Integrative description of Mesobiotus anastasiae sp. nov. (Eutardigrada, Macrobiotoidea) and first record of Lobohalacarus (Chelicerata, Trombidiformes) from the Republic of South Africa Author Tumanov, Denis V. 49E22A70-C27B-485B-941E-577666EE65F9 Department of Invertebrate Zoology, Faculty of Biology, Saint Petersburg State University, 199034, Universitetskaya nab. 7 / 9, Saint Petersburg, Russia. & Marine Research Laboratory, Zoological Institute of the Russian Academy of Sciences, 199034, Universitetskaja nab. 1, Saint Petersburg, Russia. d.tumanov@spbu.ru text European Journal of Taxonomy 2020 2020-12-04 726 726 102 131 http://dx.doi.org/10.5852/ejt.2020.726.1179 journal article 3766 10.5852/ejt.2020.726.1179 a3bd24c5-60d2-4617-ba43-a96c97382e64 2118-9773 5628080 CCB2BAEF-06D4-4FAD-A018-27A54553D3DA Mesobiotus anastasiae sp. nov. urn:lsid:zoobank.org:act: 8651D164-4CBC-41DD-A5C4-FF9E584F8088 Figs 1–6 , Tables 2–3 Etymology I dedicate this new species to my daughter, Anastasia. Material examined Holotype REPUBLIC OF SOUTH AFRICA • ♀ ; Cape Town , top of Table Mountain ; approx. 33°57′43.9″ S , 18°24′38.0″ E ; = 1000 m a.s.l. ; 10 Jan. 2008 ; I. Nikolaeva leg.; moss on soil, wet depression with a small temporary pond ; SPbU 256(8) . Paratypes REPUBLIC OF SOUTH AFRICA • 5 ♀♀ , 3 ♂♂ , 22 eggs ; same collection data as for holotype; SPbU 256(1) to 256(6) and 256(9) to 256(18) • 2 adults , 7 eggs ; same collection data as for holotype; SEM stub; SPbU_Tar16 . Comparative material PEOPLE’S REPUBLIC OF CHINA • 7 eggs , paratypes of Mesobiotus mauccii ( Pilato, 1974 ) ; Canton; moss on tree bark ; UNICT 2132 . Morphological description Body elongated ( Fig. 1 A–B) (morphometrics in Table 2 , raw morphometric data are provided in Supplementary file 2). Fresh specimens uncolored or whitish with slightly greenish gut content, transparent after fixation in Hoyer’s medium. Four specimens with eyes ( Fig. 1A ), usually welldiscernible after slide mounting, three specimens without eyes; in two specimens processed for DNA extraction the presence or absence of eyes was not registered. Dorsal cuticle in LM with poorly visible network-like pattern ( Fig. 2A ). SEM investigation revealed presence of flat tubercles all over the body surface ( Fig. 2 B–C). Additionally, the body surface is covered with regularly distributed small granules (invisible in LM; Fig. 2B ). In the dorso-lateral position, above the bases of the hind legs, zones of concentrated granules are present ( Fig. 2C ). All legs with granulated areas. Legs I–III with small granulated areas on the external surfaces, near the claw bases ( Fig. 4A ), invisible or extremely poorly visible in LM ( Fig. 4D , black arrowhead), the internal leg surfaces without granulation, with indistinct pulvinus ( Fig. 4B , white asterisk). Legs IV with better-developed granulation dorsally ( Fig. 2D ) and around the claw bases ( Fig. 4G ). Buccal-pharyngeal apparatus of Macrobiotus type ( Fig. 3A ) with ventral lamina and ten peribuccal lamellae. Oral cavity armature (OCA) of modified krynauwi type (according to Kaczmarek et al . 2020 ) with three bands of teeth visible in LM. Evident first (anterior) band consists of a single uneven line of relatively large and slightly longitudinally elongated teeth ( Fig. 3F , J–K, O, black arrowheads). Second band consists of a wide zone of small dot-like teeth ( Fig. 3G, I, L, N ), teeth of the anterior-most and (rarely) posterior-most rows of the second band are slightly larger than others, sometimes teeth of the anterior-most row are very slightly longitudinally elongated. Third band comprises three dorsal and three ventral transverse ridges ( Fig. 3H, J, M, O ). Medio-ventral ridge is usually more or less clearly divided into two separate parts ( Fig. 3J, O ). Pharyngeal bulb with apophyses, three macroplacoids and a large microplacoid ( Fig. 3A ). Macroplacoid length sequence is 2 <3=1. First macroplacoid is anteriorly Fig. 1. Mesobiotus anastasiae sp. nov. , total view. A . Holotype, ♀ (SPbU 256(8)). Dorso-ventral view, PhC. B . Paratype (SPbU_Tar16). Lateral view in SEM. Scale bars = 50 µm. Table 2. Summary of morphometric data for Mesobiotus anastasiae sp. nov. Measurements are given in µm, pt values in % (the pt index is the percentage ratio between the length of a structure and the length of the buccal tube).

CHARACTER	N	RANGE µm pt		MEAN µm pt		SD µm pt		Holotype µm pt
Body length	7	284–426	809–1002	366	909	44	83	426	1002
Buccopharyngeal tube
Buccal tube length Stylet support insertion point Buccal tube external width Buccal tube internal width Ventral lamina length	9 9 7 7 9	35.1–45.5 26.5–35.8 4.1–6.0 2.8–4.0 21.9–30.7	– 75.3–78.7 11.3–13.1 7.6–8.8 62.2–67.4	40.8 31.5 5.1 3.4 26.2	– 77.1 12.2 8.2 64.1	3.5 3.1 0.6 0.4 2.7	– 1.2 0.6 0.4 1.7	42.6 33.1 5.4 3.6 27.1	– 77.8 12.7 8.5 63.8
Placoid lengths
Macroplacoid 1 Macroplacoid 2 Macroplacoid 3 Microplacoid Macroplacoid row Placoid row	9 9 9 9 9 9	3.7–6.1 2.8–4.4 3.2–6.4 2.0–3.3 11.7–18.3 13.9–21.5	10.0–13.4 8.0–10.3 9.1–14.6 5.3–7.7 33.0–41.9 39.0–49.0	4.8 3.6 4.7 2.6 15.1 18.2	11.7 8.9 11.4 6.3 36.8 44.3	0.9 0.5 1.1 0.5 2.5 2.9	1.3 0.7 1.9 0.9 3.2 3.9	5.2 4.4 5.4 3.3 16.6 19.7	12.2 10.3 12.6 7.7 38.9 46.2
Claw 1 lengths
External primary branch External secondary branch Internal primary branch Internal secondary branch	5 5 7 7	7.6–10.8 5.9–8.0 7.3–10.5 5.9–8.3	21.6–24.5 16.8–19.1 20.8–23.8 16.1–18.2	9.4 7.2 9.0 7.0	23.1 17.7 22.2 17.3	1.2 0.8 1.0 0.8	1.2 0.9 1.0 0.9	9.9 7.4 9.4 6.8	23.4 17.5 22.0 16.1
Claw 2 lengths
External primary branch External secondary branch Internal primary branch Internal secondary branch	6 6 6 6	8.0–10.8 6.6–8.4 7.4–10.1 6.0–7.7	22.4–24.7 17.2–19.3 21.0–23.8 16.2–19.4	9.4 7.4 9.0 7.0	23.5 18.5 22.6 17.5	1.0 0.7 1.1 0.6	0.9 0.7 1.1 1.1	10.2 8.0 10.1 7.7	23.9 18.9 23.8 18.1
Claw 3 lengths
External primary branch External secondary branch Internal primary branch Internal secondary branch	6 6 7 7	7.8–10.9 6.1–8.8 7.1–10.7 5.8–8.2	22.0–25.1 17.2–20.1 20.2–24.6 16.4–18.4	9.5 7.5 9.4 7.2	23.5 18.4 22.8 17.4	1.1 0.9 1.3 0.9	1.3 1.1 1.4 0.9	10.3 8.0 10.1 7.8	24.1 18.7 23.8 18.4
Claw 4 lengths
Anterior primary branch Anterior secondary branch Posterior primary branch Posterior secondary branch	6 6 5 5	9.8–12.3 7.0–8.6 11.7–12.9 8.0–9.4	24.7–28.2 17.0–20.3 27.2–30.2 18.7–20.6	11.1 7.9 12.3 8.6	26.4 18.8 28.6 19.9	1.0 0.6 0.6 0.6	1.2 1.1 1.2 0.7	11.4 8.6 12.9 8.7	26.9 20.3 30.2 20.4

narrowed, third macroplacoid without distinct subterminal constriction, but with strong terminal protrusion, directed towards pharynx lumen ( Fig. 3 B–E, black arrowheads). Claws of Mesobiotus type with minute stalk, distinct distal part of the basal portion, short common tract and developed internal septum, defining a distal part. Primary and secondary branches diverge at a point near half the claw height, main branches with long accessory points, which at a large distance from the main claw ( Fig. 4C , E–F, H). Claws of fourth pair of legs slightly longer than claws of first three pairs of legs ( Fig. 4H ). All claws with smooth lunules ( Fig. 4C , E–F, H). Anterior (internal) and posterior (external) claws of legs IV are similar in shape, with equally sized lunules. Poorly developed bar-like cuticular thickenings are present below claw bases of the first three pairs of legs ( Fig. 4C , black arrowhead). Claws of legs IV are connected with a wide horseshoe-like structure ( Fig. 4F , black arrowhead). Fig. 2. Mesobiotus anastasiae sp. nov. , cuticular sculpture. A . Paratype (SPbU 256(18)). Sculpture of the dorsal body surface, PhC. B–D . Paratype (SPbU_Tar16). B . High magnification of the sculpture of the dorsal body surface, SEM. C . Dorso-lateral zone of concentrated cuticular dots, SEM. D . Dot-like sculpture on the dorsal side of hind legs, SEM. Scale bars A, C, D = 5 µm; B = 2 µm. Fig. 3. Mesobiotus anastasiae sp. nov. , bucco-pharyngeal apparatus. A, E–O . Paratype, ♀ (SPbU 256(9)). B–D . Holotype, ♀ (SPbU 256(8)). A . Total dorso-ventral view of the bucco-pharyngeal apparatus, DIC. B . Ventral row of macroplacoids, focused on their outer surface, DIC. C . Ventral row of macroplacoids, focal plane shifted deeper into the parynx, black arrowheads indicate appendices of the third macroplacoid bent inward, DIC. D . Lateral rows of macroplacoids, black arrowheads indicate appendices of the third macroplacoid bent inward, DIC. E . Ventral row of macroplacoids, focused on their outer surface; note the absence of the preterminal constriction of the third macroplacoid, DIC. F–O . Oral cavity armature, dorsal (F–H, K–M) and ventral view (I–J, N–O), black arrowheads indicate teeth of the first band, F–J = PhC, K–O = DIC. Scale bars: A = 10 µm; B–O = 5 µm. Fig. 4. Mesobiotus anastasiae sp. nov. , paratype (SPbU_Tar16). Legs. A . External surface of leg III with developed dot-like sculpture, SEM. B . Internal surface of leg II without dot-like sculpture, white asterisk – pulvinus on the inner side of the leg, SEM. C . Claws of leg I, black arrowhead indicates barlike cuticular thickening (holotype), PhC. D . Leg II, black arrowhead indicates dot-like sculpture on the external surface (SPbU 256(1), paratype), PhC. E . Claws of leg II, black arrowhead indicates dotlike sculpture on the external surface of the leg, SEM. F . Claws of leg IV, black arrowhead indicates horseshoe-like structure (SPbU 256(13), paratype), PhC. G . Leg IV, dot-like sculpture on the ventral surface (holotype), PhC. H . Claws of leg IV, SEM. Scale bars = 5 µm. Fig. 5. Mesobiotus anastasiae sp. nov. , eggs. A–F . Paratype (SPbU 256(15)). G–I . Paratype (SPbU 256(4)). A . Total view of the egg surface, PhC. B . Total view of the optical section of the embryonated egg, PhC. C–D . Details of the egg surface, DIC, PhC. E . Optical section of the egg process, black arrowhead indicates pore, DIC. F–I . Optical sections of different egg processes, black arrowheads indicate collar, PhC. Scale bars: A–B = 20 µm; C–D = 10 µm; E–I = 5 µm. Fig. 6. Mesobiotus anastasiae sp. nov. , paratype (SPbU_Tar16). Eggs. A–C . Total view of the egg, SEM. D–F . Details of the egg surface, white arrowheads indicate pores above the collar, white arrows indicate pores below the collar, SEM. Scale bars: A–C = 20 µm; D–F = 5 µm. Table 3. Measurements (in μm) of selected morphological structures of eggs of Mesobiotus anastasiae sp. nov. Abbreviations: N = number of eggs/structures measured, range refers to the smallest and the largest structure among all measured specimens; SD = standard deviation.

CHARACTER	N	RANGE	MEAN	SD
Egg bare diameter	13	61.7–74.7	66.7	4.0
Egg full diameter	13	84.6–108.5	95.9	7.1
Process height	39	14.6–21.3	17.7	1.8
Process base width	39	11.8–18.6	15.5	1.6
Process base/height ratio	39	63%–117%	88%	9%
Inter-process distance	39	0.0–3.8	1.3	0.9
Number of processes on egg circumference	13	12–15	13.4	0.9

Eggs spherical, white, ornamented and laid freely ( Figs 5 A–B, 6A–C; morphometrics in Table 3 ). Chorion with conical processes that can be attributed to the “ sharp wide cones with collars ” and “ reticular design with “bubbles” ” morphotypes (according to Kaczmarek et al . 2020 ). Egg processes with wide bases and thinned and flexible apices ( Figs 5 C–I, 6D–F). Basal parts of the processes with bilayered walls, with a net of trabecular structures between the internal and external layers, forming irregular rounded meshes of different size, so the processes seem to be reticulated in LM ( Fig. 6 C–I). Apical parts of the processes with bubble-like internal structure ( Fig. 5 E–I), often branching ( Figs 5 E–F, 6E–F). The SEM observations revealed circular wrinkles on the outer surface of the processes ( Fig. 6 D–F). Basal parts of the processes with well-developed collar elevated above the egg surface ( Figs 5 G–I, black arrowheads; 6D–F). Large pores ( 1–3 µm in diameter), visible in LM and in SEM, are present on the surface of all processes, near the collar, in two rows: one row of larger pores above the collar and second row of smaller pores below the collar ( Figs 5 C–E, black arrowhead; 6D–F). Process bases are smooth, without a crown of granules or teeth. Egg surface between the processes without areolation, with a system of Fig, 7. Mesobiotus mauccii ( Pilato, 1974 ) . Paratype egg (UNICT 2132). A–B . Egg processes, white arrowhead indicates polygonate mesh-like pattern, black arrowheads indicate the collar, black arrows indicate pores, DIC. Scale bars: 10 µm . irregularly distributed ridges and small pores between them ( Fig. 6 D–F). In some eggs underdeveloped small processes are present among normal processes ( Fig. 6B, E ). Reproduction The new species is dioecious.Adult males were identified by having testis filled with spermatozoa, visible under PhC on mounted slides. Males of M. anastasiae sp. nov. exhibit no secondary sexual dimorphism. DNA sequences Sequences of good quality for the four aforementioned molecular markers were obtained from one specimen (voucher slide SpbU 256(11)): COI sequence (GenBank: MT904513 ), 658 bp long; 18S rRNA sequence (GenBank: MT903468 ), 1030 bp long; 28S rRNA sequence (GenBank: MT903612 ), 740 bp long; ITS-2 sequence (GenBank: MT903470 ), 419 bp long.