Milnesium M. krzysztofi M. reductum Tumanov, 2006 M. reticulatum M. tardigradum M. tetralamellatum Pilato & Binda, 1991 M. beasleyi M. krzysztofi M. reticulatum M. krzysztofi M. beasleyi M. krzysztofi M. beasleyi M. krzysztofi M. reticulatum M. reticulatum M. beasleyi M. reticulatum M. beasleyi M. reticulatum M. beasleyi M. katarzynae M. beasleyi M. katarzynae M. beasleyi M. katarzynae M. beasleyi M. katarzynae M. beasleyi M. katarzynae M. beasleyi M. katarzynae M. beasleyi M. katarzynae M. alabamae M. beasleyi M. alabamae M. beasleyi M. alabamae M. granulatum M. beasleyi M. granulatum M. beasleyi M. granulatum M. beasleyi M. reductum M. beasleyi M. reticulatum M. tardigradum M. tardigradum M. beasleyi M. tardigradum M. tetralamellatum M. tetralamellatum M. beasleyi M. tetralamellatum M. beasleyi Milnesium M. beasleyi Current knowledge on Turkish tardigrades with a description of Milnesium beasleyi sp. nov. (Eutardigrada: Apochela: Milnesiidae, the granulatum group) Kaczmarek, Łukasz Jakubowska, Natalia Michalczyk, Łukasz Zootaxa 2012 3589 49 64 [151,374,1827,1852] Polychaeta Aphroditidae Milnesium Animalia Phyllodocida 1 50 Annelida species beasleyi sp. nov.   Material examined. Holotype(female) and 6 paratypes( 5 femalesand 1 male) in sample A.   Description (measurements and statistics in Table 1).Body ( Figs 1–2) yellowish (before and after preparation). Eyes were absent in 5 of the 7 examined individuals (=71%, including the male), after mounting in Hoyer’s medium. Cuticle without granulation or pores, but the dorsum covered with numerous tiny and shallow polygonal or rounded depressions (pseudopores), 0.1–0.4 μm in diameter ( Figs 3–5). These depressions, visible under PCM as light spots with undefined edges, are a manifestation of a fine reticulum, therefore placing the species within the  granulatumgroup (Michalczyk et al.2012ab); see also the Differential Diagnosis below. Weak sculpturing is also present on the outer surfaces of all legs ( Fig. 5). Six peribuccal papillae (ventral papilla smallest) and six peribuccal lamellae around the mouth opening present. Two cephalic papillae positioned laterally. Bucco-pharyngeal apparatus of the  Milnesium type( Figs 6–7). Buccal tube slightly funnel-shaped, wider anteriorly (on average the posterior diameter is 92% of the anterior diameter). Pharyngeal bulb elongated, pearshaped and without placoids or septulum. Claws of the  Milnesium type, slender ( Figs 8–9). Primary branches on all legs with small, but distinct accessory points detaching from the branch at its greatest curvature ( Fig. 9). Secondary claws of all legs without rounded basal thickenings. Secondary branches of external claws I–III and posterior claws IV with two points, and secondary branches of internal claws I–III and anterior claws IV with three points ( i.e.claw configuration: [2-3]- [3-2], Figs 8–9). Single, long transversal, cuticular bars under claws I–III present ( Fig. 8).   FIGURES 1–2.  Milnesium beasleyi  sp. nov.habitus: 1—female (holotype), 2—male (paratype). Both dorso-ventral views, scale on Fig. 2 same as on Fig. 1.   FIGURES 3–5.  Milnesium beasleyi  sp. nov.cuticular sculpture: 3–4—dorsal cuticle at increasing magnification, 5—cuticle of the outer part of leg III. All paratypes.   FIGURES 6–7.  Milnesium beasleyi  sp. nov.buccal apparatus: 6—female (holotype), 7—male (paratype). Both dorso-ventral views, scale on Fig. 7 same as on Fig. 6. Although eggs were not found, we should probably expect them to be oval, smooth and deposited in exuvium, i.e. following the character of all currently known  Milnesiumspecies in which eggs were described.   Remarks.We only found one male, which was generally similar to the females. However, apart from the diagnostic secondary sexual trait in this genus (secondary branches of claws I in the shape of large and robust hooks, Fig. 10), the male also differed from the females of this species by a narrower buccal tube and shorter papillae, both in absolute and relative terms (see Table 1).   TABLE 1.Measurements and ptvalues of selected morphological structures of six females and a male from the type population of  Milnesium beasleyi  sp. nov.Statistics include females only (N = number of specimens or structures measured; RANGE = the smallest and the largest structure found among all specimens measured; SD = standard deviation; * = abnormally developed spurs (see also remarks)). N RANGE MEAN SD HolotypeMale CHARACTER µm ptµm ptµm ptµm ptµm ptAlthough the majority of specimens had a typical claw configuration [2-3]-[3-2], two females exhibited minor claw abnormalities ( Fig. 11). One female had small basal spurs on external claws I-III and posterior claws IV ([3- 3]-[3-3]) and the other female had small basal spurs on posterior claws IV only ([2-3]-[3-3]). These basal spurs were always less curved (sometimes completely straight) and smaller than those on other claws (<2.0 μm vs.>3.0 μm) and thus could be without doubt classified as developmental aberrations. Nevertheless, the presence of such claw abnormalities highlights the importance of examining a number of  Milnesiumspecimens to ensure correct identifications.    Body length 5 418–810  1240–1674 653  1513 162  173 602  1465 481  1336  Peribuccal papillae length 5 7.1–10.0  19.6–21.3 8.7  20.5 1.1  0.7 8.3  20.2 4.0  11.1  Lateral papillae length 5 7.5–10.3  19.6–23.7 9.4  21.5 1.1  1.6 ?  ? 5.1  14.2  Buccal tube  Length 6 33.7–49.5 – 43.6 – 6.1  – 41.1  – 36.0  –  Stylet support insertion point 6 22.1–31.4  61.6–65.6 28.0  64.2 3.7  1.6 26.9  65.5 23.6  65.6  Anterior width 6 11.9–20.1  35.3–41.8 17.4  39.7 3.3  2.4 15.8  38.4 10.4  28.9  Standard width 6 10.5–19.3  31.2–39.8 16.2  36.9 3.5  3.3 14.4  35.0 9.3  25.8  Posterior width 6 11.2–19.2  33.2–39.6 16.0  36.6 3.0  2.3 14.3  34.8 9.9  27.5  Standard width/buccal tube length ratio 6 31%–40%  – 37% – 3%  – 35%  – 26%  –  Posterior/anterior width ratio 6 90%–96%  – 92% – 3%  – 91%  – 95%  –  Claw 1 lengths  External primary branch 5 13.9–22.0  40.6–46.7 18.7  43.8 3.5  2.7 16.7  40.6 ?  ?  External base + secondary branch 6 11.2–18.5  33.2–38.7 15.6  35.7 2.8  2.1 13.8  33.6 13.8  38.3  External spur* 1 1.8–1.8  4.4–4.4 1.8  4.4  –  –  –  –  –  –  Internal primary branch 4 16.1–20.8  39.2–44.0 18.8  41.9 2.2  2.3 16.1  39.2 ?  ?  Internal base + secondary branch 6 10.8–17.6  31.9–36.3 14.8  33.8 2.6  1.7 13.1  31.9 13.7  38.1  Internal spur 6 3.1–6.0  9.2–12.4 4.7  10.8 1.1  1.4 3.8  9.2 ?  ?  Claw 2 lengths  External primary branch 6 16.8–26.9  46.2–54.3 21.8  49.8 3.8  2.7 19.0  46.2 17.2  47.8  External base + secondary branch External spur* 6 1 12.2–19.3 1.9–1.9  32.1–39.8 4.6–4.6 16.1 1.9  36.8 4.6 2.9 –  2.6 – 13.2 –  32.1 – 14.0 –  38.9 –  Internal primary branch 6 15.0–25.0  44.5–50.5 20.4  46.7 3.5  2.0 18.8  45.7 17.7  49.2  Internal base + secondary branch 6 11.6–18.1  33.2–37.9 15.3  35.1 2.5  1.7 14.3  34.8 13.4  37.2  Internal spur 6 4.2–7.4  11.0–15.3 5.7  12.9 1.4  1.8 5.1  12.4 6.3  17.5  Claw 3 lengths  External primary branch 4 18.5–26.1  45.0–52.7 22.2  49.2 3.5  3.3 18.5  45.0 20.0  55.6  External base + secondary branch 5 13.9–19.0  33.8–39.7 17.1  37.4 2.2  2.4 13.9  33.8 13.7  38.1  Internal primary branch 4 18.7–25.5  45.5–51.5 21.6  48.0 3.0  2.7 18.7  45.5 19.0  52.8  Internal base + secondary branch 5 13.6–17.9  32.7–36.9 16.1  35.2 1.8  2.1 13.6  33.1 12.4  34.4  Internal spur 4 3.7–7.1  7.7–14.3 5.4  11.5 1.7  3.1 4.2  10.2 7.0  19.4  Claw 4 lengths  Anterior primary branch 5 17.9–28.4  53.1–57.4 23.6  55.3 4.1  1.9 23.3  56.7 21.0  58.3  Anterior base + secondary branch 5 12.7–19.9  37.5–42.5 17.1  39.9 3.1  2.3 15.4  37.5 14.0  38.9  Anterior spur 5 4.2–6.7  10.3–13.6 5.1  12.1 1.0  1.6 5.2  12.7 5.5  15.3  Posterior primary branch 5 20.2–31.8  59.0–64.2 25.9  60.8 4.4  2.2 25.3  61.6 22.5  62.5  Posterior base + secondary branch 5 14.1–22.3  39.7–46.7 18.3  42.8 3.3  2.7 16.3  39.7 15.2  42.2  Posterior spur* 2 1.8–1.9  4.0–4.4 1.9  4.2 0.1  0.3 – – – –   FIGURES 8–9.  Milnesium beasleyi  sp. nov.normally developed female claws: 8—claws II (paratype), 9—claws IV (holotype), scale on Fig. 9 same as on Fig. 8.   FIGURES 10–11.  Milnesium beasleyi  sp. nov.10—modified claws I of the only male found in the sample (paratype), 11—posterior secondary branch IV with an abnormal, short and straight point c (indicated by an arrow, paratype), scale on Fig. 11 same as on Fig. 10.  Locus typicus.37°14′52′ N, 31°55′42′ E, 1480 masl: the Tinaztepe Mağarasi cave, near the road D695, 25 km to the south from the city Seydişehir, Konya Province, Turkey, moss from stone.   Etymology.We dedicate this species in memory of Professor Clark Beasley ( 06.06.1942.- 24.06.2012.), an American tardigradologists and the first ZootaxaAssociate Editor for Tardigrada. Clark was also our friend and colleague who supported us throughout our adventure with tardigrades, especially in the early stages, and for this we will always be grateful to him.    Typedepositories. Holotype(slide T1/12) and six paratypes(slides T1/3, T1/11, T1/13 and T1/14) mounted in Hoyer’s medium are preserved at the Department of Animal Taxonomy and Ecology, A. Mickiewicz University, Umultowska 89, 61–614 Poznań, Poland.  Differential diagnosis.  Milnesium beasleyi  sp. nov.by having a sculptured dorsal cuticle belongs to the  granulatumgroup within the genus ( Michalczyk et al.2012a, b). Including this new species, the group now consists of six species, which is equivalent to 32% of all known  Milnesiumtaxa. The remaining five species with sculptured cuticle are:  M. alabamae Wallendorf & Miller, 2009,  M. granulatum Ramazzotti, 1962,  M. katarzynae Kaczmarek et al., 2004,  M. krzysztofi Kaczmarek & Michalczyk, 2007and  M. reticulatum Pilato et al., 2002. Cuticular sculpture in all known species of the group appears under PCMas bright dots or polygons with blurred edges. Scanning Electron Microscope (SEM) observations revealed that these dots/polygons are in fact shallow depressions in the cuticle (pseudopores) and therefore their edges can never be fully focused under PCM( Kaczmarek & Michalczyk 2007, see also Michalczyk & Kaczmarek 2006). Interestingly, these depressions have in past been mistaken for granulation—hence the name of the nominal species for the group (see Michalczyk et al.2012a). In all species of the group (apart from  M. beasleyi  sp. nov.) the cuticular depressions are densely arranged and the cuticle between very often forms a reticulum. In the new species, however, the depressions are less dense and more irregularly scattered. Large distances between the depressions, therefore, do not create an apparent reticulum.    Milnesiumspecies can also be grouped by claw configuration ( Michalczyk et al.2012a), and the new species, with the [2-3]-[3-2] configuration, falls into the second group in terms of species number (6 of 18 (33%), including the new species). Other members of the [2-3]-[3-2] group are:  M. krzysztofi,  M. reductum Tumanov, 2006,  M. reticulatum,  M. tardigradum sensu stricto Doyère, 1840and  M. tetralamellatum Pilato & Binda, 1991. Currently, excluding  M. beasleyi  sp. nov., there are only two species with both sculptured cuticle and the [2-3]- [3-2] claw configuration:  M. krzysztofiand  M. reticulatum. The new species differs specifically from:   M. krzysztofiby: smaller and more scattered cuticular depressions (diameter 0.1–0.4 µm in  M. beasleyi  sp. nov. vs.0.5–1.5 µm in  M. krzysztofi) that do not form the reticulum, the absence of rounded basal thickenings under secondary claws, and slightly longer spurs on internal claws I–III and the anterior spur IV (in normally developed animals, see remarks) (3.1–7.4 µm [ pt 9.2–15.3] in  M. beasleyi  sp. nov. vs.2.5–3.4 µm [ pt 8.0–10.1] in  M. krzysztofi).   M. reticulatumby: the lack of dorsal gibbosities, the presence of six peribuccal lamellae (four in  M. reticulatum), a larger body length (418–810 µm in  M. beasleyi  sp. nov. vs.270–405 µm in  M. reticulatum), stylet supports inserted in a more anterior position ( pt 61.6–65.6in  M. beasleyi  sp. nov. vs. 68.5–69.8in  M. reticulatum), the absence of rounded basal thickenings under secondary claws and longer claws on legs II–IV (both in absolute and relative terms).   M. beasleyi  sp. nov.is similar to the species listed below by having a sculptured dorsal cuticle, but differs from them by:   M. katarzynae: a different claw configuration ([2-3]-[3-2] in  M. beasleyivs.[2-2]-[2-2] in  M. katarzynae), smaller cuticular depressions (diameter 0.1–0.4 in  M. beasleyi  sp. nov. vs.0.5–1.0 µm in  M. katarzynae), the absence of rounded basal thickenings under secondary claws, a larger adult body length (418–810 µm in  M. beasleyi  sp. nov. vs.285–295 µm in  M. katarzynae), stylet supports inserted in a more anterior position [ pt 61.6–65.6in  M. beasleyi  sp. nov. vs. 73.3–78.3in  M. katarzynae], a longer buccal tube ( 33.7–49.5 in  M. beasleyi  sp. nov. vs. 28.5–30.4 in  M. katarzynae), a wider standard diameter of the buccal tube (10.5–19.3 µm [ 31.2–39.8] in  M. beasleyi  sp. nov. vs.6.6–7.6 µm [ 21.7–26.6] in  M. katarzynae), and longer external claws on legs I–IV (both in absolute and relative terms).   M. alabamae: a different claw configuration ([2-3]-[3-2] in  M. beasleyivs.[3-3]-[3-3] in  M. alabamae), the presence of accessory points on primary branches and smaller cuticular depressions (diameter 0.1–0.4 µm in  M. beasleyi  sp. nov. vs.0.5–1.0 µm in  M. alabamae).   M. granulatum: a different claw configuration ([2-3]-[3-2] in  M. beasleyivs.[3-3]-[3-3] in  M. granulatum), smaller cuticular depressions (diameter 0.1–0.4 µm in  M. beasleyi  sp. nov. vs.0.5–1.5 µm in  M. granulatum), and the absence of rounded basal thickenings under secondary claws.   M. beasleyi  sp. nov.is similar to the species listed below by having the [2-3]-[3-2] claw configuration, but differs from them all by having a sculptured dorsal cuticle and specifically from:   M. reductumby the presence of accessory points on the primary branches of claws, stylet supports inserted in a more anterior position (  61.6–65.6in  M. beasleyi  sp. nov. vs. 65.3–69.3in  M. reticulatum) and the absence of rounded basal thickenings under secondary claws,   M. tardigradum s. s.by peribuccal and cephalic lateral papillae of an almost equal length (peribuccal papillae longer in  M. tardigradum s. s.), longer lateral papillae (length 7.5–10.3 µm [ pt 19.6–23.7] in  M. beasleyi  sp. nov. vs.4.5–5.7 µm [ pt 13.8–17.2] in  M. tardigradum s. s.) and the absence of rounded basal thickenings under secondary claws.   M. tetralamellatumby the presence of six peribuccal lamellae (four in  M. tetralamellatum), a narrower buccal tube (10.5–19.3 µm [ pt 31.2–39.8] in  M. beasleyi  sp. nov. vs.22.3 µm [ pt 49.1] in  M. tetralamellatum(specimen 645 µm long)) and the absence of rounded basal thickenings under secondary branches of claws. Finally, we would like to draw the attention to the lack of basal thickenings at the bases of all secondary claws in  M. beasleyi  sp. nov.As this trait is not always reported in  Milnesiumdescriptions, currently it is not possible to conclude whether the new species is unique in this character. However, whenever this character has been addressed the thickenings have always been present, which makes baseless claws of  M. beasleyi  sp. nov.at least rare within the genus.