BARBARIA WEGLARSKAE GĄSIOREK, WILAMOWSKI,

VONČINA & MICHALCZYK, SP. NOV.

(FIGS 4, 7L, 8L, 9L, 10L, 11M, N; TABLE 5)

Zoobank registration: u r n: l s i d: z o o b a n k. org:act: F083907B-741F-48C0-A127-2FC28482C9AA

Morphometric data: w w w. t a r d i g r a d a. n e t / register/0110.htm

Ty p e m a t e r i a l: Holotype (adult female, slide AR.059.04) and two paratypes (adult females, slides AR.059.01, 05).

Type locality: 48°25′42″S, 71°44′48″W, 803 m asl: Argentina, Patagonia, Santa Cruz Province, Río Chico Department, vicinity of La Florida; lichen from on in the Andean Patagonian forest (see also Table 1) .

Etymology: A patronym honouring Professor Barbara Węglarska, 20.02.1922 – 02.10.2020, whose death left a void in the community of tardigradologists. A noun in the genitive case.

Description: Adult females (i.e. from the third instar onwards, measurements in Table 5). Orange body with large, red eyes; the entire pigment and eyes dissolve quickly after mounting in Hoyer’s medium. Body massive (Fig. 4A). Cylindrical, Echiniscus - type cephalic papillae (secondary clavae) and (primary) clavae; cirri embedded in bulbous cirrophores. Cirrus A is long (> 50% of the body length) and with evident, conical cirrophore (Fig. 4A, B).

Dorsal plate sculpturing of the bigranulata - type, composed of pillars present in all plate portions (pseudogranulation) and pores and pseudopores present in different elements of armour (Figs 4A, B, 7L, 8L, 9L, 10L). Pseudopores can be present or absent exclusively in the anterior portion of the median plate 2 and paired segmental plates and the entirety of median plate 3 (Fig. 4A, B). Minute pores of equal size are regularly distributed in the remaining plate portions (Figs 4A, B, 7L, 8L, 9L, 10L). The cephalic plate large, with a pronounced chalice-shaped anterior incision and lateral sutures demarcating roughly triangular lateralmost portions of the plate (Fig. 4A, B). Thin cervical plate with developed pillars and pseudopores. Lateral sutures in the scapular plate demarcate lateralmost, rectangular portions with identical sculpturing as on the rest of the plate (Fig. 4A, B). Median plates 1 and 3 unipartite (the latter strongly reduced and partially covered by the caudal plate), median plate 2 bipartite (Figs 4A, B, 8L). Paired segmental plates with broad, transverse, unsculptured bands (Figs 4A, B, 9L). The caudal plate with short, poorly sclerotized incisions clearly joined by a transversal suture (Figs 4A, B, 10L). Ventral cuticle with minute endocuticular pillars distributed evenly throughout the entire venter; a pair of small, subcephalic plates present (Fig. 4C–D). Sexpartite gonopore (Fig. 4E) placed anteriorly to a trilobed anus between legs IV.

Pedal plates I–IV with evident pillars and pseudopores (Figs 4A, 11M, N). Evident pulvini on outer sides of all legs. Dentate collar IV with numerous irregular short teeth (Fig. 11N). A tiny spine I and a tubby papilla IV (Fig. 11N). Claws slightly heteronych, because primary spurs on internal claws IV are positioned higher than those on claws I–III (Fig. 11M, N). The shape and angle at which spurs diverge from branches are almost identical on all limbs. Cuticular bars below claw bases on the inner side of legs present. Buccal apparatus with a rigid tube and round pharynx containing placoids. Lacking stylet supports.

Adult males, juveniles, larvae or eggs not found.

Differential diagnosis: There are only two other species of Barbaria with a cirrus A /body length ratio> 50% [the titles and values in the last five rows of table 1 in Michalczyk & Kaczmarek (2007) are mismatched in the case of sexes treated separately, but the ratio statistics for all measured specimens of B. ganczareki stands valid: min = 15%, max = 23%, mean = 19%]: B. jenningsi and B. ranzii, but B. weglarskae can be distinguished from:

• Barbaria jenningsi, by the type of perforation in the dorsal plates [dominant pores in B. weglarskae (Fig. 7L) vs. pseudopores in B. jenningsi (Figs 7E, F, 8F, 9F, 10F)] and claw isomorphy [anisonych/ slightly heteronych in B. weglarskae (Fig. 11M, N) vs. strongly heteronych in B. jenningsi (Fig. 11E, F)].

• Barbaria ranzii, by the pedal plate sculpturing [with evident pillars in B. weglarskae (Fig. 11M, N) vs. without pillars in B. ranzii (Fig. 11R)], the shape of papilla IV [tubby in B. weglarskae (Fig. 11N) vs. elongated in B. ranzii] and by the presence of secondary spurs directed upwards on external claws IV [absent in B. weglarskae (Fig. 11N) vs. present in B. ranzii (Fig. 11R)].

MOLECULAR PHYLOGENY

Phylogeny based on the five concatenated markers brought fully resolved relationships between all eight analysed species of Barbaria, which form clades characterized by low intraspecific and large interspecific genetic variability (Fig. 5). The topology is as follows: B. madonnae is a sisterspecies to all other sequenced Barbaria spp., which are clustered in two clades: the first comprising ( B. paucigranulata ( B. danieli + B. charrua)) and the second grouping (( B. weglarskae + B. jenningsi) + ( B. ollantaytamboensis + B. bigranulata)).

INTRA- AND INTERSPECIFIC GENETIC VARIABILITY

Regarding COI sequences deposited in GenBank, the data are available only for B. bigranulata and B. jenningsi . COI p -distances between populations of B. bigranulata and the previously published data for a population from Chile (HM193406; Jørgensen et al., 2011) ranged between 2.6 and 2.9% (alignment length = 585 bp). Analogous index for the pair B. weglarskae – B. jenningsi (KP013596; Velasco-Castrillón et al., 2015) was 18.9% (alignment length = 472 bp).

More than one haplotype per marker has been found for all markers, but only in a few species. The intraspecific p -distances are as follows: 18S rRNA: 0.1% (in B. bigranulata and B. charrua); 28S rRNA: 0.1–0.4% ( B. bigranulata, B. ollantaytamboensis); ITS 1: 0.5 % ( B. bigranulata), 0.1 – 1.0 % ( B. charrua); ITS2: 0.2% ( B. bigranulata), 0.2–0.4% ( B.ollantaytamboensis); COI: 0.1–2.3% ( B. bigranulata) and 0.7% ( B. charrua).

Interspecific p -distances in the analysed dataset are as follows:

• 18S rRNA: 0.0–2.5% (1.2% on average), with the most similar being B. charrua (MZ820796) and B. danieli (MZ820800); and the least similar being B. madonnae (MZ820803) and B. ollantaytamboensis (MZ820804).

• 28S rRNA: 0.0–4.3% (2.4% on average), with the most similar being B. charrua (MZ820814) and B. danieli (MZ820818); and the least similar being B. madonnae (MZ820821) and B. ollantaytamboensis (MZ820823).

• ITS1: 0.3–10.3% (5.5% on average), with the most similar being B. charrua (MZ820833) and B. danieli (MZ820836); and the least similar being B. madonnae (MZ820839) and B. bigranulata (MZ820828).

• ITS2: 1.6–10.8% (8.1% on average), with the most similar being B. charrua (MZ822380) and B. danieli (MZ822384); and the least similar being B. danieli + B. madonnae (MZ822384, MZ822387) and B. ollantaytamboensis (MZ822388 –91).

• COI: 9.1–20.4% (16.0% on average), with the most similar being B. charrua (MZ820850) and B. danieli (MZ820853); and the least similar being B. ollantaytamboensis (MZ820855) and B. weglarskae (MZ820860).

MORPHOLOGICAL EVOLUTION

Mapping morphological traits on to the phylogeny suggests that the ancestor of Barbaria was most probably covered with uniform dorsal sculpturing comprising both pillars and pores (Fig. 6A) and welldelimited pedal plates on legs I–III with densely packed pillars and pseudopores (Fig. 6B). Its papillae on legs IV were elongated (Fig. 6C), meaning that they were much longer than wide. Internal claws were exhibiting heteronychy or lacking primary spurs (Fig. 6D). In other words, among the extant species, B. bigranulata is morphologically overall the most similar to the last common ancestor of the analysed species.