Pietermaritzburg 3200, South Africa 2School of Biological and Conservation Sciences, University of KwaZulu-Natal, Private Bag X01, Scottsville, Pietermaritzburg, 3209, South Africa 3NSW Department of Primary Industries, Wagga Wagga Agricultural Institute, PMB Wagga Wagga, NSW 2650, Australia

Received 7 January 2008; accepted for publication 21 April 2008

The land snail superfamily Orthalicoidea, although generally assumed to be of Gondwanan origin, is considered by the majority of recent authors to be absent from the African continent. However, two poorly-known African genera, Aillya and Prestonella, have historically been referred to the orthalicoid family Bulimulidae s.l. Anatomical study of Aillya has subsequently shown it to be morphologically distinct from the Bulimulidae and referable to a family of its own, outside the Orthalicoidea, but Prestonella has remained an enigmatic taxon of unknown affinity. Using molecular and morphological evidence, we demonstrate conclusively that Prestonella is indeed a member of the Bulimulidae s.l. We thus conﬁrm that this family is represented in Africa, and that it has a classical disjunct, tri-continental southern distribution. Thus, either the origin of the family must at the least predate the separation of Africa and South America in the Mid Cretaceous (under a vicariance scenario) or there must have been subsequent dispersal between the isolated Gondwanan fragments. In view of the limited dispersal ability of terrestrial snails, we consider the former more likely. Anatomically, Prestonella exhibits many character states thought to be plesiomorphic, suggesting a relationship with the subfamily Bulimulinae. Bayesian analysis of nuclear DNA sequence data places it as sister group (posterior probability = 1.0) to an Australasian clade comprising Bothriembryon and Placostylus. However, taxon sampling within the Orthalicoidea is currently inadequate to permit meaningful resolution of subfamilial affinity using molecular data. Similarly, although those orthalicoid taxa for which molecular data are available comprise a well-supported clade, the relationships of this clade to other stylommatophoran clades remain unresolved. © 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 96, 203–221.

ADDITIONAL KEYWORDS: Africa – Bulimulidae – Gondwana – molecular phylogeny – morphology.

INTRODUCTION tion and it is generally considered to be of Gond- wanan origin (Breure, 1979; Solem, 1998). Orthalicoid Whether or not the terrestrial snail superfamily diversity is greatest in the Americas (primarily the Orthalicoidea [= Bulimulacea/oidea] occurs in Africa Neotropics), with smaller radiations also occurring has remained an unresolved question. Globally, the in southern Australia and New Zealand/Melanesia superfamily exhibits a southern continental distribu- (Breure, 1979). However, Pilsbry (1902: ix) stated that ‘there is yet no trace of Bulimulidae in Africa’ and recent authors have maintained this opinion (Solem, *Corresponding author. E-mail: [email protected] 1959, 1998; Peake, 1978; Breure, 1979; Tillier, 1989;

Schileyko, 1999a). Nonetheless, in the intervening idae. In general, the possibility that Prestonella may period, two somewhat enigmatic African taxa, truly be an orthalicoid (bulimulid) taxon has been although not currently considered bulimulids, have rejected in most subsequent studies, or at least not been referred to the orthalicoid family Bulimulidae, considered due to insufficient information (Peake, namely Aillya Odhner, 1927 and Prestonella Connolly, 1978; Breure, 1979; Solem, 1998; Schileyko, 1999a). 1929. Vaught (1989) was the only recent author to main- Despite its aberrant semislug form, Odhner (1927) tain Prestonella with orthalicoid taxa. Ultimately, believed that his new genus Aillya exhibited ana- this confusion stems from one fact, as succinctly tomical similarity with the Bulimulidae and Con- stated by Solem (1959: 123), that ‘determination of nolly (1929), when proposing the new genus its [Prestonella] affinities must await dissection of Prestonella, likewise referred the taxon to this the animal’. Downloaded from https://academic.oup.com/biolinnean/article/96/1/203/2448044 by guest on 27 September 2021 family, basing his decision on unpublished anatomi- Clearly, the original material of Prestonella studied cal observations made by Hugh Watson. Thiele by Connolly (and Watson) included preserved soft (1931) maintained the bulimulid affinities of both parts but, subsequently, very little material of any of Prestonella and Aillya, referring them to the subfam- the three described species has been collected (and ily Amphibuliminae, and Zilch (1959–60) was of a none at all in the last 50 years) and thus no preserved similar opinion, although he raised the Amphibulimi- material has been available for anatomical study. nae to the rank of family, within the superfamily Resolution of the long-standing problem concerning ‘Bulimulacea’ [= Orthalicoidea]. Van Mol (1978), the phylogenetic affinity of Prestonella therefore however, subsequently investigated the internal hinged upon finding further living specimens so that morphology of the type species of Aillya in greater the anatomy could be re-examined. detail, and found it to differ significantly from that Recent targeted field work has resulted in the of bulimulids. He suggested that Aillya represented rediscovery of living colonies of two Prestonella a family of its own, as proposed by Baker (1955), but species, including the type species, enabling us to postulated affinity with the Helicarionidae and study the anatomy of the soft parts in detail. In the Urocyclidae, rather than with the elasmognaths present study, we discuss the implications of our (Succineidae and Athoracophoridae), as suggested observations in relation to the systematic affinities by Baker (1955). Some recent authors have gone one of Prestonella. In addition, we obtained DNA step further, placing Aillya in its own superfamily sequence data from the nuclear rRNA gene cluster (Nordsieck, 1986; Vaught, 1989; Bouchet et al., 2005) and included this in a re-estimation of the sty- or infra-order (Schileyko, 1999a). In any event, the lommatophoran molecular phylogeny performed by fact that Aillya is neither a bulimulid nor of orthali- Wade, Mordan & Naggs (2006). Because Prestonella coid stock now appears to be well established. has been linked to the Urocyclidae, but this family Although Connolly (1929) stated that Hugh was not represented in the Wade et al. (2006) analy- Watson was to publish his anatomical observations sis, we added a sample of urocyclid taxa (represent- on Prestonella, this did not happen and, aside from ing three subfamilies) to the data set, as well as a Connolly’s comments on the jaw and illustration of representative of the Australian bulimulid genus the radula teeth (Connolly, 1939), neither comprising Bothriembryon Pilsbry, 1894, which proved to be particularly informative phylogenetic indicators, the morphologically similar to Prestonella. Attempts to anatomy of Prestonella has remained undocumented. obtain material of the rare West African genus Its familial affinities have therefore never been con- Aillya were not successful. The few specimens avail- clusively established and all subsequent statements able have not proved suitable for DNA extraction in this regard have lacked solid foundation. The (A. De Winter, pers. comm.). In the light of this new chequered and inconclusive taxonomic history of morphological and molecular data, we re-evaluate the genus is detailed elsewhere (Herbert, 2007). In the phylogenetic relationships of Prestonella,inan summary, the most recent, albeit speculative, attempt to establish whether or not it exhibits affin- hypotheses proposed are those of Bruggen (1977, ity with the Orthalicoidea: Bulimulidae, and thus 1978, 1986) and Schileyko (1999a). Bruggen sug- determine whether or not this Gondwanan group is gested that Prestonella may in fact represent a dis- represented on the African continent. tinct family, the Prestonellidae, perhaps derived from ‘the cosmopolitan succineids or the urocyclids (cf. Aillyidae)’, whereas Schileyko (1999a), pre- INSTITUTIONAL ABBREVIATIONS sumably in the light of Connolly’s discussion of BMNH, Natural History Museum, London; NMSA, Prestonella together with Aillya, considered that Natal Museum, Pietermaritzburg, South Africa; SAM, Prestonella might be to some degree related to South African Museum, Cape Town, South Africa; Aillya, and thus perhaps also referable to the Ailly- WAM, Western Australian Museum, Perth.

MATERIAL AND METHODS dilute NaOH, rinsed in distilled water, dehydrated in MATERIALS ethanol and air-dried on stubs with double-sided carbon tape. Shells and radulae for scanning electron In an attempt to obtain material for both morphologi- microscopy examination were coated with gold- cal and molecular study, the senior author, in colla- palladium and examined at low accelerating voltage boration with Mary Cole (East London Museum), (5–10 kv) in a Hitachi S-570 scanning electron undertook a ﬁeld trip to the Somerset East and microscope. Cradock area, Eastern Cape, South Africa, in order to search for specimens of Prestonella bowkeri (Sowerby, 1890) and Prestonella nuptialis (Melvill & Ponsonby, MOLECULAR METHODS

1894). The sites selected focused on localities listed Downloaded from https://academic.oup.com/biolinnean/article/96/1/203/2448044 by guest on 27 September 2021 Laboratory techniques for these species by Connolly (1939). Material of P. Total genomic DNA was extracted from approximately bowkeri was collected at the type locality, Glen Avon 20 mg of muscle tissue from the posterior portion of Falls, Somerset East, Eastern Cape (32°40.887′S: the foot, by means of the Qiagen DNeasy Tissue Kit. 25°38.13′E), and that of P. nuptialis at the summit of Two sets of polymerase chain reaction (PCR) primers Elandsberg Mountain, Cradock area, Eastern Cape originally described by Wade & Mordan (2000) (32°07.488′S: 25°42.348′E). Living specimens were (i.e. LSU1–LSU3 and LSU2–LSU4) were used for drowned overnight in sealed tubes and preserved in PCR ampliﬁcation and DNA sequencing of partial 75% ethanol for anatomical dissection. For molecular 5.8S, complete internal transcribed spacer (ITS)2 and sequencing, small fragments of the foot were excised partial 28S rRNA genes for both P. bowkeri and and preserved in 99% ethanol, before the specimens P. nuptialis, as well as the four urocyclid species were drowned. Material of further species included and Bothriembryon indutus. The PCR products were in the molecular analysis for comparative pur- sequenced directly in both directions using the ABI poses was obtained from the Natal Museum mol- PRISM Big Dye Terminator v.3.0 Ready Reaction luscan tissue bank [Urocyclidae: Sheldonia poeppigii Cycle Sequencing Kit, and fractionated on an Applied (Pfeiffer, 1846), Sheldonia phaedima (Melvill & Biosystems 3100 Genetic Analyser. Ponsonby, 1892), Gymnarion sp., and Elisolimax ﬂa- vescens (Keferstein, 1866)] and the Western Austra- lian Museum [Bothriembryon indutus (Menke, 1843)]. Data set assembly Specimen details are provided in Table 1. We obtained the full DNA sequence alignment pre- All dissections were performed under a Wild M4 sented by Wade et al. (2006) from the senior author. dissecting microscope with drawing tube. Radulae The data set comprised 167 taxa and 823 bp of unam- were extracted by maceration of the buccal mass in biguously aligned nuclear ribosomal DNA sequence

Table 1. Details of specimens, provenance, collectors and museum voucher numbers for tissue samples sequenced during the present study

Family Species Provenance Collector Voucher number

? Prestonella bowkeri Glen Avon, Somerset East, D. Herbert NMSA: V9816 (Sowerby, 1890) South Africa ? Prestonella nuptialis Cradock area, South D. Herbert NMSA: V9778 (Melvill & Ponsonby, Africa 1894) Bulimulidae Bothriembryon indutus Walyunga National Park, C. Whisson & WAM: S33057 (Menke, 1843) Western Australia H. Morrison Urocyclidae: Urocyclinae Elisolimax ﬂavescens Pietermaritzburg, South D. Herbert NMSA: W2297 (Keferstein, 1866) Africa & L. Davis Urocyclidae: Gymnarion sp. Graskop, South Africa H. Musgrave NMSA: W2798 Gymnarioninae (?introduced) (?Gymnarionidae) Urocyclidae: Sheldoniinae Sheldonia poeppigii Pietermaritzburg, South D. Herbert NMSA: W4882 (Pfeiffer, 1846) Africa & L. Davis Urocyclidae: Sheldoniinae Sheldonia phaedima Dargle, South Africa D. Herbert NMSA: W1781 (Melvill & Ponsonby, 1892)

© 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 96, 203–221 206 D. G. HERBERT and A. MITCHELL data. Our seven new sequences (GenBank accession set to derive a smaller data set focused on taxa numbers EU622016–EU622023) were in the size relevant to the resolution of question under investi- range 929–1343 bp. The 929-bp sequence obtained for gation. The superfamilies selected for reduction S. poeppigii is missing data from the ITS region; represented stylommatophoran clades shown to be however, this region was excluded by Wade et al. unrelated to Prestonella in our NJ analysis and which (2006) for all species because it was too variable to exhibit a morphology substantially different to that of align across all taxa. Our seven new sequences were Prestonella (‘achatinoids’, Arionoidea, Clausilioidea, unambiguously aligned to the Wade et al. (2006) data Helicoidea, and Orthurethra). The taxa chosen to set by hand using BioEdit (Hall, 1999), to give an represent these clades are shown in Table 2. With the alignment comprising 174 taxa and 823 bp. Guided by exception of these reduced clades, the data set com- the results of an initial neighbour-joining (NJ) analy- prised all taxa included in the analysis of Wade et al. Downloaded from https://academic.oup.com/biolinnean/article/96/1/203/2448044 by guest on 27 September 2021 sis of the 174 taxa data set, the results of Wade et al. (2006). This smaller data set comprised 83 taxa. (2006) and morphology-based comparison, we then Because our data set was largely a subset of that of selected 83 taxa for inclusion in a second data set of Wade et al. (2006), and we wished to compare our a size amenable to Bayesian analysis (see below). results with theirs directly, we followed their analysis in selecting the GTR + gamma model. Tree space was Phylogenetic analyses explored using four chains of Markov Chain Monte An initial NJ analysis was performed on the 174 taxa Carlo analysis for 3 million generations, sampling data set using PAUP*4.0b10 (Swofford, 2000) under every 2000 generations. A consensus tree was built the LogDet model. Parameters of the General Time using the last 1000 trees (burn-in = 501). Node Reversal (GTR) model [the model of sequence evolu- support was considered to be strong at posterior Ն tion used by Wade et al. (2006)] were estimated for probability (PP) values 0.98 [bootstrap percentage Ն this initial NJ tree, then ﬁxed and a full NJ analysis, (BP) 90%] and moderate at PP values of 0.95 (BP including a 1000-replicate bootstrap analysis, per- circa 80%). formed by estimating distances under maximum like- lihood using the GTR model. RESULTS Bayesian analyses were performed using MrBayes, version 3.1 (Huelsenbeck & Ronquist, 2001). Prelimi- TAXONOMIC SUMMARY nary analysis using the 174 taxa data set indicated Genus Prestonella Connolly, 1929 that it would take approximately 250 days of analysis Prestonella Connolly, 1929: 232. Type species (original for convergence. Prior to running a full Bayesian designation) ‘Buliminus bowkeri’[sic] Sowerby, analysis, we therefore sub-sampled the 174 taxa data 1890.

Table 2. Higher level clades identiﬁed by Wade et al. (2006) and the exemplar species selected to represent them in our Bayesian analysis of the reduced data set

Clade Family Species

Achatinoid Achatinidae Archachatina marginata (Swainson, 1821) Coeliaxidae Coeliaxis blandi (Pfeiffer, 1852) Subulinidae Subulina striatella (Rang, 1831) Streptaxidae Gonaxis quadrilateralis (Preston, 1910) Helicoidea Helicidae Helix pomatia Linnaeus, 1758 Bradybaenidae Bradybaena similaris (Férussac, 1821) Hygromiidae Cernuella virgata (Da Costa, 1778) Camaenidae Polydontes undulata (Férussac, 1821) Clausilioidea Clausiliidae Clausilia bidentata (Ström, 1765) Clausiliidae Nenia tridens (Schweigger, 1820) Arionoidea Arionidae Arion hortensis Férussac, 1819 Philomycidae Meghimatium bilineatum (Benson, 1842) Orthurethra Partulidae Partula suturalis Pfeiffer, 1855 Cerastidae Cerastus schweinfurthi (Martens, 1895) Enidae Mastus pupa (Bruguière, 1792) Vertiginidae Vertigo antivertigo (Draparnaud, 1801)

Specimen details provided by Wade et al. (2006).

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Figure 1. Shells of the three Prestonella species. A, Prestonella bowkeri, lectotype of Bulimus (Mesembrinus?) bowkeri Sowerby, 1890, Glen Avon, Somerset East, Eastern Cape, South Africa, length 19.5 mm (BMNH 89.11.4.9). B, Prestonella nuptialis, Cradock, Eastern Cape, South Africa, length 13.3 mm (NMSA 1592). C, Prestonella quadingensis, paratype, Quthing, Lesotho, length 11.4 mm (BMNH, 1937.12.30.7079-80).

Figure 2. Distribution of Prestonella species. The genus is endemic to southern Africa, occurring along the southern edge of the Great Escarpment in South Africa and in Lesotho (1000–1750 m a.s.l.); circles, Prestonella bowkeri; triangles, Prestonella nuptialis; square, Prestonella quadingensis; contour at 1000 m.

The genus Prestonella is endemic to southern Africa (Herbert, 2007). The adult shell is succineiform to and is represented by three described species (Fig. 1), elongate-bulimiform and has few distinctive teleo- P. bowkeri (Sowerby, 1890) (Fig. 1A), P. nuptialis conch features, no doubt contributing to the confusion (Melvill & Ponsonby, 1894) (Fig. 1B), and P. quadin- regarding the familial affinities of the genus. gensis Connolly, 1929 (Fig. 1C), occurring in south- western Lesotho and along the southern edge of the Great Escarpment in South Africa (1000–1750 m MORPHOLOGY a.s.l.) (Fig. 2). A detailed taxonomic revision of the Our discussion of the morphology of Prestonella genus has been completed and is provided elsewhere focuses on the type species, P. bowkeri, with compara-

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Figure 3. Prestonella bowkeri (Sowerby, 1890), protoconch, Nuweveldberg, Beaufort West (SAM, A8171). A, scale bar = 0.5 mm. B, scale bar = 0.2 mm. tive notes on P. nuptialis where pertinent. Further details of the morphology of P. nuptialis are provided by Herbert (2007). No preserved material of P. quadingensis is available.

Protoconch (Fig. 3): Diameter somewhat variable, 1.6–2.2 mm, comprising approximately 2.0 whorls; the first whorl is set at a pronounced angle, rising to the start of the second whorl which is initially flat- sided with an almost ridge-like apical angle, becoming more evenly convex toward the end of the whorl; sculptured with widely spaced collabral axial riblets, approximately 0.20 mm apart, riblets approximately one-fifth the width of their intervals; intervals sculptured by numerous close-set spiral striae (Fig. 3B); riblets usually straight, but somewhat wavy in occa- Figure 4. Prestonella bowkeri (Sowerby, 1890), living sional individuals; strength of sculpture variable, specimen, Glen Avon Falls, Somerset East, shell length almost obsolete in some specimens, and frequently 18.0 mm (NMSA V9816). badly eroded; change from embryonic to adult sculpture moderately abrupt and protoconch-teleoconch perpendicular to mid-line in central region, curving junction, thus relatively distinct. slightly backwards or forwards laterally (probably External features (Fig. 4): Head-foot more or less uni- related to specimen mounting), but not V-shaped; formly pale apricot-buff to yellow (dark brown in P. approximate formula R + L(10–11) + M(±30), but nuptialis); tentacles well-developed, but not particu- tooth form smoothly intergrading across radula width larly long, withdrawn by progressive inversion from with no clear delimitation between lateral and mar- tip; genital aperture ventral and slightly posterior to ginal series (Fig. 5A); anterior radula teeth worn right ommatophore; peripodial groove not evident down to mere stubs. (holopodous); hind end of foot blunt ending, with Rachidian with a single, trigonal cusp (Fig. 5B, C), neither a caudal pit nor appendage; sole not obviously somewhat smaller than those of innermost lateral subdivided longitudinally; no mantle lobes extending teeth, cusp base (anterior portion of tooth) truncate over shell. and sometimes indented medially; tooth base-plate symmetrical with a raised tubercle at posterior angle Radula (Fig. 5): Dentition extremely fine; radula up on both sides, evidently interlocking with correspond- to 3.4 mm in length and 1.1 mm wide, with up to 230 ing indentations in the cusp base of rachidian of transverse rows of teeth, each with up to 100 teeth, the next posterior row, in peg-and-pocket manner depending on the size of the animal; rows more or less (Fig. 5C). Innermost lateral teeth (numbers 1–10) also

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Figure 5. Prestonella bowkeri (Sowerby, 1890), radula (NMSA V9816). A, central ﬁeld and inner marginals, scale bar = 50 mm. B, rachidian (R) and inner laterals, scale bar = 25 mm. C, rachidian (R), scale bar = 15 mm. D, marginals, scale bar = 50 mm. E, inner marginals, scale bar = 20 mm. F, outer marginals, scale bar = 20 mm. monocuspid (Fig. 5A, B); shape of cusp somewhat approximately tooth 10 (start of latero-marginal variable and cusp frequently not symmetrical about series), the outer edge of the cusp buds off a basal its mid-line, outer edge slightly longer than inner one; ectocone (Fig. 5A, E); a somewhat smaller endocone base of cusp more rounded with an irregular pattern buds off the inner cusp margin from tooth number of indentations; base-plate asymmetrical, a distinct 13–14 outward; endocone nearer to mesocone apex tubercle present only at the outer posterior angle. At than ectocone and smaller; endocone and ectocone

© 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 96, 203–221 210 D. G. HERBERT and A. MITCHELL becoming progressively larger relative to mesocone (stegognathous cf. Hausdorf, 1998), translucent cor- toward margin of radula, but ectocone remaining neous yellow; central plate broadly triangular; seven more basal than endocone. Teeth progressively to 12 plates on either side of central one, elongate and smaller from tooth 30 outward (outer marginal series) with more or less parallel sides; inner of two plates and more widely spaced both between and within overlapping its outer neighbour. Details of plate mor- rows (Fig. 5D, F), the cusps more elongate and phology are more evident when viewed by transmit- pointed, generally quite variable in form; ectocone ted light (Fig. 6A). frequently biﬁd or even triﬁd. The radula of P. nuptialis is essentially similar Pulmonary anatomy (Fig. 7): Typically sigmure- (Herbert, 2007), but with fewer teeth per row and a throus. Kidney short, triangular, cardiac edge of Downloaded from https://academic.oup.com/biolinnean/article/96/1/203/2448044 by guest on 27 September 2021 more obvious distinction between the laterals and similar length to pericardium; renal pore at anterior marginals, formula R + L15 + M(20–30). angle, opening into the primary ureter, which runs posteriorly along the right side of kidney to the apex Jaw (Fig. 6): Crescent-shaped to almost U-shaped, of pulmonary cavity, and then, turning anteriorly to composed of fused, imbricating chitinous plates form the secondary ureter, which runs forward along-

Figure 6. Prestonella bowkeri (Sowerby, 1890), jaw (NMSA V9816). A, drawing of light microscope preparation of entire jaw; B, SEM of left half of jaw, scale bar = 100 mm.

Figure 7. Prestonella bowkeri (Sowerby, 1890), pulmonary anatomy (NMSA V9816). an, anus; aur, auricle; int, intestine; kid, kidney; pc, pericardium; pn, pneumostome; pu, primary ureter; pv, pulmonary vein; rec, rectum; su, secondary ureter; urp, ureteric pore; ven, ventricle.

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Figure 8. Prestonella bowkeri (Sowerby, 1890), reproductive anatomy (NMSA V9816). A, entire genital tract. B, apical portion of flagellum and penial retractor muscle. C, penis with sheath partially removed to reveal kinked mid region. D, transverse section of penis. E, transverse section of epiphallus. alb, albumen gland; bc, bursa copulatrix; bcd, bursa copulatrix duct; el, epiphallus lumen; eph, epiphallus; flg, flagellum; ga, genital atrium; hd, hermaphrodite duct; od, oviduct; ot, ovotestis; pen, penis; pl, penis lumen; pr, prostate gland; prm, penial retractor muscle; ps, penis sheath; vag, vagina; vd, vas deferens. side rectum to open at edge of the pneumostome; sac, accessory glands, appendages, and stimulatory secondary ureter closed for most of its length except organs). Penial complex long and tapering (up to in the distal region, just inside the pneumostome, 18 mm in total length), extending far back into the where it opens to form a V-shaped notch. Pulmonary body cavity and comprising a relatively short penis, a vein distinct, with side branches on both sides, those longer, more slender epiphallus and an even more on the right, between the pulmonary vein and the slender flagellum, coiled into a number of loops. Junc- secondary ureter, more numerous; on the left, a sec- tions between these elements are not well differ- ondary vein runs parallel to the main pulmonary entiated. Penis itself somewhat variable in shape, vein. Pulmonary anatomy of P. nuptialis is essentially approximately 2.2 mm in length and approximately the same, but the kidney is somewhat broader and 1.0 mm at its widest; basal portion enclosed in thin the main pulmonary vein more oblique on account of sheath, inside which it may be weakly kinked or the less elongate shell (Herbert, 2007). folded into an S-shape (Fig. 8C), embedded in much connective tissue; a cuff-like sheath at the penis base Genital system (Fig. 8): Distal reproductive tract rela- not present. Penis wall muscular, the lumen with five tively simple, lacking auxiliary structures (e.g. dart irregularly pleated, longitudinal ridges (pilasters),

© 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 96, 203–221 212 D. G. HERBERT and A. MITCHELL lined by a columnar epithelium (Fig. 8D), one ridge shown) and only results of GTR analyses are pre- usually larger than the others, all becoming less sented. Both NJ and Bayesian analyses (Fig. 9) of distinct distally at junction with epiphallus. Epiphal- the nuclear DNA sequence data identified higher- lus subcylindrical, 3–5 mm long and approximately level taxa (superfamilies and families) identical to 0.5 mm wide, tapering towards its junction with those found by Wade et al. (2006), likewise grouping flagellum, at which point the vas deferens inserts these in a monophyletic Stylommatophora. However, (marked only by a slight change in thickness); inter- as with the tree of Wade et al. (2006), little renal wall with four smooth longitudinal ridges creating solution was evident at deeper levels within the an X-shaped lumen (Fig. 8E), a layer of rounded sub- non-achatinoid Stylommatophora (NJ BP < 50% and epithelial cells separating this from the outer muscle PP < 0.50 at many nodes). With regard to the place- layer. Flagellum continues posteriorly as a thin, ment of Prestonella, both analyses gave identical Downloaded from https://academic.oup.com/biolinnean/article/96/1/203/2448044 by guest on 27 September 2021 blind-ending tube, twisting through several coils and results, grouping it within the Orthalicoidea and in terminating in an extremely short penial retractor the same position. The two Prestonella species group muscle (Fig. 8B), which attaches to body wall (dia- as sister taxa (BP = 100%; PP = 1.0) and the genus phragm) beneath the posterior quarter of pulmonary itself is sister taxon to a clade comprising the Aus- cavity. Vas deferens very slender and thin-walled, not tralasian genera Placostylus and Bothriembryon.The visible without close scrutiny; commencing at base of level of support for the monophyly of Prestonella + prostatic portion of the sperm-oviduct and initially (Placostylus + Bothriembryon) was high (BP = 100%; running within wall of free oviduct, emerging for a PP = 1.0), but that for the Australasian clade short distance in the angle between the vagina and (Placostylus + Bothriembryon) was lower (BP = 81%; penis base, then merging with penis wall and con- PP = 0.66). tinuing, embedded in outer layers of the penis and The South American bulimulid taxa (Bulimulus and epiphallus wall, to its insertion at the junction of the Drymaeus) cluster as a well supported (BP = 100%; epiphallus and flagellum. Sperm-oviduct approxi- PP = 1.0) but internally unresolved clade, which forms mately 10.0 mm in length, oviducal portion thicker the sister group to the African-Australasian taxa than prostatic portion, but distinction between the (BP = 58%; PP = 0.99). The Neotropical amphibulimid two is not clear. Duct of bursa copulatrix arises from genus Gaeotis, branches off basally, completing the free oviduct close to its junction with the genital a moderately well-supported orthalicoid clade atrium and the vagina is thus very short; duct as (BP = 87%; PP = 1.0) [but excluding Cerion (Cerion- long or longer than sperm-oviduct (approximately idae), which Nordsieck (1986) and Bouchet et al. (2005) 12.0 mm) and relatively wide; runs closely applied to referred to the Orthalicoidea]. As in the analysis of sperm-oviduct and terminates in ovate bursa copula- Wade et al. (2006), the Orthalicoidea in turn clusters trix lying ventral and posterior to the kidney and with a weakly supported clade (BP < 50%; PP = 0.74) pericardium. Genital atrium not enlarged and with a comprising the reputedly acavid genus Leucotaenius single opening to exterior. Retractor muscle of optic and a well-supported (BP = 100%; PP = 1.0) elasmog- tentacle passes between penis and sperm-oviduct. nath clade (Succinea and Athoracophorus). However, Ovotestis racemose, embedded in the upper whorl node support for the sister group relationship of digestive gland; hermaphrodite duct initially thin- between the Orthalicoidea and (Leucotaenius + walled and straight, becoming thicker and highly (Athoracophorus + Succinea)) was low [BP < 50%; convoluted before narrowing again prior to its junc- PP = 0.72 in our tree, PP = 0.48 in the tree of Wade tion with talon (fertilization pouch and receptaculum et al. (2006) (C. Wade, pers com)], again reflecting lack seminis). Talon comprising a small swelling on of resolution at deeper levels. surface of albumen gland with a narrow duct extend- The four urocyclid taxa included for the first ing into tissues of the albumen gland to join with the time in the nuclear DNA data set, belonging to albumen gland duct and the start of the sperm- three genera (Elisolimax, Gymnarion,andShel- oviduct (not examined in detail). donia), cluster together as a monophyletic group The genital system of P. nuptialis differs little from (BP < 50%; PP = 0.98) within the Limacoidea s.l., that described for P. bowkeri. Its penial complex and well separated from Prestonella. The three genera sperm-oviduct are somewhat less elongate and the selected belong to three different urocyclid sub- vas deferens follows a more twisted course within the families (Urocyclinae, Gymnarioninae, and Sheldo- penis wall (Herbert, 2007). niinae) and reflect the diversity evident within the family. This urocyclid clade, however, lies within the Helicarionidae, rendering the latter paraphyl- MOLECULAR DATA etic. Although the Limacoidea s.l., is well supported The NJ analyses performed under LogDet and GTR (BP = 86%; PP = 1.0), internal resolution at deeper models yielded almost identical results (trees not levels is poor.

Carychium tridentatum Laemodonta sp. 1.0 Non-stylommatophoran 0.94 Aplysia punctata Siphonaria pectinata Pulmonata 0.84 Laevicaulis alte (outgroup) 0.99 Atopos australis Melampus luteus Gonaxis quadrilateralis 0.99 1.0 Archachatina marginata Achatinoid clade 0.55 Coeliaxis blandii Subulina striatella Acavus phoenix Testacella scutulum Megalobulimus oblongus Corilla adamsi Caryodes dufresnii 1.0 Dorcasia alexandri Trigonephrus globulus Dorcasiidae Chlamydephorus burnupi 0.99 Downloaded from https://academic.oup.com/biolinnean/article/96/1/203/2448044 by guest on 27 September 2021 1.0Rhytida stephenensis Schizoglossa sp. Rhytididae 1.0 Otoconcha dimidiata 0.77 Laoma sp. Punctoidea Suteria ide Cerion sp. 0.63 1.0 Anguispira alternata Discidae 0.76 Discus rotundatus Euglandina rosea 0.74 Haplotrema vancouverense 0.72 Polydontes undulata 1.0 Bradybaena similaris 1.0 1.0 Helicoidea 0.65 Cernuella virgata Helix pomatia Leucotaenius proctori 0.74 1.0 Athoracophorus bitentaculatus 1.0 Succinea sp. 0.72 Succinea striata Elasmognatha Gaeotis nigrolineata Amphibulimidae 1.0 Bulimulus guadalupensis 1.0 Bulimulus sporadicus 0.99 Drymaeus discrepans 1.0 Prestonella bowkeri Orthalicoidea 1.0 Prestonella nuptialis Bulimulidae s.l. 0.66 Bothriembryon indutus 0.67 Placostylus ambagiosus Placostylus eddystonensis 0.99 Clausilia bidentata Nenia tridens Clausiliidae 1.0 Solatopupa similis 0.87 1.0Chondrina avenacea 0.74 Chondrina clienta Partula suturalis 1.0 Orthurethra 0.74Mastus pupa 0.70 Cerastus schweinfurthi 0.89 Vertigo antivergo 0.84 Arion hortensis Meghimatium bilineatum Arionoidea Deroceras reticulatum 0.86 Tandonia budapestensis Vitrea crystallina 0.64 Limacoidea s.l. 1.0 Plutonia laxata Vitrina pellucida 1.0 Oxychilus cellarius 0.78Oxychilus alliarius 1.0 Oxychilus helveticus 1.0 Trochomorpha pallens Videna gouldiana sp. 0.92 Hiona 1.0 Euconulus fulvus 0.90 Louisia barclayi 1.0 Asperitas inquinata 1.0 Craterodiscus pricei 1.0 Cryptozona bistrialis Ratnadvipia sp. Ariophantidae Euplecta gardeneri 1.0Harmogenanina argentea 0.74 Fastosarion brazieri Helicarionidae 0.95 Plegma caelatura Helicarionoidea Rhysotina hepatizon 0.99 0.68 Elisolimax flavescens 0.98 Gymnarion sp. Urocyclidae 0.1 1.0Sheldonia phaedima Sheldonia poeppigii

Figure 9. Bayesian tree for the Stylommatophora based on the reduced data set of 83 taxa, and 823 unambiguously aligned nucleotide sites of nuclear DNA. Posterior probabilities (‘clade credibility values’; PP) shown to the right of nodes (polytomies = PP < 0.50). Taxa highlighted in bold are those for which new sequence data has been obtained and which have been added to the data set of Wade et al. (2006).

DISCUSSION of the families with which it has been tentatively LACK OF AFFINITY WITH SUCCINEIDAE, associated by recent authors (Succineidae, Urocy- UROCYCLIDAE, AND AILLYIDAE clidae, Aillyidae). The Succineidae differ in having a The morphology of the soft parts of Prestonella is heterurethrous rather than sigmurethrous pulmo- clearly not consistent with a close relationship to any nary anatomy, the jaw is elasmognathous, the method

© 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 96, 203–221 214 D. G. HERBERT and A. MITCHELL of optic tentacle retraction is by contraction and Prestonella within the Orthalicoidea, in a strongly partial inversion (Burch & Patterson, 1969) rather supported (BP = 100%; PP = 1.0) sister-group relation- than by progressive inversion from the tip, the foot ship with an Australasian clade comprising Bothri- is aulacopod, and its sole longitudinally tripartite embryon and Placostylus. Although this was initially (Solem, 1978; Barker, 2001). Any similarity between surprising, in view of recent speculation concerning Prestonella and succineids therefore concerns only the the affinities of Prestonella and the supposed absence overall shell physiognomy. Although the molecular of orthalicoid taxa in Africa, it provided an excellent data suggest a distant relationship between the clade pointer for further morphological comparison, which, containing Succinea and that proving to contain as we discuss below, turns out to be fully congruent Prestonella, this relationship is weakly supported with the molecular data. (BP < 50%; PP = 0.72), extremely deep, and is sub- Some of the most informative morphological char- Downloaded from https://academic.oup.com/biolinnean/article/96/1/203/2448044 by guest on 27 September 2021 tended by very long branches (discussed further acters available for phylogenetic reconstruction of below). stylommatophoran relationships are thought to be Members of the Urocyclidae have an aulacopod those of the genital tract, of which the distal com- foot with a distinct peripodial groove, a conspicuous ponent is more varied and more easily studied, and caudal pit and associated appendage, a longitudinally therefore better known. A considerable amount of tripartite foot sole, well-developed mantle lobes, elon- information concerning the distal genital tract of gate marginal radula teeth, an oxygnath jaw, and orthalicoid species is available (Pilsbry, 1946a, b; frequently have accessory structures associated with Kondo, 1948; Odhner, 1951; Van Mol, 1971; Turner the distal genitalia [epiphallic caecum, penial gland, & Clench, 1972; Breure, 1978, 1979; Breure & sarcobelum, atrial diverticulum, oviducal (capsular) Coppois, 1978; Parodiz, 1979; Breure & Eskens, gland] (Van Mol, 1970; Van Goethem, 1977; Hausdorf, 1981; Breure & Schouten, 1985; Kershaw, 1986; 1998). In addition, the urocyclid shell bears almost no Salgado & Leme, 2000). It is clear that the relatively resemblance to that of Prestonella, being subconical to simple structure of the genitalia of Prestonella has lenticular, or reduced to an internal vestige in limaci- much in common with that of certain orthalicoid form genera. The inclusion of urocyclid taxa in our taxa. Of particular significance are: (1) the lack of molecular data set, as expected from morphological accessory structures (e.g. dart sac, accessory glands, features (Hausdorf, 1998), saw them cluster as a appendages and stimulatory organs); (2) the extreme monophyletic entity (well supported in the Bayesian length of the phallus complex; (3) the limited differ- analysis (PP = 0.98), but not so in the NJ analysis entiation between the penis, epiphallus and flagel- BP < 50%) within the Limacoidea s.l. and they are lum; (4) the short penial retractor muscle which clearly of helicarionoid affinity. Indeed, they cluster inserts at the flagellum tip; (5) the extremely thin within the Helicarionidae, suggesting that the latter, vas deferens which is for the most part embedded in as currently interpreted, is not monophyletic. Recent the wall of the penis and epiphallus; and (6) the long analysis of mitochondrial DNA sequence data has also bursa copulatrix duct, which lacks a diverticulum. supported the helicarionoid affinities of the Urocyl- With strong evidence in support of affinity between idae (Hyman, Ho & Jermiin, 2007). Prestonella and the Orthalicoidea emerging from both The Aillyidae differ from Prestonella in terms of the molecular data and reproductive tract morphology, we shape and orientation of the kidney (which resembles can move on to consider how other morphological that of the Succineidae), have a far less elongate features fit in with this hypothesis. Although signifi- penial complex, a short bursa copulatrix duct and cant primarily at rather deeper levels and perhaps an expanded vagina, well-developed mantle lobes, and ancestral character (Wade, Mordan & Clarke, 2001), a more auriform shell (Van Mol, 1978). However, the sigmurethrous nature of the pulmonary cavity of because molecular data are not available for this Prestonella, with the secondary ureter closed for taxon, its relationships within the Stylommatophora almost its entire length, is similar to that of orthali- remain poorly understood. From a morphological per- coid taxa, as is the holopodous foot and undivided foot spective, therefore, Prestonella must either belong to sole. More informative perhaps is the form of the another sigmurethrous family, or a family of its own protoconch. In this regard, the somewhat elevated (Bruggen, 1978). protoconch of Prestonella, with the first whorl set at an angle (tilted) and the second whorl flat-sided with an acute shoulder and a deep suture, is distinctive. In REFERRAL TO THE ORTHALICOIDEA many sigmurethrous families, the embryonic shell is The results obtained through the inclusion of nuclear less elevated, more globose, and has a shallow suture, DNA sequence data from Prestonella in our re- but protoconchs closely resembling that of Prestonella estimation of the stylommatophoran molecular phy- have been illustrated for several bulimulid genera logeny published by Wade et al. (2006) clearly place (Bothriembryon, Naesiotus, Neopetraeus, Scutalus,

© 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 96, 203–221 PRESTONELLA: AN AFRICAN ORTHALICOID SNAIL 215 and Thaumastus; Breure, 1977, 1978, 1979; Breure of Prestonella is typical of orthalicoid taxa and & Coppois, 1978; Kershaw, 1986). In terms of proto- resembles that of bulimulid species illustrated by conch microsculpture, Breure has illustrated a Breure & Schouten (1985), in particular species of number of different forms, all essentially variations Drymaeus (Breure & Eskens, 1981). on a theme with dominant axial sculpture. That of It is clear that these additional morphological char- Naesiotus, comprising widely spaced axial riblets with acters are consistent with the genital anatomy and fine, close-set spiral threads in their intervals, is molecular data. There is thus evidence available from strikingly similar to the protoconch of P. bowkeri nuclear DNA and a suite of independent morphologi- (compare Fig. 3 with Breure, 1978: pl. 12, fig. 2 and cal characters indicating that Prestonella exhibits Breure & Coppois, 1978: figs 1–6). close affinity with the Orthalicoidea and in particular Unlike its prosobranch counterpart, the pulmonate with the Bulimulidae s.l., and we propose to refer it to Downloaded from https://academic.oup.com/biolinnean/article/96/1/203/2448044 by guest on 27 September 2021 radula has proved to be of limited value to date in this family. The suggestion that it represents a dis- providing phylogenetically informative characters. On crete family-level entity itself (Bruggen, 1978, 1985) the one hand, it appears to be relatively conservative, is not supported (unless further phylogenetic work yet, on the other, it appears to show considerable indicates the need to subdivide the Bulimulidae s.l. adaptive plasticity related to diet (e.g. the occurrence into smaller family-level entities). Hugh Watson’s of a beloglossan radula in many unrelated carnivo- unpublished observations that led Connolly (1929) to rous families). In comparison with phytophagous Sty- refer his new genus to the Bulimulidae were evidently lommatophora, the radula of Prestonella is unusual in sound. Interestingly, Sowerby (1890), in his original that the rachidian and inner lateral teeth are mono- description of the species that Connolly (1929) subse- cuspid rather than the more usual bi- or tri-cuspid. quently chose as the genotype of Prestonella, referred Breure & Gittenberger (1982) have identified such it to Bulimus (Mesembrinus?) [= Drymaeus]. Although structural modification as being another example of he did not specify his reasons, clearly he had inter- diet-related adaptation, in this case associated with a preted the significant conchological features correctly. rock-scraping mode of feeding. Although no observa- This represents the first well-supported record of tions on feeding behaviour have been made for Pre- the Orthalicoidea from Africa and enables us to stonella, this is consistent with the microhabitat in confirm, contrary to published opinion regarding which both P. bowkeri and P. nuptialis occur (i.e. bare orthalicoid/bulimulid biogeography (Pilsbry, 1902; rock surfaces) and with the heavy wear evident on the Solem, 1959, 1998; Peake, 1978; Breure, 1979; Tillier, anterior radula teeth. Radulae of this kind are known 1989; Schileyko, 1999a), that the group is represented from several unrelated families, including the Bulim- in the African fauna. We are now able to show that the ulidae, Camaenidae, Chondrinidae, and Clausiliidae. Bulimulidae s.l. has a classic tri-continental distribu- However, despite being an adaptive and thus poten- tion, with representatives in central and southern tially convergent character, closer examination of America, Australasia, and southern Africa (Fig. 10). the illustrations provided by Breure & Gittenberger Although the acavoid families (including inter alia the (1982) shows the overall form of the radula teeth in Acavidae, Caryodidae, Dorcasiidae, Megalobulimidae Prestonella to be more similar to that of the bulimulid and Strophocheilidae) are collectively represented in taxa than to that of the other families mentioned. these three southern continents, the individual fami- This is confirmed by comparison with illustrations of lies are of more restricted distribution. The Buli- further orthalicoid radulae given by Breure (1978); mulidae may perhaps be the only stylommatophoran and (Breure & Eskens 1981). The general form and family with strong claims to a Gondwanan origin, proportions of the tooth cusps and their base plates in which is known to occur in all three of these southern Prestonella closely resembles that of the Bulimulidae. land masses (Peake, 1978: 453). It remains unre- In the latero-marginal series, there is a progres- corded in Madagascar and India/Sri Lanka. Other sion from inner monocuspid to outer tricuspid teeth, families with wide distributions in both the Old with the latter resembling the laterals of non-rock- and New World (e.g. Charopidae and Streptaxidae) scraping bulimulids such as species of Discoleus and (Bruggen, 1987) show a less relictual pattern of occur- Drymaeus (Breure, 1978: pl. 22, fig. 8; Breure & rence and are perhaps more likely to have achieved Eskens, 1981: pl. 3, fig. 1), notably with regard to the such widespread distributions, at least in part, basal ectocone and more distal endocone. through passive dispersal, facilitated by the small size Solem (1978) considered variation in stylommato- of many species. Hausdorf (2000) demonstrated a phoran jaw morphology too great to be phylogeneti- significant negative correlation between body size and cally informative. Jaw-related characters have been extent of distribution in limacoid snails, and diminu- included in phylogenetic analyses (Hausdorf, 1998), tive size is evidently also important for other snail but they appear to exhibit frequent exceptions and taxa in which aerial dispersal is evident (Vagvolgyi, homoplasies. Nonetheless, the stegognathous jaw 1975; Kirchner, Krätzner & Welter-Schultes, 1997).

© 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 96, 203–221 216 D. G. HERBERT and A. MITCHELL Downloaded from https://academic.oup.com/biolinnean/article/96/1/203/2448044 by guest on 27 September 2021

Figure 10. Global distribution (cross-hatched areas) of Orthalicoidea: Bulimulidae s.l.

RELATIONSHIPS WITHIN THE ORTHALICOIDEA axially ribbed sculpture of the protoconch, and the With strong evidence suggesting that Prestonella is lack of apertural dentition. Species with a shell referable to the Orthalicoidea: Bulimulidae s.l.,a resembling that of Prestonella occur in a number of further question that needs to be addressed concerns bulimuline genera, namely: Bostryx, Bothriembryon, its relationships within this diverse and widely dis- Drymaeus, Scutalus, Simpulopsis,andStenostylus. tributed group. This question, however, is confounded The derived characters present in the other suprage- by lack of consensus regarding the composition and neric taxa, but absent in Prestonella and the Bulim- ranking of the various suprageneric taxa within ulinae, are summarized in Table 3. Although the the Orthalicoidea (Herbert, 2007: 4). In reviewing similarity between Prestonella and members of the his extensive morphological studies, Breure (1979) Bulimulinae concerns character states that Breure employed the term Bulimulidae to encompass what is (1979) considered plesiomorphic (and which may thus now generally termed the Orthalicoidea. This he be of limited phylogenetic significance), they suggest divided into five subfamilies, distributed geographi- that Prestonella represents a relatively little modified cally as shown in Table 3. He also summarized the stage in bulimulid evolution. anatomical diversity evident within the group as a Within the bulimulid clade, it is of interest that the whole and provided a tentative phylogeny. When species so far included in the molecular analysis assessed in the light of Breure’s data (Table 3), the cluster in two well-supported sub-clades showing morphological character states exhibited by Pre- clear continental structure. Bothriembryon has not stonella, although frequently plesiomorphic, are gen- previously been included in molecular analyses of this erally most similar to those found in the Bulimulinae. nature, but was included here to provide greater Such plesiomorphic character states shared by Pre- representivity from the Australasian region and stonella and the Bulimulinae include the long cylin- because, like Prestonella, it has a relatively conser- drical penis, the lack of clear differentiation between vative genital morphology (Breure, 1978; Kershaw, the penis, epiphallus and flagellum, the short penial 1986), which led Breure to regard it as the only Old retractor muscle, the long bursa copulatrix duct, the World component of the Bulimulinae. With the excep- lack of a distinct genital atrium, the absence of a tion of Gaeotis and Placostylus, all the orthalicoid penial appendage and cuff-like basal penial sheath, genera for which nuclear DNA data are available the simple, longitudinally ridged lumen of the penis exhibit the relatively conservative anatomy of bulim- and epiphallus (no lateral pouches or tubes), the uline taxa. Species with such anatomy thus occur smooth (nongranular) jaw with numerous plates, the on all three southern Gondwanan land masses. That

Table 3. Geographical distribution and signiﬁcant morphological character states of suprageneric taxa within the Bulimulidae sensu Breure (1979)

Taxon Distribution Morphological character states

Bulimulinae Southern USA, Neotropics, Typically snail-like; pulmonary anatomy sigmurethrous; South America and transverse rows of radula teeth more or less straight; jaw southern Australia smooth (nongranular) with numerous plates; penis cylindrical; ﬂagellum present; penial retractor thin and short; bursa copulatrix duct long; genital atrium small; penial appendage

absent; lumen of penis simple and longitudinally ridged (no Downloaded from https://academic.oup.com/biolinnean/article/96/1/203/2448044 by guest on 27 September 2021 lateral pouches or tubes); protoconch often axially ribbed; aperture lacking dentition; columellar muscle present. Amphibuliminae Caribbean Highly modiﬁed semi-slugs; pulmonary anatomy heterurethrous; protoconch smooth; transverse rows of radula teeth V-shaped (except Peltella); elements of penial complex relatively well demarcated; protoconch smooth; columellar muscle absent. Odontostominae Eastern South America Reduced number of jaw plates (< 20); pulmonary venation weak, shell aperture dentate. Orthalicinae Neotropics Transverse rows of radula teeth V-shaped; jaw with columnar internal structure; genital atrium enlarged (sometimes with separate male and female openings); penis conical and with an appendix or gland; protoconch smooth. Placostylinae Melanesia and northern Jaw surface granular; penis club-shaped; ﬂagellum reduced or New Zealand absent; penial retractor broad and thick; sperm-oviduct torted.

The Bulimulinae (given ﬁrst) is a large and diverse group often exhibiting more than one character state; the states listed here are thought to represent the plesiomorphic condition (Breure, 1979); for the other taxa, only the character states which differ from those of the Bulimulinae are listed (data mostly from Van Mol, 1971; Breure, 1974, 1979).

Placostylus, with its relatively derived morphology, Discoleus and Plectostylus in South America (Breure, should cluster within this bulimuline group is sig- 1979). Whether the Orthalicinae and Odontostominae niﬁcant, and is consistent neither with Breure’s represent New World derivatives of this stock or phylogeny (Breure, 1979), nor with recent ranking independent orthalicoid lineages of an earlier origin, of placostyline taxa as a family distinct from the like the Amphibulimidae, remains to be established. Orthalicidae/Bulimulidae (Neubert & Janssen, 2004; The relevant sequence data are not currently avail- Bouchet et al., 2005). It is, however, consistent able. Only when these relationships have been clari- with Solem’s observation that the morphology of ﬁed will we be able to resolve the nomenclatural placostyline genera ‘merged into the Bulimulinae uncertainties that currently bedevil this group. At gradually enough for no subfamily separation to be present, although morphological data may supply desirable’ (Solem, 1988). valuable insight regarding the subfamilial affinities Given the limited number of orthalicoid taxa of orthalicoid genera, they appear to provide little included in the molecular data set, caution must be resolution concerning the relationships between the exercised in drawing conclusions from this analysis. suprageneric taxa. However, when more species Data for members of the Odontostominae and have been included in the molecular data set, a Orthalicinae are lacking. This notwithstanding, as a re-assessment of morphological characters may be working hypothesis, we suggest that each of the major instructive. Gondwanan land masses (excluding Antarctica) currently contains a bulimuline lineage exhibiting little morphological novelty, originating from a shared, pre- GONDWANAN CONSIDERATIONS fragmentation stock, from which further, more derived The continental structuring of the bulimulid clade lineages have subsequently evolved in some areas is consistent with the southern relictual origin pro- [none (remaining) in Africa, Placostylus and related posed for the Orthalicoidea. However, the nature of genera in Australasia]. These little-derived lineages the relationship, namely (South America + (southern are today represented by Bothriembryon in Australa- Africa + Australasia)), is not a pattern congruent with sia, Prestonella in southern Africa, and perhaps the accepted geological scenario, in which Africa’s

© 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 96, 203–221 218 D. G. HERBERT and A. MITCHELL most recent contact with other Gondwanan fragments Although monophyletic and well-supported, the was with South America (Sanmartín & Ronquist, phylogenetic affinities of the orthalicoid clade within 2004). Nor is it one of the other commonly occurring the Stylommatophora remain unresolved. Almost patterns listed by Sanmartín & Ronquist (2004). If certainly, the Orthalicoidea is derived relative to the sister group relationship between Prestonella and the ‘achatinoid’ group, and probably basal to the the Australasian taxa that we have identiﬁed persists Orthurethra + Arionoidea + Limacoidea group (Wade when more taxa are added to the analysis, then a et al., 2006) but, with poor resolution at deeper more complex explanation must be sought. levels within the stylommatophoran clade, we can On the basis of Early Tertiary fossils, Solem (1979) establish no more than a weakly supported associa- suggested that the Bulimulidae [s.l.] arose in North tion with the Madagascan Leucotaenius (Acavidae) America and subsequently spread into South and the elasmognaths (Succinea and Athoracopho- Downloaded from https://academic.oup.com/biolinnean/article/96/1/203/2448044 by guest on 27 September 2021 America and Australasia. However, the fossils con- rus). Node support for this clade was not strong cerned are dubiously bulimulid and fossil bulimulid (PP = 0.72), but increased relative to that in the taxa of a similar age (Eocene) are also known Wade et al. (2006) Bayesian tree (PP = 0.48; C. Wade, from South America (Parodiz, 1969). In any case, if pers. comm.). However, it is not clear whether this is the group is truly Gondwanan, then it must have due to the inclusion of additional taxa, to the reduc- arisen well before the Tertiary, by which time the tion in the size of our data set, or to small differ- fragmentation of the super-continent was already ences in the analysis parameters. As noted by Wade well advanced. Breure (1979) dated the origin of the et al. (2006), the sister group relationship between Bulimulidae at 80–95 Mya, but, in the process, Leucotaenius and the elasmognaths is enigmatic, assumed bulimulid taxa to be absent from Africa. in view of the acavoid morphology of Leucotaenius Our new observations demonstrate that, under (Mead, 1985). However, Mead’s assessment of the a vicariance scenario, the ancestral orthalicoid/ morphology of Leucotaenius focused on demonstrat- bulimulid stock must predate this, given that Africa’s ing that it was not of achatinid affinity, and his last contact with any of the other Gondwana frag- referral of the genus to the Acavidae was somewhat ments was with South America in the region of the tentative. In view of the molecular evidence, a reap- Gulf of Guinea, from which it separated 95–110 Mya praisal of Leucotaenius morphology is warranted and (Pitman et al., 1993; Sanmartín & Ronquist, 2004), the possibility that it represents a long isolated during the Albian to Early Cenomanian. However, taxon of orthalicoid origin needs to be considered. the separation of South America and southern Africa The extreme length of the branch subtending the was initiated well before this, perhaps coincident elasmognath clade suggests that great caution must with the origin of the Paraná-Entendeka plume at be taken in drawing conclusions regarding its phy- approximately 132 Mya (Valanginian), resulting in logenetic placement (Wade et al., 2006). The position the opening of the proto-South Atlantic (Pitman of the Cerionidae within the Stylommatophora and et al., 1993; Lawver, Gahagan & Dalziel, 1998; Jokat its possible relationship with the Orthalicoidea (Nor- et al., 2003). If the sister group relationship observed dsieck, 1986; Bouchet et al., 2005) is not resolved. between Prestonella and the Australasian clade is The clustering of a group of Gondwanan families genuine, then an even earlier origin must be con- (Acavidae, Rhytididae, Chlamydephoridae, Dorcasi- templated; one involving East Antarctica, which idae, Caryodidae, Megalobulimidae, Corillidae) iden- separated from southern Africa 162–165 Mya tiﬁed by Wade et al. (2001) was not apparent in our (McLoughlin, 2001). Such a scenario would require Bayesian tree, nor that of Wade et al. (2006). In our that the origin of the Stylommatophora predate the tree, these families diverged independently from Upper Jurassic (150 Mya) estimate proposed by an unresolved polytomy at the base of the non- Tillier, Masselot & Tillier (1996). achatinoid clade, with the exception of the Chlamy- Clearly, Prestonella and its ancestors have been dephoridae and Rhytididae, which formed a well- isolated in Africa (possibly restricted to southern supported clade (PP = 0.99). Nonetheless, an affinity Africa) for an extensive period of time (at the least between Gondwanan taxa within a broadly inter- since the Early to Mid Cretaceous). As suggested by preted acavoid group is evident from morphology Wade et al. (2001), more recent trans-oceanic, sweep- (Zilch, 1959–60; Tillier, 1989; Emberton, 1990; Schi- stakes dispersal between the separate Gondwana leyko, 1999b), and both Watson (Connolly, 1915: 133) fragments, although perhaps more common than was and Bequaert (1948: 4) have suggested that the previously accepted (Sanmartín & Ronquist, 2004; Orthalicoidea [Bulimuloidea/Bulimulidae] is perhaps McGlone, 2005; de Queiroz, 2005; Waters & Craw, the group most closely related to the Acavoidea/ 2006), appears to be improbable for snails of rela- Acavidae. Although this is an appealing hypothesis, tively large size (compared to many charopids and it does not appear that the nuclear rRNA gene streptaxids; see above). cluster is a suitable marker for clarifying such

© 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 96, 203–221 PRESTONELLA: AN AFRICAN ORTHALICOID SNAIL 219 relatively ancient relationships within the non- Breure ASH, Eskens AAC. 1981. Notes on and descriptions achatinoid Stylommatophora. of Bulimulidae (Mollusca, Gastropoda), II. Zoologische Ver- handelingen 186: 3–111, 8 pls. Breure ASH, Gittenberger E. 1982. The rock scraping ACKNOWLEDGEMENTS radula, a striking case of convergence. Netherlands Journal of Zoology 32: 307–312. For assistance in the field, we thank the Brown Breure ASH, Schouten JR. 1985. Notes on and descriptions family at Glen Avon farm (Somerset East), the of Bulimulidae (Mollusca, Gastropoda), III. Zoologische Ver- Michaud family at Plankfontein farm (Cradock), and handelingen 216: 3–98. particularly Mary Cole (East London Museum), who van Bruggen AC. 1977. A preliminary analysis of the non- planned our field work in the Eastern Cape and marine Gastropoda Euthyneura families. Proceedings of the Downloaded from https://academic.oup.com/biolinnean/article/96/1/203/2448044 by guest on 27 September 2021 collected additional material in the Beaufort West Fifth European Malacological Congress. Malacologia 16: area. Linda Davis skilfully prepared the anatomical 75–80. illustrations from camera lucida drawings and Adnan van Bruggen AC. 1978. Land molluscs. In: Werger MJA, ed. Moussalli provided valuable comment on the manu- Biogeography and ecology of southern Africa. The Hague: script. We also acknowledge Jon Ablett and Kathie W Junk, 877–923. Way for facilitating access to BMNH type material, van Bruggen AC. 1985. The terrestrial molluscs of Lesotho Shirley Slack-Smith and Corey Whisson (WAM) for (southern Africa), a first contribution, with detailed notes tissue of Bothriembryon indutus, and the University on Archachatina machachensis (Mollusca, Gastropoda). of KwaZulu-Natal for use of the facilities at its Proceedings of the Koninklijke Nederlandse Akademie van Molecular Biology Unit and Centre for Electron Wetenschappen, Series C, 88: 267–296. Microscopy. Chris Wade very kindly sent us the DNA van Bruggen AC. 1986. Aspects of the diversity of the land sequence alignment from Wade et al. (2006). This molluscs of the Afrotropical region. Revue de Zoologie Afric- research was funded largely through a grant from the aine 100: 29–45. National Research Foundation, South Africa, and a van Bruggen AC. 1987. Aspects of the biogeography of some terrestrial animals. In: Hovenkamp P, Gittenberger E, Hen- NSW BioFirst award to A.M. nipman D, de Jong R, Roos MC, Sluys R, Zandee M. eds. 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