Pseudopus apodus levantinus ssp. nov. Jablonski, Ribeiro-Junior , Meiri, Maza, Mikulicek , Jandzik

Jablonski, Daniel, Ribeiro-Junior, Marco Antonio, Meiri, Shai, Maza, Erez, Kukushkin, Oleg V., Chirikova, Marina, Pirosova, Angelika, Jelic, Dusan, Mikulicek, Peter & Jandzik, David, 2021, Morphological and genetic differentiation in the anguid lizard Pseudopus apodus supports the existence of an endemic subspecies in the Levant, Vertebrate Zoology 71, pp. 175-200 : 175

Pseudopus apodus levantinus ssp. nov. Jablonski, Ribeiro-Junior, Meiri, Maza, Mikulicek, Jandzik

Holotype.

TAU-R 16895 (Fig. 6 , Table S7), an adult male, collected on the 16th of May, 2014 by Erez Maza, at Giv’at Ada (גבעת עדה), Israel (32.52°N, 35.01°E, Fig. 7 ). The molecular-genetic data of the holotype are available for ND2 (MW400924), Cyt b (MW400903), and microsatellites (Fig. 2 , Table S1, S2).

Paratypes.

TAU-R 17076 (MW400927, MW400906), an adult male, collected on 19th May 2014 by Talia Oron, at Biranit (בירנית), Israel (33.05°N, 35.34°E); TAU-R 17235 (MW400928, MW400907), an adult male, collected on 6th May 2015 by Ron Elazari, at Hadera (חֲדֵרָה), Israel (32.47°N, 34.89°E); TAU-R 17895 (MW400930, MW400909), an adult male, collected on 18th April 2016 by Ofer Shimoni, on Mt. Gilboa’ (הר הגלבוע), Israel (32.45°N, 35.43°E); TAU-R 19403, an adult male, collected on 16th May 2019 by Amir Arnon, at Ramat HaNadiv (רמת הנדיב), Israel (32.55°N, 34.95°E) (Figs 2 , 8 , Table S1, S2, S7).

Diagnosis.

A large Pseudopus (up to 610 mm snout-vent length, 1,367 mm total length and a mass of 1,100 g) that can be distinguished from the other two subspecies by a combination of the following characters (means followed by standard deviations; Table 4 ): (1) preanal scales (PAN) 10; (2) long body (SVL; 490.22 mm ± 76.15); (3) long tail (TL; 600.70 mm ± 107.04); (4) long head (HL1; from the ear aperture to the tip of the snout, 47.6 mm ± 7.99); pileus length (PL; mean 41.98 mm ± 6.78); and distance from the frontal shield to the tip of the snout (FSL; 17.27 mm ± 2.96); (5) wide head (HW1; maximum width, 32.04 mm ± 6.34); and width at the level of the posterior edge of the eyes orbits (HW2; 28.21 mm ± 4.95); (6) relatively long limb rudiments (RL; 5.55 mm ± 1.53) (see and Tables 3 and 4 for comparisons to P. a. apodus and P. a. thracius ).

Comparisons.

Pseudopus apodus levantinus ssp. nov. can be distinguished from P. a. apodus and P. a. thracius by having 10 preanal scales (PAN) in all morphologically examined specimens (vs. 5-11, with the mean of 8 in P. a. apodus , and 5-8 and mean of 6 in P. a. thracius ). Other differences can be used in combination to support differences between P. a. levantinus ssp. nov., P. a. apodus , and P. a. thracius : longer body (SVL; 610 mm maximum length and mean of 490.22 mm in P. a. levantinus ssp. nov., vs. 487 mm maximum and mean of 387.61 mm in P. a. apodus , and 410 mm maximum and mean of 360.06 mm in P. a. thracius ); longer distance between the ear aperture and the tip of the snout (HL1; 63.75 mm maximum length and mean of 47.60 in P. a. levantinus ssp. nov., vs. 58 mm maximum and mean of 38.61 mm in P. a. apodus , and 49.50 maximum and mean of 39.55 mm in P. a. thracius ); longer pileus (PL; 54.79 mm maximum length and mean of 41.98 in P. a. levantinus ssp. nov., vs. 46.30 mm maximum and mean of 35.82 mm in P. a. apodus , and 28.80 mm maximum and mean of 36.93 mm in P. a. thracius ); longer distance between the anterior frontal scale to the tip of snout (FSL; 23.39 mm maximum length and mean of 17.27 mm in P. a. levantinus ssp. nov., vs. 18 mm maximum and mean of 13.90 mm in P. a. apodus , and 15.98 mm maximum and mean of 12.58 mm in P. a. thracius ); wider head (HW1; 44.90 mm maximum width and mean of 32.04 mm in P. a. levantinus ssp. nov., vs. 36.5 mm maximum and mean of 24.47 mm in P. a. apodus , and 29.50 mm maximum and mean of 24.35 mm in P. a. thracius ); wider distance between the posterior edge of the orbits (HW2; 39.33 mm maximum width and mean of 28.21 mm in P. a. levantinus ssp. nov., vs. 28.76 mm maximum and mean of 19.44 mm in P. a. apodus , and 26.30 mm maximum and mean of 22.33 mm in P. a. thracius ); and longer limb rudiments (RL; 8.56 mm maximum length and mean of 5.55 mm in P. a. levantinus ssp. nov., vs. 6.55 mm maximum and mean of 3.5 mm in P. a. apodus , and 4.8 mm maximum and mean of 3.31 mm in P. a. thracius ). The remaining differences are presented in Tables 3 - 4 .

Description of the holotype.

An adult male ( TAU 16895; Fig. 6 , Table S7), specimen in a good state of preservation in 70% ethanol, midbody oval, and robust. All measurements of the holotype were taken post-preservation. Snout-vent length 532 mm, tail length 602 mm, weight 900 g. The number of transversal dorsal scale rows at midbody 12 (DST), the number of transversal ventral scale rows 10 (VST). Head large, length (HL1) 59.01 mm, clearly distinct from the neck. The pileus length is 50.52 mm, maximum head width 42.25 mm. The right part of the mouth is slightly open. The number of supralabials on both parts of the head is 11. Head and body scales smooth. Slightly keeled scales are visible on the ventral part of the tail. Rostral curved toward the top of the head. Frontal scale well visible and big with the length 17.73 mm and width 13.15 mm. Eyes oval, both closed. The ear and nose openings are visible. The hind limb rudiments are well visible with a length 7.42 mm. The complete tail is clearly differentiated from the body by the cloacal opening and 10 preanal scales on the ventral side. There are 154 subcaudal scale rows (SCR) overall, which is very similar to the specimens with regenerated tails. It has 106 longitudinal dorsal scale rows (DSL), and 121 ventral longitudinal scale rows (VSL).

The coloration of the holotype in life was not recorded. The coloration of the holotype in preservative is brownish or slightly orange with some of the scales on the body that have darker coloration creates an impression of tiny dark spots on the body. The head is lighter than the body, which is especially apparent on its dorsal side. The dorsal side of the body is light brown to gray, again with the impression of darker spots on the tail.

Variation.

Details on variation among the type specimens of P. a. levantinus ssp. nov. are presented in Table S7. The overall morphology and coloration of the paratypes ( TAU 17076, 17235, 17895) are very similar to that of the holotype; TAU 19403 was recently collected and has an evident dark-brown coloration pattern ( TAU 19403, Fig. 8 ).

Etymology.

No name is available for the glass lizards from the Levantine region. We hence suggest a new name, Pseudopus apodus levantinus , as a reference to the isolated and allopatric distribution of this subspecies exclusively in the Levant region. This region covers present-day (western) Syria, Lebanon, (north-western) Jordan, Israel, West Bank, Cyprus, and most of Turkey south-east of the middle Euphrates, which is almost identical to the known distribution range of the new subspecies. The term “Levant” is derived from the Italian “Levante”, meaning “rising” and implying the rising of the Sun in the east as an equivalent to the Arabic “al-Mashriq” (المشـرق) and the Hebrew “Mizrāḥ” ((מִזְרָחboth meaning “east”.

Distribution.

According to the genetic data, P. a. levantinus ssp. nov. occurs in southern Turkey, western Syria, northern and central Israel ( Jandzik et al. 2018; this study). Based on the published distribution data, the new subspecies could also be expected in the Mediterranean regions of Lebanon and north-western Jordan (In den Bosch et al. 1998; Disi et al. 2001; Hraoui-Bloquet et al. 2002; Rifai et al. 2005; Sindaco and Jeremcenko 2008), although we did not examine specimens from these regions. We therefore hypothesize that it ranges from southern Turkey (Hatay province) to central Israel (throughout the Mediterranean zone but excluding the deserts to the south and east; Figs 3 and S4b). Its northern distribution edge is presumably south of the Nur Mountains, the biogeographic barrier for the split between Anatolian and Levantine populations of other species ( Jandzik et al. 2013; Tamar et al. 2015; Jablonski and Sadek 2019; Šmíd et al. 2021). This, however, needs further investigation as there is a possibility that the new subspecies could be found further north in Mersin or Adana provinces (Figs 1 , S4 and Baran et al. 1988; Sindaco and Jeremcenko 2008; see also Hofmann et al. 2018). The southern limit of the subspecies range is near the border of the Negev desert, around Lahav (31.38°N, 34.87°E), and Kisufim (31.37°N, 34.40°E) in Israel, and Dhibam (31.47°N, 35.80°E) in Jordan (Fig. S4; Disi and Amr 1998; Roll et al. 2017, the Steinhardt Museum records).

Habitat and ecology.

The new subspecies is known in Mediterranean habitats of the Levant (see the type locality Fig. 7 ), preferring relatively well-shaded light woodlands (maquis and garrigue), dry and warm hillsides, stream banks, and agricultural fields. It is found from ~50 m below sea level (in Hefzibah, Israel; 32.52°N, 35.42°E) to the hilly sub-mountain areas in Israel, Lebanon, Syria, and Turkey (elevation maximum ~1000 m; In den Bosch et al. 1998; Disi et al. 2001; Rifai et al. 2005; Werner 2016; the Steinhardt Museum records). Pseudopus a. levantinus ssp. nov. is a leaf-litter diurnal carnivore (Bar and Haimovich 2012). It is oviparous and females typically lay a single clutch comprised of 3-12 eggs (data from the records of the Zoological Research Garden, Tel Aviv University). Its habitat characteristics, ecology, and diet were studied in Jordan and presented by Rifai et al. (2005), however data from other countries (see Distribution) are scarce (see Arbel 1984; In den Bosch et al. 1998; Bouskila and Amitai 2001; Bar and Haimovich 2012; Werner 2016).

Diversity.

The specimens analyzed in this study showed certain variation in both genetic and morphological markers. Some intra-lineage variation in P. a. levantinus ssp. nov. can be found in ND2 and Cyt b sequences as well as in microsatellites (see details in the Results). The haplotype pattern of nuclear genes PRLR and RAG1 indicates incomplete lineage sorting among all three subspecies, with one recorded heterozygote of P. a. levantinus ssp. nov. ( Jandzik et al. 2018). The calculated genetic distances among the three main mtDNA lineages (~2.6-4.0%) and the estimated time of divergence in Pseudopus mostly based on the nuclear loci (~8 to 3 Mya) support the subspecies status of the Levantine population ( Jandzik et al. 2018, Lavin and Girman 2019). This is in accordance with subspecific taxonomy of the closely related species Anguis colchica (approx. split between 2.8-2.5 Mya, genetic distances ~3.1-4.7% among defined subspecies/lineages based on ND2 mtDNA fragment; Gvoždík et al. 2010; Jablonski et al. 2016), while the genetic distance among the Anguis species are significantly higher (7.0% between A. fragilis and A. colchica and even 9.2% between A. fragilis and A. veronensis ; Gvoždík et al. 2013). We also recorded some variation in the measurements and coloration both among the three subspecies (Figs 5 , 9 ) and within P. a. levantinus ssp. nov. Males of P. a. levantinus ssp. nov. differ from females by having relatively wider heads (HW1-3, SW, IWM; Table S6). The coloration of this subspecies is similar to the nominotypic subspecies ( Rifai et al. 2005) but more data are needed to allow for drawing any more robust conclusions. A melanistic individual has been recorded in the Levantine population ( Jablonski and Avraham 2018).

Conservation.

Based on the data presented here, the distribution range of P. a. levantinus ssp. nov. covers approximately 30,000 km2. Together with human overpopulation and accelerated development in the Mediterranean parts of the Levant, high traffic density (most individuals nowadays brought to the Steinhardt Museum of Natural History in Tel Aviv are road-kills), development of mass tourism, extensive use of pesticides in the agricultural areas, proliferation of human commensals such as domestic cats, cattle egrets, dogs, rats, and golden jackals, and challenging political situation have potential to worsen the conservation status of this endemic subspecies. Pending a formal assessment, we preliminary recommend the IUCN category of Vulnerable (VU) based on the criteria A2c,e and strongly encourage further surveys benefiting from international collaboration allowing to open a dialogue across the conflict zones (e.g. EcoPeace Middle East, http://ecopeaceme.org; Roulin et al. 2017). Interestingly, recent data suggest that populations of P. apodus from the Levant were likely a part of modern human diet for millennia (Natufian culture, around 13,050 to 7,550 BC), which provides a rare piece of evidence of a long-term civilization pressure on the local biota ( Lev et al. 2020).

Figures

Figure 1. Geographic distribution of the samples and specimens used in the molecular-phylogenetic (A, B) and morphological analyses (C). Some specimens used in morphological analyses were from unknown localities, so those are not indicated in the map. Color scheme corresponds with the one used in Fig. 2 and follows Jandzik et al. (2018). For more details on samples, specimens, and localities, refer to Tables S 1 - 3. The distribution range follows Sindaco and Jeremcenko (2008) and Jandzik et al. (2018). The pictured individuals represent currently recognized subspecies originating from (left to right) Lefkimi, Greece (P. a. thracius), Hirbat Tzura, Israel (P. a. levantinus ssp. nov.), and Tyolyok, Kyrgyzstan (P. a. apodus). Photos by Daniel Jablonski and David David.

Figure 2. Phylogenetic relationships of Pseudopus apodus reconstructed from concatenated and single locus trees of ND 2 & Cyt b sequences and presented as collapsed Bayesian trees (A, C, E) and haplotype networks (B, D, F). The numbers at the tree nodes represent Bayesian Posterior Probabilities / Maximum Likelihood Bootstraps showing branch support (for details see Figures S 1 and S 2). The numbers at the end of main lineages or with haplotypes show the sampling code (DJab acronym) or GenBank numbers (see also Table S 1). Circle sizes in the haplotype networks are proportional to the relative frequency of the haplotypes and small black circles represent the missing haplotypes.

Figure 3. Maps showing the localities and sampling numbers related to the collection with DJab acronym or GenBank number (see Table S 1) of the genetically investigated populations of Pseudopus apodus levantinus ssp. nov. (ND 2 & Cyt b) in the Levant with the haplotype networks for each DNA marker. Circle sizes in the haplotype networks are proportional to the relative frequency of haplotypes and small black circles represent missing haplotypes. The expected distribution range (orange) of the Levantine subspecies (see also Figure S 4) follows Sindaco and Jeremcenko (2008) and Jandzik et al. (2018).

Figure 5. Principal Component Analysis (PCA) of the examined phenotypic characters among the adult specimens of Pseudopus apodus apodus (red dots), P. a. thracius (blue dots), and P. a. levantinus ssp. nov. (yellow dots): A - adult females and males together, B - females only, C - males only. The oval outlines correspond to 95 % confidence intervals. For A, PC 1 explains 98.7 % of variance, PC 2 0.59 %; for B, PC 1 explains 98.9 % of variance, PC 2 0.45 %; for C, PC 1 explains 98.6 %, PC 2 0.59 % (see details in Table S 5), respectively.

Figure 6. Holotype (TAU 16895) of Pseudopus apodus levantinus ssp. nov. from Giv'at Ada (הדע תעבג), Israel. A - dorsal view, B - ventral view, C - dorsal view of the head, D, E - lateral views, F - ventral view. For details and measurements see Tables S 1, S 2 and S 7.

Figure 7. Habitat of Pseudopus apodus levantinus ssp. nov. at the type locality Giv'at Ada (גבעת עדה) in Israel (photo by Ilana Rosenstein).

Figure 8. Paratypes of Pseudopus apodus levantinus ssp. nov. from Israel showing A - dorsal part of the body, B - ventral part of the body, C - dorsal part of the head, D - ventral part of the head. For the locality details and measurements see Tables S 1, S 2, and S 7.

Figure 9. Color and pattern variation in the Pseudopus apodus subspecies: A - adult male of P. a. apodus in a typical steppe habitat from Samarkand, Uzbekistan (photo by Daniel Jablonski). B - juvenile individual of P. a. apodus from Kyz-Kermen, Bakhchisarayi, Crimea (photo by Oleg V. Kukushkin). C - adult female of P. a. thracius from Dadia, Greece (photo by D. Jablonski). D - juvenile individual of P. a. thracius from National Park Paklenica, Croatia (photo by D. Jablonski). E - adult female of P. a. levantinus ssp. nov. from Antakya, Turkey (photo by David Jandzik). F - juvenile individual of P. a. levantinus ssp. nov. from Ness Ziyona, Israel (photo by David David).