Because of the paucity of anatomical information on diverse species of Xenopus, we provide summaries for the genus, each subgenus, and two species groups. In addition, we provide accounts for specific species, including six new species that we describe below.

Species within the subgenus Silurana are readily distinguished from one another using nucleotide sequences from mitochondrial DNA or autosomal DNA, and by using a combination of karyotype and vocalization data. Below, we describe one new species (Silurana new tetraploid 1 sensu Evans et al. [ 23 ]) and for Silurana new tetraploid 2 sensu Evans et al. [ 23 ] we resurrect the name Xenopus calcaratus Peters [ 54 ] from synonymy with X. tropicalis.

These subgenera are further distinguished by the parasites they host. For example, the monogenean Protopolystoma, the digenean Dolfuschella, and the tapeworm Cephalochlamys are represented by multiple species in subgenus Xenopus, but do not infect species of the subgenus Silurana, and the monogenean Gyrdicotylus and the digeneans Oligolecithus and Progonimodiscus each have different species specific to each host subgenus [ 52 ]. The camallanid nematodes occur in both host subgenera, but their phylogenetic relationships suggest independent colonization of each [ 53 ]. We recognize this clade as the subgenus Silurana within the genus Xenopus with an aim of promoting consistency with the large body of research focused on the model organism in Xenopus tropicalis.

Comparison of osteology of subgenera Xenopus (left, holotype specimen of Xenopus amieti, MHNG 2030.80 and Silurana (right, X. calcaratus, CAS 207759). Differences include (a) paired nasal bones in subgenus Silurana but not subgenus Xenopus, (b) absence of the vomer bones in the palate of subgenus Silurana but not subgenus Xenopus and (c) fusion of the first two presacral vertebrae in subgenus Silurana but not subgenus Xenopus.

Comparison of external morphology of subgenera Xenopus (right, specimen of X. victorianus CAS 250836 [DCB-202]) and Silurana (left, X. calcaratus CAS 207759) including in Silurana rougher skin, relatively smaller eyes, relatively shorter subocular tentacle (in comparison to sympatric Xenopus species), and relatively less of eye covered by lower eyelid (top), relatively shorter feet (middle), and ventrally fused cloacal lobes, claw on prehallux, and lack of skin ridge on first pedal digit from the prehallus (bottom).

A summary phylogeny inferred by comparing the mitochondrial and autosomal gene trees depicted in Figs 1 and 2 . New and resurrected species described here are in red. S, A, L, and M refer to subgenus Silurana, and the amieti, laevis, and muelleri species groups within subgenus Xenopus respectively. Dotted lines indicate paternal ancestral lineages. Circles over internal nodes indicate allopolyploidization events; shapes on branch tips indicate ploidy of extant species; colored next to these shapes circles indicate call type inferred from this study and Tobias et al. [ 42 ]. Letters over red dots refer to ancestors whose homeologous lineages are labled in Fig 2 . Daggers indicate lost ancestors, including up to three diploid species (assuming allotetraploidization in subgenus Xenopus) and at least three tetraploid ancestors (A, B, and C).

Labeling follows Fig 1 with the addition that tetraploid homeologs are indicated with α and β and, for octoploid and dodecaploids, each of these homeologs classes is further divided into two or three categories indicated by numbers. Homeologs of new and resurrected species are in red. Letters in circles indicate homeologous lineages inferred to be descended from six tetraploid ancestral species A–F. Data are lacking from X. petersii, X. poweri, and X. victorianus, which form a clade with X. laevis, and from X. fraseri.

New and resurrected species detailed here are indicated in red, possible additional new species are indicated with blue, and species with paraphyletic mtDNA are indicated with gray. Dots subtending nodes indicate posterior probabilities as indicated by the key with some values over terminal clades omitted for clarity. The scale is in units of millions of years.

Genetic data from the mitochondrial and nuclear genomes strongly support a clade of species that is sister to all other living species of Xenopus (Figs 1 – 3 ) [ 9 , 16 , 20 – 23 ]. We use Silurana as a subgenus following Kobel et al. [ 15 ]. Silurana contains four species from West and Central Africa: X. tropicalis, X. epitropicalis, and two other species described below (one new, one resurrected). In general, these medium-sized species in the subgenus Silurana are distinguished from species in the subgenus Xenopus by the following combination of external morphological features ( Fig 4 ): (1) cloacal lobes fused ventrally; (2) keratinous claws on prehallux as well as the first three toes; (3) many small spicules across the dorsum; (4) lack of a dermal ridge extending along the first toe from the prehallux; (5) many scattered tubercles on the plantar surface; (6) relatively short feet; (7) relatively small eyes; (8) relatively little of the eye covered by the lower eyelid; (9) relatively shorter subocular tentacle in comparison to the sympatric species in the subgenus Xenopus; (10) generally fewer plaques in each row of the lateral-line system than in the subgenus Xenopus, though the ranges can overlap between taxa; (11) tadpoles with relatively long barbels and generally fewer small melanophores [ 50 ]. In addition, species of Silurana are diagnosable by features that require molecular or internal morphological study, reviewed in [ 12 ], including a haploid karyotype of 10 chromosomes [ 51 ], fusion of the first two presacral vertebrae [ 11 ], paired (unfused) nasal bones [ 11 ], and absence of the vomer bones in the palate [ 11 ], and thus also vomerine teeth ( Fig 5 ).

Morphological differentiation among adult Xenopus varies from substantial to subtle, with few species having unique distinguishing characteristics. With a few exceptions, body size (SVL) and lateral-lines are insufficient for differentiating most of these species (Tables 1 – 3 ). Xenopus gilli is distinguished by its unique dorsal pattern consisting of longitudinal dark brown blotches separated by pale brown coloration. Xenopus longipes is distinguished by feet that are large relative to its small body size. The four species previously recognized as Xenopus laevis, for example [ 15 ], which are now designated X. laevis, X. petersii, X. poweri, and X. victorianus [ 2 ], are distinguished both by the large body size of adults, especially the population of X. laevis from the Cape Region in South Africa [ 45 ], and by the large size of the eyes relative to body size. Xenopus muelleri and X. fraseri are the only described extant species in the genus with vomerine teeth [ 46 – 48 ], though see comments below regarding the new tetraploid species of the muelleri species group.

Species of Xenopus have compressed bodies that are oblong and ovoid in dorsal view. The head is subtriangular, and the rostrum projects just beyond the lower jaw, though species vary in the degree to which the rostrum is blunt or pointed. The canthus rostralis is typically flat to weakly concave, the loreal region is generally flat, and the internarial region varies from flat to weakly concave. All species lack a tongue and have a single opening to the Eustachian tubes. The floor of the mouth is typically wrinkled and covered in small pustules. The posteroventrally directed choanae are large, rounded, and largely or entirely visible in ventral view. Premaxillary and maxillary teeth are present, but vomerine teeth are nearly always absent. The nares are prominent ellipsoid slits directed dorsally; a small sheet of skin projects from the margins of each naris, the extent and morphology of which varies among species but often features a laterally projecting nubbin. The size of the eye relative to the head varies among species as does the extent to which the eye is covered by the lower eyelid. In preservative, the pupil is typically round. Each eye is encircled by lateral-line plaques located on a raised ring of skin, though the degree to which this ring is observable varies based on specimen preservation. A subocular tentacle extends from the lateral margin of each eye; the length of this tentacle varies among species and is absent in two species (X. gilli and X. largeni). All species lack an externally visible tympanic annulus. The skin is generally smooth, although it can be covered by small spicules (especially those in the subgenus Silurana). A prominent feature of all adult Xenopus is the lateral-line system; the individual plaques (each comprising multiple sensory organs) resemble stitches. Distinct lateral-line rows extend across the skin of the head and dorsal, lateral, and ventral body. In both males and females, the medial surfaces of the manual digits are covered by small black punctiform spicules. In males, these are accentuated into nuptial pads comprising sheets of darkly pigmented spicules on the manual digits as well as on the upper arm, forearm, and sometimes axillary region. The forelimbs are typically moderately robust and have elongate manual digits that lack webbing (in contrast to other African pipids, the dwarf clawed frogs Hymenochirus and Pseudhymenochirus). The relative length of the manual digits varies among species, though in all species these digits typically terminate in small bulb-like tips. Being primarily aquatic frogs, these species have large hind limbs with fully webbed feet (i.e., extending to either the toe tip or base of the keratinous claw). The extent to which the pedal webbing is pigmented varies among species. As in most other frogs, the digits of the foot are longer than those of the hand, with the fourth toe being the longest and the first the shortest. As their common name of African clawed frogs suggests, all species have dark brown or black keratinous claws on the first three pedal digits; similar to the hand, those pedal digits that lack keratinous claws terminate in bulb-like tips. Many species have a keratinous claw on the prehallux, which in combination with other characters, can be diagnostic of particular species groups. The hands and feet lack subarticular tubercles, though scattered pustules are found on the plantar surface of some species. Female Xenopus are often identifiable by their protruding cloacal lobes, the number and/or fusion of which varies among species. In life, the coloration of most species ranges from grays to browns, sometimes with patterning or markings that are indicative of particular species.

All species in the genus Xenopus have size dimorphism (females larger than males), fully webbed feet, a dorsoventrally compressed body, relatively smooth skin, and lateral-line organs. The tadpoles are suspension feeders that are morphologically similar across species and notable for their slit-like anteriorly directed mouth, a pair of spiracles, conspicuous barbels, and lack of keratinized mouthparts. The two subgenera (Silurana and Xenopus) are distinguished by a number of morphological, genetic, karyotype, and host-parasite characters (see below).

Habitat and range.—Based on samples with available genetic data, X. tropicalis is widespread across West Africa, extending from Sierra Leone east to at least into western Cameroon [ 1 , 59 ]. We lack genetic data for samples from central Cameroon, though previously Tinsley et al. [ 1 ] recognized the easternmost extent of this species as the Sanaga River.

Variation.—Variation in the lateral-line based on specimens from across the geographic range of this species in West Africa (n = 15, given as mean and range): orbital– 10 (8–12); oral– 11 (9–12); medial– 18 (16–21); lateral– 20 (17–22); ventral– 18 (16–21).

Comments on syntypes.—The four syntypes are two late-stage tadpoles and two metamorphs in stages corresponding to NF 57–60 based on the staging developed by Nieuwkoop and Faber [ 73 ] for X. laevis ( S9 – S14 Figs). Two specimens, BMNH 1947.2.24.84 and 86 are incomplete due to damage of the posteriormost tail (respectively, NF 59 at 46 mm total length, and NF 58 at 37 mm total length). The latest stage individual (BMNH 1947.2.24.83) is at NF 59/60 with a total length of 54 mm, whereas the earliest stage specimen (BMNH 1947.2.24.85) has a total length of 63 mm. As indicated in the original description, these larval specimens have elongate barbels as is typical of species in the subgenus Silurana. The coloration of the latest stage individual (BMNH 1947.2.24.83) is medium brown with scattered darker brown variegations ( S9 – S14 Figs), with a venter that is pale creamy beige and a small stark white region at the opercular region. The specimen lacks pigmentation on the pedal webbing and has three dark keratinous pedal claws as well as a similar claw on the prehallux. The other specimens are largely consistent in appearance, and all appear to have faded in coloration over time. We designate the latest stage individual BMNH 1947.2.24.83 as the lectotype.

Diagnosis.—Xenopus tropicalis exhibits all of the morphological features of subgenus Silurana described above, and is diagnosable from other species in the subgenus by unique nucleotide substitutions in mitochondrial and autosomal DNA (mitochondrial DNA is paraphyletic; Figs 1 and 2 and S1 and S2 ), by having a trill-type call, and by being the only diploid in the genus. In addition, it differs from other species of Silurana by having higher intensity modulation (~38) of its call in contrast to other species (which is ~10), Table 4 , [ 42 ]. As noted above, characters previously proposed as diagnostic between X. epitropicalis and X. tropicalis, including adult body size, number of lateral-line plaques around the eye, and coloration (Fischberg et al., 1982; Kobel et al., 1996), are not useful for distinguishing these two species.

Syntypes.—BMNH 1947.2.24.83–86, metamorphs and tadpoles, “West Africa, Lagos,” now Federal Republic of Nigeria, coll. R.B.N. Walter. To stabilize the identity of the nomen, we designate BMNH 1947.2.24.83 (a late stage metamorph) as the lectotype.

Remarks.—In addition to wild-caught individuals with associated genetic data, we also have DNA sequence from CAS 250558, a specimen from the laboratory colony established in Geneva. Similarity to recently collected specimens confirms that previous authors have in fact published on the same entity that we describe here as X. mellotropicalis.

Etymology.—For the species epithet, we have combined the Greek word μέλλω (mello), often interpreted as indicating that something is “about to” happen [ 71 ], to tropicalis, which forms part of the specific epithet of the other two other species in the subgenus Silurana (X. tropicalis and X. epitropicalis). This species epithet, which is an adjective, suggests the long delay in a formal description of this species that has been referenced in the literature for nearly thirty years beginning with Graf & Fischberg [ 62 ]. In 1993, a series of publications referred to this species as “Xenopus paratropicalis,” or “Silurana paratropicalis” but this is not a valid name, for review see [ 72 ]

Habitat and range.—Based on surveys of specimens with genetic data, X. mellotropicalis is found in both disturbed and forested areas in Central Africa, including the Congo Republic [ 61 ], Cameroon, Gabon, and Democratic Republic of Congo ( Fig 10 ). It is likely also found in mainland Equatorial Guinea and southwestern Central African Republic. It lives in sympatry with several Xenopus species, including X. epitropicalis in the Republic of Congo [ 61 ], as well as X. cf. fraseri 1, and X. cf. fraseri 2 sensu [ 23 ] in DRC and Cameroon, respectively; see below for descriptions of the latter two species.

Coloration in life.—Based on color photographs of NCSM 78871 ( Fig 8 ), the dorsum of X. mellotropicalis is pale greenish and grayish brown in life with scattered medium variegations. The venter is a pale gray with hints of pale yellow blotches, especially posteriorly and on the ventral hind limbs.

Coloration of holotype (in alcohol).—Dark brownish gray on dorsum and limbs (Figs 7 and S6 – S8 ). The darker coloration of the dorsum is somewhat paler on the anterior; there is a thin pale gray interocular line; the coloration anterior and posterior to the interocular line is similar. There are no prominent spots on the dorsum, but the pattern tends to be finely variegated. The venter is orange-gray and darker gray towards the head; variegations on the ventral thighs are darker than those on the venter.

Measurements.—Female specimens reach a maximum SVL of 58 mm (mean: 53 mm; n = 14), and males reach a maximum SVL of 51 mm (mean: 45 mm; n = 12) ( Table 2 ). See S1 Table for more measurements.

Relative lengths of fingers: II ≈ III > IV > I; relative lengths of fingers when adpressed: II > III > IV > I; thigh length 111% of crus length; relative length of toes: IV > V ≈ III > II > I; foot and toes along metatarsals and digits with scattered prominent pustules on ventral surfaces.

Skin smooth; small prominent asperities on snout and scattered over much of body and limbs; small tubercles across plantar surface; punctiform but well separated lateral-line plaques around eye; lateral-line plaques most prominent on dorsal and lateral surfaces and extending onto the ventral surface, and oral and ventral plaques difficult to observe due to similarity of coloration with venter; counts of lateral-line rows as follows: orbital– 11, oral– 12, medial– 17, lateral– 17, ventral– 13; male nuptial pads are well developed appearing as dark keratinous patches on ventral surface of the arm and forearm, and extending along metacarpals and digits.

Description of the holotype.—Medium-sized (SVL 48 mm), moderately robust male ( Table 1 , Figs 6 and S3 – S5 ); rostral tip blunt and rounded in dorsal view; eyes not projecting beyond margins of orbit in dorsal view and projecting slightly beyond dorsal margin of head in lateral view; subocular tentacle short, length less than half eye diameter; eye diameter 42% of interorbital distance, 83% of eye–narial distance, and 1.4 times distance from naris to rostral tip; internarial distance 46% of interorbital distance; no vomerine teeth.

Diagnosis.—Xenopus mellotropicalis is a tetraploid species with a burst-type call that exhibits all of the morphological features of subgenus Silurana described above ( Fig 11 ). It differs from other species of Silurana in the following ways: from all species by unique nucleotide substitutions in mitochondrial and autosomal DNA (Figs 1 and 2 and S1 and S2 ); from X. calcaratus by more pulses in the call, more defined lateral-line plaques, and generally lacking prominent dark spots on the dorsum (common in the Bioko population of X. calcaratus); from X. epitropicalis by lacking a biphasic call, and having shorter interpulse intervals; from X. tropicalis by being tetraploid, by lacking a trill-type call and having less intensity modulation in the call, and generally lacking prominent dark spots on the dorsum. Xenopus mellotropicalis also differs from X. tropicalis and X. epitropicalis by the peptides present in its norepinephrine-stimulated skin secretions [ 70 ].

Paratypes.—Gabonese Republic: NCSM 78871, adult female, Ogooué-Ivindo Province, Rougier Gabon Forestry Concession, N 0.2018 E 12.2693, 221 m, 16 October 2011, coll. B. L. Stuart, R. C. Bell, T. Ogombet, U. Eyagui, P. Endazokou; NCSM 78881, juvenile, same collectors, N 0.0426 E 12.2983, 21 October 2011. Republic of Cameroon: Centre Region: NMP6V 74568, adult female, Ebogo, N 3.3913 E 11.4663, 628 m, 17 October 2009, coll. V. Gvoždík; Est Region: NMP6V 74718, subadult female, Kika, N 1.9419° E 15.6269°, 337 m, 30 May 2010, coll. V. Gvoždík, O. Kopecký; Sud Region: MHNG 2644.58 (AMNH 17288), male laboratory animal descended from animals collected in Nkoemvone, N 2.8800° E 11.1500°, ~575 m, unknown date, coll. H. R. Kobel; ZFMK 87790–1, sex unknown, Nkoelon, Campo Region, N 2.3500° E 10.6167°, 76 m, October 2007, coll. M. Barej, J. Wurstner; Democratic Republic of Congo: CAS 250558, female, Bas-Congo Province, Malemba, S 5.83°, E 12.57° (estimated), ~150 m, collection date unknown, coll. D. Rungger (specimen from captive population originally housed in Laboratoire de Génétique Animale et Végétale, Université de Genève; possibly wild caught).

Remarks.—Mitochondrial DNA sequences from our X. epitropicalis sample ( S2 Table ) are derived from the lab colony in Geneva from which the species was described.

Habitat and range.—Based on samples with genetic data, X. epitropicalis is known from near the type locality in Kinshasa, Democratic Republic of Congo, to the northeast of this locality along the Congo River near the confluence with the Kwa River, and from Pointe Noire, Republic of Congo [ 61 ], where it was recorded syntopically with the new tetraploid of Silurana described below. There are records that may be attributable to X. epitropicalis, though without genetic data, from northeastern Democratic Republic of Congo [ 1 ]. Most previous records of X. epitropicalis, including from Cameroon and Gabon [ 1 ], most likely represent the new tetraploid species of Silurana described below.

Vocalization.—Xenopus epitropicalis has a biphasic call, which is a unique call type in the subgenus, and has longer interpulse intervals (~22 msec) than other species of Silurana (~10 msec) [ 42 ].

Coloration of holotype (in alcohol).—Dorsum is grayish medium brown with fine mottling of dark brown across dorsum and hindlimbs, becoming paler on the thigh and inguinal region ( S9 – S14 Figs). There is a pale gray blotch representing the incomplete interocular line across a darker gray rostrum. The venter and limbs are a uniform dusky gray cream, becoming darker gray towards the head. The plantar surface is uniformly grayish brown. The lateral-line plaques are generally without pigmentation and appear pale in coloration.

Measurements.—Female specimens reach a maximum SVL of 68 mm (mean: 56 mm; n = 12), and males reach a maximum SVL of 51 mm (mean: 43 mm; n = 12) ( Table 2 ). The original description [ 55 ] gives the maximum SVL of 72 mm in females and 53 mm in males, both of which measurements are slightly larger than those of the specimens we examined. See S1 Table for more measurements.

Skin smooth; small prominent asperities on snout and scattered over much of body and limbs, and strongly developed on the forelimbs; many small tubercles across plantar surface; punctiform, globular, and closely spaced lateral-line plaques around eye; lateral-line plaques most prominent on dorsal and lateral surfaces and extending onto ventral surface; oral and ventral plaques projecting from skin; counts of lateral-line rows as follows: orbital– 12, oral– 11, medial– 21, lateral– 23, ventral– 19.

Description of the holotype.—Large-sized (SVL 68 mm), robust female ( S9 – S14 Figs; Table 1 ); rostral tip blunt and somewhat rounded in dorsal view; eyes not projecting beyond margins of orbit in dorsal view and projecting slightly beyond dorsal margin of head in lateral view; subocular tentacle short, length less than half eye diameter; eye diameter 37% of interorbital distance, 93% of eye–narial distance, and 1.5 times distance from naris to rostral tip; internarial distance 47% of interorbital distance; no vomerine teeth.

Diagnosis.—Xenopus epitropicalis is a tetraploid species with a biphasic-type call [ 42 ] that exhibits all of the morphological features of subgenus Silurana described above. It differs from other species of Silurana in the following ways: from all species by unique nucleotide substitutions in mitochondrial and autosomal DNA (Figs 1 and 2 and S1 and S2 ); from both X. calcaratus and the new tetraploid described below by having a biphasic call and longer interpulse intervals; and from X. tropicalis by being tetraploid, lacking a trill-type call, and having less intensity modulation in the call. Characters previously proposed as diagnostic between X. epitropicalis and X. tropicalis, including adult body size, number of lateral-line plaques around the eye, and coloration [ 15 , 55 ] are not useful for distinguishing these two species because of overlapping patterns of variation ( Table 3 ).

Holotype.—BMNH 1982.462, female, Democratic Republic of Congo, “au confluent de la Funa et de la Kemi, à 8 km au sud du centre de Kinshasa (Zaïre),” [confluence of the Funa and Kemi rivers, 8 km south of the centre of Kinshasa (Zaire)]; 350 m; S 4.3° E 15.3°, August 1978, coll. V. Nzingula.

Dorsal view is on the left and ventral view is on the right, including the lectotype specimen of Xenopus (Silurana) calcaratus (ZMB 8255A) from Cameroon and a specimen from Bioko Island (CAS 207759), holotype of X. (S.) mellotropicalis (NCSM 76797), and the holotypes of the new dodecaploid species from subgenus Xenopus: X. eysoole (MCZ A-138016) and X. kobeli (MCZ A-148037). The type specimen of X. calcaratus was preserved with its mouth ajar.

Fig 11. MicroCT scans of skulls of two tetraploids in subgenus Silurana and two dodecaploid in subgenus Xenopus.

Remarks.—Xenopus calcaratus was described by Peters [ 54 ] based on material collected at what is now Limbe (formerly Victoria) on the coast of the Republic of Cameroon. Müller [ 58 ] suggested that X. calcaratus should be considered a junior synonym of X. tropicalis based largely on the assumption that the type specimens of X. tropicalis represent larvae and metamorphs of the same species as the types of X. calcaratus. The type locality of X. tropicalis is Lagos in present-day Nigeria and all evidence suggests that populations of Silurana from western Nigeria and farther west in Africa are referable to X. tropicalis [ 59 ]. This tetraploid species found on Bioko Island and coastal Cameroon, including Limbe ( Fig 10 ), is morphologically consistent with the types of X. calcaratus ( Fig 11 ). Thus, we resurrect Xenopus calcaratus [ 54 ] from synonymy with X. tropicalis [ 49 ] for this distinct evolutionary lineage.

Type localities with black dots inside symbols. For X. fischbergi white circles with slashes indicate specimens from which we have genetic data (including the holotype), unfilled circles are specimens from Tinsley et al. [ 1 ] and those other collections from which we lack genetic data, including one field sample from Uganda–indicated by a hexagon–in which the X. fischbergi-specific parasite Protopolystoma occidentalis was detected (J. A. Jackson & RCT, unpublished).

Habitat and range.—Based on surveys of specimens with genetic data, this species is known only from low elevations on Bioko Island (Equatorial Guinea) and coastal Cameroon near Mt. Cameroon ( Fig 10 ). In portions of both localities, X. calcaratus can be found syntopically with X. cf. fraseri 1, sensu [ 23 ], which is described as a new species below.

(A) X. calcaratus, NMP6V 74746 (VG05-S; female) from Cameroon, (B) X. mellotropicalis, CAS 255058 (BJE 3652) from Republic of Congo, (C) X. fischbergi non-vouchered sample (BJE 3873), (D) X. parafraseri, CAS 249961 (BJE 3060) from Cameroon, (E) X. allofraseri, MCZ A-148162 (BJE 3486) from Bioko Island, Equatorial Guinea, (F) X. kobeli, MCZ A-148038 (BJE 3076) from Cameroon, (G) X. eysoole, MCZ A-148097 (BJE 3220), from Cameroon.

Axes are labeled only in panel A. For some of the recorded individuals, specimen IDs are available including X. kobeli (field ID: BJE 3073), X. allofraseri (MCZ A-148176), X. parafraseri (CAS 249961), X. eysoole (MCZ A-148129 or MCZ A-148130), and X. calcaratus (field ID: VG09-368 or VG09-369).

Vocalization.—Xenopus calcaratus has a burst-type call ( Table 4 , Fig 8 ). Similar to other species of Silurana [ 42 ], the call of X. calcaratus has only one dominant frequency.

Variation.—Variation in the lateral-line based on the referred specimens above from Bioko Island (n = 17, given as mean and range): orbital– 10 (9–13); oral– 10 (9–12); medial– 16 (15–19); lateral– 18 (15–21); ventral– 17 (14–21) ( Table 3 ).

Pictures of resurrected and new species in life. This is a truncated version of Fig 7 and is meant for preview purposes. Please refer to S6 – S8 Figs in the Supporting information for the full size version.

Coloration in life.—Based on above specimens from mainland Cameroon, X. calcaratus ranges from medium grayish brown to dark gray, sometimes with dark brown spots on the dorsum (Figs 7 and S6 – S8 ). The interocular bar tends to be pale gray, rather indistinct, and the rostrum is a somewhat darker coloration than the dorsum. The venter and ventral surface of the limbs are pale yellowish gray with indistinct marbling. The posterior parts of the venter, thighs, and throat may be darker in some individuals.

Coloration of lectotype (in alcohol).—Dorsum is medium-dark brown with few distinct spots that are most prominent on the hind limbs (Figs 6 and S3 – S5 ). The darker coloration of the dorsum becomes somewhat paler anteriorly and there is a thin, pale, and indistinct interocular line. The venter ranges from a medium to pale brown, becoming darker near both the gular and inguinal regions; both the forelimbs and hind limbs are paler in coloration than the remaining venter. The lateral-line plaques are generally without pigmentation and thus appear pale in coloration.

Measurements.—Female specimens reach a maximum SVL of 59 mm (mean: 44 mm; n = 19, Table 2 ), but the size of males remains unclear because of the difficulty in determining sex for most specimens from Bioko. Two likely males from Bioko (CAS 207618, CAS 207756) have SVL of 45.2 and 46.1 mm, respectively. Additional measurements of the lectotype show that the crus (23.1 mm) is longer than the thigh (22.1 mm), the tarsus (16.7 mm), and the 4th toe (measured from the prehallux to toe tip, 15.7 mm).

Skin generally smooth; small isolated punctiform asperities across dorsal surface of head, body, forelimbs, and hind limbs; few small tubercles on plantar surface; punctiform and closely spaced lateral-line plaques around eye; lateral-line plaques (18 on both sides) most prominent on dorsal and lateral surfaces and extending onto ventral surface; oral and ventral plaques difficult to observe.

Pictures of type specimens of resurrected and new species including X. calcaratus ZMB 8255A (lectotype), X. mellotropicalis NCSM 76797 (holotype), X. allofraseri CAS 207765 (holotype), X. eysoole MCZ A-138016 (holotype), X. kobeli MCZ A-148037 (holotype), X. parafraseri MCZ A-148034 (holotype), and X. fischbergi CAS 255060 (holotype). Scale bar is 5 mm. This is a truncated version of Fig 6 and is meant for preview purposes. Please refer to S3 – S5 Figs in the Supporting information for the full size version.

Description of lectotype (ZMB 8255A).—Proportions were estimated via photographs if no exact values are given in parentheses. Large-sized (SVL 53 mm), moderately robust female (Figs 6 and S3 – S5 ; Table 1 ; determination of sex based on body size, protruding cloacal lobes, and lacking enlarged posteromedial processes of hyoid plate); rostral tip blunt and slightly squared in dorsal view; eyes weakly projecting beyond margins of orbit in dorsal view and slightly inset from dorsal margins of head in lateral view; subocular tentacle short, length slightly greater than half of eye diameter; eye diameter ~33% of interorbital distance, ~80% of eye–narial distance; internarial distance ~50% of interorbital distance; no vomerine teeth.

Diagnosis.—Xenopus calcaratus is a tetraploid species with a burst-type call that exhibits all of the morphological features of subgenus Silurana described above. Most individuals are a medium to dark brown with a pale interocular bar (more distinct in the Bioko population), and a rostrum tending to darker brown coloration than the rest of the dorsum. Specimens from Bioko Island often exhibit small scattered and irregularly shaped dark brown spots on the dorsum and hindlimbs, which has not been recorded in Cameroonian specimens. Xenopus calcaratus differs from other species of Silurana in the following ways: from all species by unique nucleotide substitutions in mitochondrial and autosomal DNA (Figs 1 and 2 and S1 and S2 ); from X. epitropicalis by having a burst-type instead of a biphasic call [ 42 ], having shorter interpulse intervals, and having somewhat less pedal webbing pigmentation; from X. tropicalis by being tetraploid, having a burst-type instead of a trill-type call and having less intensity modulation in the call; and from the tetraploid new species of Silurana described below by fewer pulses in the burst-type call, by less defined lateral-line plaques, and by having large prominent dark brown spots on the dorsum in some specimens.

Referred Specimens.—Republic of Equatorial Guinea, Bioko Island: Bioko Norte Province: CAS 207615–19 (N 3.7110° E 8.6666°; ~55 m), 18 October 1998; Bioko Sur Province: Arena Blanca Road: CAS 207752–53 and 207755–58 (N 3.5201°, E 8.5859°; ~65 m), 207759–64 (N 3.5276°, E 8.5794°; ~30 m), 14 October 1998; coll. L. G. Henwood, J. V. Vindum. Republic of Cameroon: South West Region, Bakingili, lava flow: NMP6V 74630/1-4 (two subadults, probably females, and two adult males), 25 November 2009, NMP6V 74746 (adult female), 11 December 2005, (N 4.0689°, E 9.0684°, ~180 m), coll. V. Gvoždík.

Syntypes.—ZMB 8255 (originally three specimens, only two adult females are known to be present in ZMB–ZMB 8255A and ZMB 74681 (formerly 8255B), ZMB 8326, originally two specimens, one adult female (ZMB 8326A) and one juvenile (ZMB 74682; formerly 8326B), ZMB 8328, originally five specimens, only four specimens are known to be present in ZMB, (ZMB 8328A-D, including one tadpole in metamorphosis), ZMB 8329 (one tadpole in metamorphosis), “Cameruns (Victoria)” [presently Limbe, Southwest Province, Republic of Cameroon], coll. Reichenow; based on Bauer et al. [ 56 ], verified and detailed by VG with help of F. Tillack (ZMB). Two syntypes were listed in Zoologischen Museums Greifswald [ 57 ], but no further details were provided. To stabilize the identity of the nomen, we designate ZMB 8255A (an adult female, SVL 54 mm) as the lectotype.

Xenopus tropicalis is diploid, whereas the other three species are tetraploid. Two species, X. calcaratus and the new species described below, are distinguished from X. tropicalis and X. epitropicalis by having burst-type calls, with X. calcaratus being distinguished by having fewer sound pulses ( Table 4 ) [ 42 ]. Other spectral features are similar between these two species with burst-type calls including the dominant frequency 1 (both ~500–600 Hz), inter-pulse interval (~3–15 msec), and intensity modulation (~9–19). Xenopus tropicalis and X. epitropicalis are distinguished from the other two species by having, respectively, a trill-type and a biphasic call [ 42 ]. The vocalizations of all four species have a similar dominant frequency and all species in subgenus Silurana have only one dominant frequency, which is more broadband and lower than species in subgenus Xenopus [ 42 ]. Xenopus epitropicalis is distinguished by having longer interpulse intervals (~22 msec) than the other species (~10 msec); X. tropicalis is distinguished by higher intensity modulation (~38) compared to the other species (which is ~10, Table 4 ) [ 42 ]. Similar to subgenus Xenopus, body size and lateral-lines are insufficient for differentiating species of subgenus Silurana (Tables 1 – 3 ). Our studies of variation of body size and the number of lateral-line plaques around the eye suggest that these are not useful for diagnosing species of Silurana in contrast to previous suggestions [ 15 , 55 ]. The number of lateral-line plaques around the eye for each species exhibits overlapping variation ( Table 3 ). We provide a detailed description of X. calcaratus and the new species, and short descriptions regarding the two existing species, X. epitropicalis and X. tropicalis.

We modify those traditional groupings in light of phylogenetic discoveries as well as recently described species. The groups that we recognize within the subgenus Xenopus are (1) the amieti species group (X. amieti, X. andrei, X. boumbaensis, X. itombwensis, X. lenduensis, X. longipes, X. pygmaeus, X. ruwenzoriensis, X. vestitus, X. wittei, and three new species described below), (2) the laevis species group (X. gilli, X. laevis, X. petersii, X. poweri, and X. victorianus; see Furman et al. [ 2 ]), and (3) the muelleri species group (X. borealis, X. muelleri, a new species described below, and possibly also X. clivii). The relationships of X. fraseri and the Ethiopian endemic X. largeni remain uncertain and we therefore do not assign them to a species group. The subgenus Xenopus can be differentiated from the four species in the subgenus Silurana by a number of morphological features (see above and Figs 4 and 5 ). Interestingly, patterns of parasite specificity match the species groups within Xenopus in that species of several parasite genera exclusively infect host species in either the laevis, amieti, or the muelleri species groups [ 74 , 75 ]. For several parasites (including the monogenean Protopolystoma and the digenean Dolfuschella), there are significant morphometric and life cycle differences between samples from different parts of the geographical range of X. laevis sensu lato [ 75 ] that match phylogenetic divisions within this clade [ 2 ]. Within the muelleri species group, there are distinct species of Protopolystoma, P. occidentalis and P. orientalis, that are respectively host specific to X. muelleri and the new tetraploid species in this group described below [ 75 , 76 ].

Previous authors divided Xenopus into subgroups including the laevis subgroup (X. laevis sensu lato, X. gilli, X. largeni), the muelleri subgroup (X. muelleri, X. borealis, X. clivii and an undescribed species X. “new tetraploid” / X. “muelleri west” that is described below), the fraseri subgroup (X. fraseri, X. pygmaeus, X. amieti, X. andrei, X. boumbaensis, X. ruwenzoriensis), the vestitus-wittei subgroup (X. vestitus, X. wittei), and longipes subgroup (X. longipes) [ 15 ]. Phylogenetic analyses reported here and elsewhere, for example [ 16 , 21 ] facilitate redefinition of these groups based on common ancestry (Figs 1 – 3 and S1 and S2 ). Apart from the monotypic longipes subgroup, monophyly is not strongly supported for any of these subgroups by either mitochondrial DNA or autosomal DNA. Monophyly of the muelleri subgroup is supported by autosomal DNA, but not mitochondrial DNA. Paraphyly of the fraseri and vestitus-wittei subgroups sensu Kobel et al. [ 15 ] is attributable to the reticulating evolutionary history of the allopolyploid species they contain. Uncertainty in the phylogenetic placement of X. largeni (Figs 2 and 3 ) means that monophyly is uncertain for the laevis subgroup sensu Kobel et al. [ 15 ].

Species in subgenus Xenopus

Amieti species group. The amieti species group comprises 14 species found across Central Africa, from Nigeria in the west to Uganda and Rwanda in the east, including three new species described below. Some species of this group are distinguished by being octoploid and dodecaploid (no other species group has species with these ploidy levels). Previously this group was referred to as the fraseri species group [15], but the phylogenetic affinities of X. fraseri remain uncertain (see below). We therefore do not recognize the fraseri species group and instead propose the amieti species group to include all of the species currently in the fraseri species group except X. fraseri, which we do not place in a species group for the time being. We additionally place the longipes species group (sensu Kobel et al., 1996), which includes only X. longipes, into the amieti species group based on inferred evolutionary relationships [16, 20–23]. Because phylogenetic relationships among the tetraploid ancestors of some of the octoploid and dodecaploid species in the amieti species group also include ancestors of species (X. vestitus, X. wittei, X. itombwensis, and X. lenduensis) [16, 20–22] that were previously placed in the vestitus-wittei group [15], we place all of these species in an expanded amieti species group to reflect this shared evolutionary history. In general, these medium to small-sized Xenopus species can be identified by the following combination of external morphological features: (1) unfused cloacal lobes; (2) prominent keratinous claw on the prehallux; (3) a skin ridge extending along the first pedal digit from the prehallux; (4) dorsal skin often with small spicules. The four species comprising what was previously recognized as the vestitus-wittei group [15] are distinguished by lacking the claw on the prehallux that is found in all other species in the amieti group. While unreceptive females in other species of subgenus Xenopus produce a release call when clasped by a male, females in the amieti species group do not [43].

Xenopus (Xenopus) eysoole, new species. urn:lsid:zoobank.org:act:D15EA055-1351-4AAC-92B3-BE546FF05C64 Bamiléké Clawed Frog Holotype.—MCZ A-138016 (DCB 34586), adult female, Republic of Cameroon, North-West Region, Elak Oku village, southern face of Mt. Oku, N 6.2427º, E 10.5011º, 1870 m, 16 August 2006, D. C. Blackburn, K. S. Blackburn, P. Huang, and M. T. Kouete (Figs 6 and S3–S5). Paratypes.—MCZ A-138017, adult female, same collection data as holotype. Republic of Cameroon: North-West Region: MCZ A-148094–5, female and male, 5 km east of Nkambe, N 6.5486° E 10.7600°, 1684 m, 29 October 2011, coll. B. J. Evans, D. M. Portik, S. B. Menzepoh; MCZ A-148127-8, MCZ A-148131, females, Bamenda fish pond farm, N 6.0143° E 10.2703°, 1441 m, 2 November 2011; MCZ A-148129-30, males, same locality data, coll. B. J. Evans, D. M. Portik, S. B. Menzepoh; NMP6V 74555, adult female, Babungo, N 6.0488° E 10.4259°, 1771 m, 22 December 2009, coll. R. Tropek, A. Kodádková; NMP6V 74745/1-2, females, Njikwa, Acha Tugi Mountains, N 6.0618° E 9.9158°, 1800 m, 23 January 2012, coll. R. Tropek, D. Leština. Diagnosis.—Xenopus eysoole is a dodecaploid species, with either a trill-type or burst-type call, that exhibits all of the morphological features of the amieti species group described above. Phylogenetic analysis of mitochondrial and autosomal data (Figs 1 and 2 and S1 and S2) suggests this species may be the sister-species of Xenopus longipes (based on close phylogenetic relationships of mitochondrial DNA and homeologs α1, β1, α3, and β3), or that these two species have a more complicated but intermingled evolutionary history (based on a close phylogenetic relationship between homeolog α2 of X. eysoole and X. amieti). See Discussion for possible explanations for this and other phylogenetic inferences. Xenopus eysoole differs substantially from X. longipes by larger body size (e.g., maximum female SVL we measured is 34 mm in X. longipes and 52 mm in X. eysoole), having fused nasals–unfused in X. longipes [15], transverse processes of the fourth presacral vertebra that are curved posteriorly–uncurved in X. longipes [15], relatively smaller otic capsules–large in X. longipes [15], and a creamy white ventral coloration in contrast to the yellow or orange in X. longipes. Xenopus longipes further differs from X. eysoole in that many individuals have clumps of melanophores on the ventral surface, sometimes forming an anastomosing network, and less well-defined lateral-line stitches. In addition, Xenopus eysoole differs from the other species within the amieti species group by having a larger body size than X. pygmaeus and longer interpulse intervals than in X. amieti and X. lenduensis, and lower dominant frequencies than X. ruwenzoriensis and X. amieti. The presence of a prehallux claw in X. eysoole further differentiates it from X. itombwensis, X. lenduensis, X. vestitus, and X. wittei. Description of the holotype.—Medium-sized (SVL 39 mm), moderately robust female (Table 1, Figs 6 and S3–S5); rostral tip blunt and rounded in dorsal view; eyes not projecting beyond margins of orbit in dorsal view and flush with dorsal margin of head in lateral view; subocular tentacle short, length less than half eye diameter; eye diameter 40% of interorbital distance, 87% of eye–narial distance, and 1.1 times distance from naris to rostral tip; internarial distance 40% of interorbital distance; no vomerine teeth (Fig 11). Skin smooth; small asperities on snout and scattered over much of body and limbs (most evident when dry); lateral-line plaques prominent on head and dorsal and lateral surfaces; both horizontally and vertically oriented lateral-line stitches on dorsum well defined. Relative lengths of fingers: III > IV > II > I; relative lengths of fingers when adpressed: III > II > I > IV; thigh length 93% of crus length; relative length of toes: IV > III > V > II > I; foot, but not toes, with many prominent pustules on ventral and lateral surfaces. Measurements.—Female specimens reach a maximum SVL of 52 mm (mean: 50 mm; n = 7), and males reach a maximum SVL of 41 mm (mean: 40 mm; n = 3). See S1 Table for more measurements. Coloration of holotype (in alcohol).—The dorsum and limbs are a medium grayish brown and lacking prominent markings (Figs 6 and S3–S5). The venter is pale grayish cream with scattered orange and gray spots on the ventral thighs. Coloration in life.—The dorsum and limbs of Xenopus eysoole is grayish brown with shades of green (Figs 7 and S6–S8); based on DCB’s field notes and photographs, the venter is a creamy white. Variation.—Variation in the lateral-line based on type specimens at MCZ (n = 13 except where noted, given as mean and range): orbital– 13 (10–14); oral– 11 (9–13); medial– 20 (17–23); lateral– 20 (18–23); ventral– 16 (13–18, n = 4). Vocalization.—Xenopus eysoole has either a trill-type or burst-type call (Table 4, Fig 8). These two call types may represent variation across populations, or differences in the social context or reproductive state when the calls were recorded. Karyotype.—Xenopus eysoole is dodecaploid with a karyotype of 2n = 12x = 108 (Fig 9). Habitat and range.—Xenopus eysoole is known from high elevations (~1400–2000 m) in the northern part of the Bamiléké Plateau in Cameroon, in the Bamenda-Banso Highlands (Fig 10). The type specimens were found in a small pool (approximately 1 m2; approximately 20 cm deep). This pool was located several meters from a fast-moving stream running through cultivated land in Elak Oku village. Astylosternus rheophilus, Cardioglossa pulchra, C. schioetzi, and Phrynobatrachus steindachneri [sensu 77] were found in a similar habitat at a nearby locality in Elak Oku village. Etymology.—In consultation with the Fon of Oku and his advisors, we have chosen a word derived from the Oku language as the species epithet. Roughly translated, the word eysoole (pronounced “ee-su-lay”) means “it will jump so hold it tightly,” and should be treated as a noun in apposition. Remarks.—Based on X-ray images, the holotype (MCZ A-138016) seem to have eaten or scavenged a similarly sized Xenopus specimen (likely the same species) before it was collected (S15 Fig), suggesting cannibalism in this species.

Xenopus (Xenopus) fraseri Boulenger 1905. Fraser’s Clawed Frog Dactylethra laevis (part) Günther ([78]; p. 2) Xenopus muelleri (part) Boulenger [79] Syntypes.—BMNH 1947.2.24.78–79, (previously BMNH 52.2.22.23–24), probably subadults, possibly males, “probably from Nigeria or Fernando Po”, coll. “Mr. Fraser”. We designate the larger specimen (BMNH 1947.2.24.78) as the lectotype. Referred Specimen.—Ghana: Northern Region, Wa: CAS 146198 (N 10.0585, W 2.5097°; 6 August 1975; coll. P. Williams. Diagnosis.—Because of both the morphological distinctiveness of the types and uncertainty in the type locality of X. fraseri, we restrict usage of X. fraseri to the type specimens and possibly one referred specimen (see below). Xenopus fraseri is distinctive among other living species of Xenopus in the combination of vomerine teeth and a prehallux claw. The presence of vomerine teeth differentiates X. fraseri from all living species of Xenopus except X. muelleri and the new tetraploid relative of X. muelleri described below [48], whereas the presence of the prehallux claw differentiates X. fraseri from X. muelleri and from some species in the amieti species group, including X. itombwensis, X. lenduensis, X. vestitus, and X. wittei. Because the name is restricted to the type specimens there is no information on the karyotype, advertisement call, or molecular variation. Description of lectotype (BMNH 1947.2.24.78).—Medium-sized (SVL 37 mm), somewhat robust, sex unknown (Table 1, Figs 12 and S9–S14); rostral tip rounded in dorsal view; eyes within margins of orbit in dorsal and lateral views; prominent subocular tentacle, length more than half eye diameter and extending nearly to upper lip; eye diameter 43% of interorbital distance, approximately equal to eye–narial distance, and approximately 1.2 times distance from naris to rostral tip; internarial distance 39% of interorbital distance; vomerine teeth present (Fig 12). Skin smooth and lacking asperities; lateral-line plaques prominent on dorsal and lateral surfaces and easily visible on the ventral surface; both horizontally and vertically oriented lateral-line plaques are well defined on dorsum and lateral surfaces of body; specimen missing all keratinous claws on left foot and only maintaining claw on third pedal digit on right foot. Relative lengths of fingers: III > IV ≈ II > I; relative lengths of fingers when adpressed: III > IV ≈ II > I; thigh length 90% of crus length; relative length of toes: III > IV > V > II > I; no asperities on plantar surface. Measurements.—The other syntype (now the paralectotype), BMNH 1947.2.24.79 (sex unknown) is somewhat smaller at 33 mm SVL. Coloration of lectotype (in alcohol).—Dorsum pale medium brown (color significantly faded) without prominent markings (S9–S14 Figs). Neuromast “stitches” are unpigmented and distinct against dorsal coloration. Venter pale yellowish gray with mostly faded melanocytes. Variation.—Variation in the lateral-line based on the lectotype and paralectotype (except when noted n = 3, given as mean and range): orbital– 13 (13–13, n = 1); oral– 14 (13–15); medial– 21 (21–21); lateral– 23 (21–25); ventral– 24 (24–24, n = 1). The paralectotype is similarly faded in coloration and missing all keratinous claws on both feet. Vocalization.—Unknown. Karyotype.—Unknown. Habitat and range.—Boulenger [47] described this species based on specimens previously cited as Xenopus muelleri [79], and before that as X. laevis, which was Dactylethra laevis at that time [78]. These two specimens were collected in West Africa by Louis Fraser, “probably from Nigeria or Fernando Po.” The collection locality for the original syntypes of Xenopus fraseri has long been uncertain [47]. As part of our work, we were able to more certainly establish the region from which these specimens probably were collected. The two specimens later described as X. fraseri were added to the catalog of the British Museum of Natural History (now the Natural History Museum, London) in 1852 (J. Streicher, pers. comm.), when their collector, L. Fraser was based in Ouidah, today’s southern Benin. The original registration numbers were 1852.2.22.23 and 1852.2.22.24 for 1947.2.23.78 and 1947.2.23.79 respectively. Although these specimens do not have unique locality information associated with them, other specimens in the Fraser collection catalogued at the same time are from four localities: “Fernando Po” (now Bioko Island, Equatorial Guinea), “Whidah” (now Ouidah, southern Benin), “Abomey” (southern Benin), and “Budagery.” The last locality probably refers to Badagry, southwestern Nigeria, which was a former slave port located between Ouidah and Lagos that was under British control in the 1840s and 1850s [80]. Fraser was based in Ouidah between July 1851 and November 1852 and then Lagos [81], and Badagry is approximately halfway between these two towns. Other than Fernando Po, these localities are in or on the fringe of the Dahomey Gap, an area for which few collections exist for Xenopus. Furthermore, based on the shared presence of vomerine teeth (Fig 12) and a prehallux claw, we have identified another specimen that is probably assignable to X. fraseri (CAS 146198) from Wa, northern Ghana, which lies to the west of the Dahomey Gap but is also in Sahel habitat. For these reasons we conclude that the type locality of X. fraseri is most likely from southern Benin or southwestern Nigeria, and not from Bioko Island.

Xenopus (Xenopus) kobeli, new species. urn:lsid:zoobank.org:act:EFB69AA8-54A3-4052-B058-D64184E1FE3C Kobel’s Clawed Frog Xenopus sp. nov. VIII sensu Tymowska (1991) Holotype.—MCZ A-148037 (BJE 3075), adult female, Republic of Cameroon, Centre Region, Meganme village, N 4.6116°, E 12.2254°, 637 m, 22 October 2011, coll. B. J. Evans, D. M. Portik, M. LeBreton (Figs 6 and S3–S5). Paratypes.—Republic of Cameroon: Centre Province: MCZ A-148035, male, MCZ A-148036, MCZ A-148038-9, females, same data as holotype; MCZ A-148059, MCZ A-148063, females, Balaoungao, N 5.2043° E 10.4289°, 1516 m, 25 October 2011; MCZ A-148060–62, males, same locality data, coll. B. J. Evans, D.M. Portik, S. Menzepoh; Ouest Region: MCZ A-148065–66, males, N 5.5703°, E 10.6097°, 1127 m, 26 October 2011, coll. B.J. Evans, D.M. Portik, S. Menzepoh; Est Region: NMP6V 74714/1, subadult male, Malapa, N 2.1028°, E 15.3566°, 388 m, 31 May, 2010, coll. V. Gvoždík, O. Kopecký. Diagnosis.—Analysis of mitochondrial data suggests that a portion of the allopolyploid genome of X. kobeli is closely related to X. ruwenzoriensis in the Albertine Rift and that the rest of its genome is most closely related to that of other species of the amieti species group in Cameroon. This species is thus distinguished from all others by unique nucleotide substitutions in mitochondrial and autosomal DNA and in the unique combination of ancestral genomes from which it is derived. Similar to X. ruwenzoriensis, X. longipes, and X. eysoole (described above), X. kobeli is distinguished from most closely related species by being dodecaploid. The trill-type call of X. kobeli distinguishes it from X. ruwenzoriensis, X. amieti, X. lenduensis, and X. pygmaeus Table 4,[42]. The fewer number of pulses in the call distinguishes X. kobeli from the trill-type calls of X. vestitus and its higher dominant frequency distinguishes it from the trills of X. allofraseri (described above) and X. wittei Table 4,[42]. Xenopus kobeli is distinguished from X. eysoole by having a trill-type instead of a burst-type call, and from the new tetraploid species in the amieti species group described below by fewer pulses per call. Both X. longipes and X. pygmaeus have smaller adult body size than X. kobeli; based on specimens we examined, maximum female SVL is 34 mm in X. longipes and 36 mm in X. pygmaeus, in comparison to 47 mm in X. kobeli. The presence of a prehallux claw in X. kobeli further differentiates it from X. itombwensis, X. lenduensis, X. vestitus, and X. wittei. Description of the holotype.—Medium-sized (SVL 42 mm), moderately robust female (Table 1, Figs 6 and S3–S5); rostral tip rounded in dorsal view; eyes projecting just beyond margins of orbit in dorsal view and beyond dorsal margin of head in lateral view; subocular tentacle moderately long, length slightly more than half of eye diameter and extending half way to upper lip; eye diameter 38% of interorbital distance, 82% of eye–narial distance, and approximately equal distance from naris to rostral tip; internarial distance 40% of interorbital distance; vomerine teeth absent (Fig 11). Skin smooth and lacking asperities; lateral-line stitches most prominent on dorsal and lateral surfaces and extending onto ventral surface; both horizontally and vertically oriented lateral-line stitches well defined on dorsum and lateral surfaces of body, but difficult to discern ventrally. Relative lengths of fingers: II > III ≈ IV > I; relative lengths of fingers when adpressed: II > III > I > IV; thigh length approximately equal to crus length; relative length of toes: IV > III > V > II > I; few scattered asperities on plantar surface. Measurements.—Female specimens reach a maximum SVL of 47 mm (mean: 44 mm; n = 6), and males reach a maximum SVL of 38 mm (mean: 35 mm; n = 6) (Table 2). Coloration of holotype (in alcohol).—Dorsum medium gray brown with a few dark brown irregularly shaped spots on the posterior dorsum (Figs 6 and S3–S5). Neuromast “stitches” are unpigmented and well defined against dorsal and lateral coloration (Figs 6 and S3–S5). Venter yellowish cream with many small and punctate orange spots and a few scattered patches of dark melanocytes on the venter and hind limbs. Many diffuse melanocytes on the plantar surface, but they are not developed into spots. Pedal webbing unpigmented. Pale gray interocular region, not well defined into an interocular bar. Coloration in life.—Based on color photographs of MCZ A-148062–63 (Figs 7 and S6–S8), the dorsum and limbs of Xenopus kobeli are brownish gray with medium gray or brown markings, and sometimes with a pale creamy gray interocular region. Variation.—Variation in the lateral-line based on specimens at MCZ (n = 12 except where noted, given as mean and range): orbital– 11 (10–12, n = 9); oral– 16 (14–19); medial– 19 (16–22); lateral– 15 (13–21, n = 4); ventral– 15 (13–21, n = 4). Vocalization.—Xenopus kobeli has a trill-type call (Table 4; Fig 8). Karyotype.—Xenopus kobeli is dodecaploid with a karyotype of 2n = 12x = 108 Fig 9, [51]. Habitat and range.—Some populations of X. kobeli are known from standing water on the southern Bamiléké Plateau at sites between 1100 and 1500 m (Fig 10), whereas others are known from similar habitats in central or southeastern Cameroon at lower elevations (~380 m). Etymology.—This species epithet is a patronym in honor of Hans Rudolf Kobel for his contributions to our understanding of the genetics and diversity of Xenopus [82].

Xenopus (Xenopus) parafraseri, new species. urn:lsid:zoobank.org:act:7DD73695-4F8F-40BF-A96C-92448EA9D973 Upland Clawed Frog Xenopus cf. fraseri 2 sensu Evans et al. [23] Xenopus fraseri-like Tinsley et al. [1] Xenopus fraseri Conlon et al. [83] Holotype.—MCZ A-148034 (field no. BJE 3068), adult female, Republic of Cameroon, Centre Region, Mfoundi Department, Old Douala Road, N 3.7931°, E 11.4170°, 715 m, 21 October 2011, coll. B. J. Evans, M. T. Kouete, D. M. Portik (Figs 6 and S3–S5). Paratypes.—Republic of Cameroon: Centre Region: MCZ A-148027, 148029, 148031–32, 148034, females, same locality data as holotype; MCZ A-148028, 148030, 148033, males, same locality data as holotype; NMP6V 74556/1, female, Ebogo, N 3.3913° E 11.4663°, 628 m, 18 October 2009, coll. V. Gvoždík; Est Region, coll. D. C. Blackburn, B. D. Freiermuth, G. F. M. Jongsma, M. T. Kouete, D. M. Portik, R. D. Tarvin; CAS 253332–33, Doumzok community forest, N 2.6445° E 14.0312°, 530 m, 13 June 2013; CAS 253366–70, Mebam community forest, N 2.6106° E 14.0234°, 550 m, 14 June 2013; Sud Region, coll. D. C. Blackburn, B. D. Freiermuth, G. F. M. Jongsma, M. T. Kouete, D. M. Portik, R. D. Tarvin; CAS 253589–91, Dja Faunal Reserve, Mekas, N 3.1738° E 12.5271°, 648 m, 21 June 2013; CAS 253609, Dja Faunal Reserve, near Mekas, N 3.1983° E 12.5228°, 23 June 2013. Gabonese Republic: NCSM 78872, Ogooué-Ivindo Province, N 0.0426° E 12.2983°, 17 October 2011, coll. B. L. Stuart, R. C. Bell, T. Ogombet, U. Eyagui, P. Endazokou; 78877–78, same collection data, 21 October 2011. Referred Specimens.—Republic of Congo: Cuvette-Ouest Department: Ndjoko, Ondou Forest, N 0.2518° E 14.1612° (NMP6V 75140/1, 1 female), N 0.2686° E 14.1592° (NMP6V 75140/2-5, 4 juveniles), N 0.2677° E 14.1618° (NMP6V 75140/6–7, 2 juveniles), 420–425 m, 11–17 January 2012, coll. V. Gvoždík. Diagnosis.—Xenopus parafraseri is a tetraploid species, with a trill-type call, that exhibits all of the morphological features of subgenus Xenopus described above. The two other closely related tetraploid species can be distinguished from X. parafraseri by unique nucleotide substitutions in mitochondrial and autosomal DNA (Figs 1 and 2 and S1 and S2), because X. pygmaeus is smaller and has a burst-type call, and X. allofraseri has a call with fewer pulses with longer interpulse intervals as well as lower dominant frequencies. Based on sequenced samples, our records of X. parafraseri are all from localities > 400 m, whereas those of X. allofraseri range from 2–1169 m (S2 Table). Xenopus parafraseri differs from the other species within the amieti species group by being larger than X. longipes and X. pygmaeus (Table 4), having a trill-type call in contrast to the burst-type call type of X. amieti, X. lenduensis, X. pygmaeus, and X. ruwenzoriensis, or the biphasic call type of X. itombwensis. The presence of a prehallux claw in X. parafraseri further differentiates it from X. itombwensis, X. lenduensis, X. vestitus, and X. wittei. Both X. parafraseri and X. allofraseri have previously been referred to as X. fraseri, yet both are different because of the presence of vomerine teeth in X. fraseri (see discussion of the X. fraseri type specimens above). Xenopus parafraseri is further distinguished from several species of Xenopus in the amino acid sequences of a PGLa and a CPF–RP peptide in epinephrine stimulated skin secretions [83]. Description of the holotype.—Medium-sized (SVL 41 mm), moderately robust female (Table 1, Figs 6 and S3–S5); rostral tip rounded in dorsal view; eyes projecting just beyond margins of orbit in dorsal view and beyond dorsal margin of head in lateral view; subocular tentacle moderately long, length slightly more than half of eye diameter and extending half way to upper lip; eye diameter 43% of interorbital distance, 86% of eye–narial distance, and 1.2 times the distance from naris to rostral tip; internarial distance 45% of interorbital distance; vomerine teeth absent (Fig 12). Skin smooth and lacking asperities; lateral-line stitches most prominent on dorsal and lateral surfaces and extending onto the ventral surface; both horizontally and vertically oriented lateral-line stitches well-defined on dorsum and lateral surfaces of body, but difficult to discern ventrally. Relative lengths of fingers: III > II ≈ IV > I; relative lengths of fingers when adpressed: III ≈ II > IV > I; thigh length 1.1 times crus length; relative length of toes: IV > III ≈ V > II > I; few scattered asperities on plantar surface. Measurements.—Female specimens reach a maximum SVL of 42 mm (mean: 37 mm; n = 14), and males reach a maximum SVL of 38 mm (mean: 34 mm; n = 3). See S1 Table for more measurements. Coloration of holotype (in alcohol).—Dorsum medium grayish brown with prominent dark brown mottling behind head, with some darker mottling on limbs (Figs 6 and S3–S5). Well defined pale brown region extending from midpoint of eyes to posterior of skull, with a medium brown snout. Neuromast stitches are unpigmented and distinct against dorsal coloration. Venter pale dusky cream, with many scattered orangish brown melanocytes and dark brown spots across throat, venter, and hind limbs. Pedal webbing with scattered dark melanocytes. Coloration in life.—Based on color photographs of NCSM 78877 and CAS 249961 (Figs 7 and S6–S8) and notes from other specimens, the dorsum and limbs of Xenopus parafraseri range from pale to olive-gray with a medium gray interocular bar and a creamy interocular region. A creamy occipital region bordered posteriorly by black markings is often present but uniformly colored specimens are also common. Variation.—Variation in the lateral-line based on specimens from Gabon (except when noted n = 32, given as mean and range): orbital– 11 (9–13, n = 30); oral– 10 (8–12; n = 29); medial– 16 (11–19); lateral– 18 (15–21); ventral– 16 (14–19, n = 17). Vocalization.—Xenopus parafraseri has a trill-type call (Table 4, Fig 8) [42]. Karyotype.—Xenopus parafraseri is tetraploid with a karyotype of 2x = 4n = 36 (Fig 9). Habitat and range.—Xenopus parafraseri occurs in southern Cameroon, central and eastern Gabon, and northwestern Republic of Congo (Fig 10). Specimens in Cameroon were collected from pools of water in agricultural plots (CAS 253767–70) and forests, and found in syntopy with X. mellotropicalis. Specimens in the Congo were found in swampy areas in pristine primary rainforest. Etymology.—For the species epithet, we have appended the Greek word παρά (para), meaning “near”, to fraseri. Both this taxon name, Xenopus parafraseri, and X. allofraseri are named to maintain the historical taxonomic relationship of these populations to the taxon X. fraseri, while recognizing that they are different and distinct evolutionary lineages.