Milnesium warrenwilsonensis, Bartels & Kayastha & Bartylak & Mioduchowska & Dmuchowska & Kaczmarek, 2025

Milnesium warrenwilsonensis sp. nov.

Figures 1–5 View FIGURE 1 View FIGURE 2 View FIGURE 3 View FIGURE 4 View FIGURE 5 and Tables 1–3 View TABLE 1 View TABLE 2 View TABLE 3

Description of the new species

Females (summary of measurements in Table 1 View TABLE 1 , raw data in SM.1). Body color light yellow ( Fig. 1A, C View FIGURE 1 ). Eyes present in 6 of 20 (30%) Hoyer’s medium fixed measured specimens. The dorsal cuticle smooth, but with pseudopores and clearly visible pseudoplates ( Figs 2–3 View FIGURE 2 View FIGURE 3 ). Pseudoplates arranged in nine transverse rows (CP: I:5; II:5; III:10; IV:10; V:12; VI:22; VII:12; VIII:19; IX:4), which are clearly visible in UVM and not fully in PCM: (I) a single trapezoid pseudoplate (divided into four more or less equal triangular parts) and an additional less visible (marked in gray in Fig. 2B View FIGURE 2 ) anteriorly situated transverse pseudoplate; (II) a large central rectangular pseudoplate (divided into five parts two anteriorly situated regular rectangles, two laterally situated triangle-like parts and single plate-shaped part in posterior part); (III) central rectangular pseudoplate (divided into four unequal rectangular parts, and two smaller parts laterally) and two large lateral square pseudoplates; (IV) large rectangular central pseudoplate (divided into six unequal rectangular parts), two small contiguous oval lateral pseudoplates and an additional two larger more distant oval lateral pseudoplates; (V) a central rectangular pseudoplate (divided longitudinally and transversely into four unequal parts: two rectangles in the middle and two long thin anterior and posterior parts) and two lateral roundish pseudoplates (divided into four unequal irregular ports); (VI) a large central rectangular pseudoplate with elongated anterior parts (divided into 14 larger and smaller parts of complex shapes) and an additional two lateral irregular pseudoplates (divided longitudinally into three unequal portions, smaller anteriorly and posteriorly and larger in the center), a less visible (marked in gray in Fig. 2B View FIGURE 2 ) rectangular pseudoplate (divided longitudinally into two smaller rectangles) visible in the posterior part of the central rectangular pseudoplate; (VII) a central rectangular pseudoplate (divided longitudinally into two equal rectangles) and additionally eight lateral pseudoplates (four square, two rectangular and two triangular), two less visible (marked in gray in Fig. 2B View FIGURE 2 ) transverse rectangular pseudoplates visible in anterior and posterior part of central plate; (VIII) a complex, trapezoid-like pseudoplate (divided into 14 larger and smaller parts of complex shapes and additionally five less visible (marked in gray in Fig. 2B View FIGURE 2 ) laterally and posteriorly arranged pseudoplates; (IX) four pseudoplates arranged transversally (internal trapezoid with elongated posterior part and the lateral triangular ( Figs 2–3 View FIGURE 2 View FIGURE 3 ).

Buccal apparatus of Milnesium type without placoids. Buccal tube cylindrical but slightly narrowed on the level of stylet supports ( Fig. 4A View FIGURE 4 ). Mouth opening surrounded with six short peribuccal papillae (with the ventral papilla being the smallest) and six triangular peribuccal lamellae almost equal in size well visible only in SEM ( Figs 4B–D View FIGURE 4 , empty arrow). Two lateral cephalic papillae also present ( Fig. 4C View FIGURE 4 , filled arrow).

Claws of Milnesium type. Primary claws with small accessory points visible both in PCM and SEM ( Fig. 5 View FIGURE 5 , filled arrow). All secondary claws with basal spur, i.e. with a [3-3]-[3-3] CC ( Fig. 5 View FIGURE 5 , empty arrow). Interior/anterior branches slightly larger than exterior/posterior branches. Cuticular bars under claws I– III present and well visible in PCM ( Fig. 5A View FIGURE 5 , filled unindented arrowhead).

Males (measurements in Table 2 View TABLE 2 ). Smaller than females ( Fig. 1B View FIGURE 1 ), with narrower buccal tube and with modified first pairs of claws into rigid hooks, as in all other Milnesium species. Eyes absent in Hoyer’s medium fixed specimens. Cuticle with pseudopores and pseudoplates. Cuticular bars under claws I–III present and well visible in PCM. All secondary claws equipped with branches, i.e. with a [3-3]-[3-3] CC.

Juveniles (measurements in Table 3 View TABLE 3 ). The life stage is indeterminate since we did not culture this species. Morphologically similar to adult females, but with larger dorsal pseudopores and weakly visible dorsal pseudoplates. Eyes absent in Hoyer’s medium fixed specimens. Other qualitative traits as in adult females.

Eggs. Smooth, oval, slightly yellowish; deposited in exuviae.

DNA molecular markers and genetic distances. We obtained good quality sequences for the amplified molecular markers: COI—544 bp long (the translation was successfully carried out with the invertebrate mitochondrial codon table and the 2 nd reading frame; GenBank: OR678203, OR678204); 18S rRNA—950 bp long (GenBank: OR677894, OR677895), and 28S rRNA—732–748 bp (GenBank: OR677896, OR677897). All three molecular markers of the new species were represented by single haplotypes.

The ranges of uncorrected genetic p-distances between Mil. warrenwilsonensis sp. nov. and other species of the genus Milnesium are as follows (SM.3–SM.5):

● COI: 16.1–20.6% (17.8% on average), with the most similar being Mil. variefidum Morek, Gąsiorek, Stec, Blagden & Michalczyk, 2016 (GenBank: KT951663 View Materials ), and the least similar being Mil. matheusi (GenBank: MN187056 View Materials ) and Mil. rastrum Suzuki, Sugiura, Tsujimoto & McInnes, 2023 (GenBank: LC721287 View Materials );

● 18S rRNA: 0.1–3.6% (2.0% on average), with the most similar being Mil. bohleberi Bartels, Nelson, Kaczmarek & Michalczyk, 2014 (GenBank: MW538123 View Materials ), and the least similar being Mil. pseudotardigradum Surmacz, Morek & Michalczyk, 2019 (GenBank: MW538129 View Materials );

● 28S rRNA: 1.5–6.9% (4.2% on average), with the most similar being Mil. bohleberi (GenBank: MW538202 View Materials ), and the least similar being Mil. iniquum Brotto-Guidetti, Morek & Garraffoni, 2024 (GenBank: PP062936).

Type locality. 35°36′34″N 82°26′22″W, 657 m asl: USA, North Carolina, Asheville, Warren Wilson College (WWC) Farm, greenshield lichen ( Flavoparmelia caperata ) mixed with other foliose lichens from a red maple tree ( Acer rubrum ), coll. Paul J. Bartels.

Type material. The series consists of the holotype (slide number USA ( WWC)-1/2) and 42 paratypes i.e.: 22 females, 6 males and 3 juveniles (slide numbers: USA-1/*, where the asterisk can be substituted by any of the following numbers: 1, 2, 3, 4); exoskeletons (after DNA extraction): 5 females and 7 males (slides numbers: USA ( WWC)-S/*, where the asterisk can be substituted by any of the following letters:A, B, C, D, E, F, G, H, I, J, K, L). In total, 43 specimens (30 females, 10 males, 3 juveniles) fixed in Hoyer’s medium on 16 microscope slides .

Etymology. The name of the species refers to the place where it was discovered i.e. Warren Wilson College Farm.

Type repositories. Holotype (female) and 19 paratypes (10 females and nine males, slides: 1/2, 1/4, S/A-L) are deposited at the Department of Animal Taxonomy and Ecology , Institute of Environmental Biology , Adam Mickiewicz University in Poznań , Uniwersytetu Poznańskiego 6, 61–614 Poznań , Poland. Eleven paratypes (nine females and two males, slide: 1/3) are deposited in the Institute of Systematics and Evolution of Animals , Polish Academy of Sciences , Sławkowska 17, 31-016, Kraków , Poland. Twelve paratypes (10 females and two males, slide: 1/1) are deposited in the Department of Biology , Warren Wilson College, Asheville, USA.