Two new species of the genus Milnesium Doyere, 1840 (Tardigrada, Apochela, Milnesiidae) from Madagascar Author Kaczmarek, Lukasz Author Grobys, Daria Author Kulpa, Adam Author Bartylak, Tomasz Author Kmita, Hanna Author Kepel, Marta Author Kepel, Andrzej Author Roszkowska, Milena text ZooKeys 2019 884 1 22 http://dx.doi.org/10.3897/zookeys.884.29469 journal article http://dx.doi.org/10.3897/zookeys.884.29469 1313-2970-884-1 725DD9DAC5FF450AABC042A06C6E5C37 5140F9C771AC5CCD9EC9247D8B103E00 Milnesium wrightae sp. nov. Figures 6-8 , 9-11 , Table 7 Material examined. Holotype and 28 paratypes, all from sample No 109: Ivohibory forest, Madagascar, moss sample from tree, coll. Marta Kepel and Andrzej Kepel. Description. Adult females ( Fig. 6 , Table 7 ) with no modified claws I. Body light yellow before fixation and transparent afterwards, eyes present only in 39% of measured specimens. Dorsal cuticle sculptured with pseudopores not arranged in bands, sparsely distributed and not forming reticular design ( Fig. 7 ). Six peribuccal papillae and six peribuccal lamellae present around the mouth opening. Two cephalic papillae positioned laterally. Peribuccal papillae slightly longer than lateral papillae. The buccal apparatus of the Milnesium type ( Figs 6 , 8 ). The buccal tube with standard width on average 62% of its length, and funnel-shaped, wider anteriorly (posterior diameter on average 91% of the anterior diameter) (Table 7 ). The pharyngeal bulb elongated, pear-shaped and without placoids or septulum. Figures 6-8. Milnesium wrightae sp. nov. 6 Habitus (ventral view) (holotype) 7 dorsal cuticle with pseudopores (holotype) 8 buccal tube (paratype). All in PCM. Table 7. Measurements and pt values of selected morphological structures of females of Milnesium wrightae sp. nov. mounted in Hoyer's medium (N - number of specimens/structures measured, RANGE refers to the smallest and the largest structure among all measured specimens; SD - standard deviation, pt - ratio of the length of a given structure to the length of the buccal tube expressed as a percentage).
Character N Range Mean SD Holotype
µm pt µm pt µm pt µm pt
Body length 17 329 - 553 - - - 448 - 60 - 515 -
Peribuccal papillae length 12 6.8 - 10.4 13.2 - 16.6 9.0 14.9 1.1 1.0 9.3 15.3
Lateral papillae length 8 5.1 - 8.4 10.0 - 13.0 6.7 11.3 1.0 0.9 6.1 10.0
Buccal tube
Length 17 44.8 - 65.6 - - - 58.4 - 6.5 - 60.9 -
Stylet support insertion point 15 31.2 - 45.8 69.3 - 73.2 40.8 70.6 4.7 1.1 43.9 72.1
Anterior width 16 14.0 - 23.0 29.5 - 37.9 19.0 32.7 2.5 2.4 20.1 33.0
Standard width 14 13.0 - 20.7 27.5 - 36.4 17.7 31.0 2.3 2.6 19.8 32.5
Posterior width 14 12.7 - 20.1 26.6 - 33.5 16.9 29.6 2.2 2.3 18.9 31.0
Standard width/length ratio 14 28% - 36% - - - 31% - 3% - 33% -
Posterior/anterior width ratio 14 88% - 97% - - - 91% - 3% - 94% -
Claw 1 lengths
External primary branch 16 11.0 - 15.2 19.5 - 24.6 13.0 21.9 1.2 1.6 13.8 22.7
External base + secondary branch 15 9.6 - 14.9 19.5 - 23.4 12.6 21.2 1.4 1.0 13.8 22.7
External spur 7 2.8 - 3.7 4.7 - 5.6 3.2 5.2 0.3 0.3 ? ?
External branches length ratio 14 87% - 103% - - - 97% - 5% - 100% -
Internal primary branch 16 10.9 - 14.0 19.4 - 24.6 12.4 20.9 0.9 1.4 12.7 20.9
Internal base + secondary branch 16 9.0 - 14.0 18.9 - 21.7 12.1 20.4 1.4 0.8 12.8 21.0
Internal spur 13 2.8 - 3.6 4.7 - 6.5 3.1 5.2 0.3 0.5 3.2 5.3
Internal branches length ratio 15 83% - 103% - - - 98% - 7% - 101% -
Claw 2 lengths
External primary branch 15 10.6 - 15.3 20.5 - 26.0 13.3 22.4 1.2 1.5 14.6 24.0
External base + secondary branch 14 9.3 - 13.7 18.8 - 21.5 12.2 20.5 1.4 0.7 12.5 20.5
External spur 8 3.1 - 4.1 4.9 - 6.7 3.4 5.5 0.3 0.6 4.1 6.7
External branches length ratio 13 78% - 103% - - - 92% - 7% - 86% -
Internal primary branch 14 10.9 - 15.0 19.2 - 24.3 12.5 21.3 1.1 1.7 13.5 22.2
Internal base + secondary branch 15 9.0 - 14.2 18.0 - 22.3 12.1 20.2 1.5 1.0 12.9 21.2
Internal spur 12 2.6 - 4.6 4.3 - 6.9 3.4 5.7 0.6 0.7 3.7 6.1
Internal branches length ratio 13 82% - 103% - - - 95% - 8% - 96% -
Claw 3 lengths
External primary branch 17 10.8 - 15.2 19.0 - 26.5 13.2 22.6 1.4 1.7 ? ?
External base + secondary branch 16 9.5 - 15.7 19.1 - 24.7 12.0 20.7 1.6 1.4 ? ?
External spur 7 3.0 - 4.0 4.9 - 6.8 3.3 5.5 0.4 0.7 ? ?
External branches length ratio 16 79% - 103% - - - 92% - 6% - ? -
Internal primary branch 17 10.7 - 14.1 19.2 - 25.0 12.4 21.3 1.1 1.7 ? ?
Internal base + secondary branch 16 9.0 - 14.1 17.8 - 21.8 11.5 19.7 1.5 1.1 ? ?
Internal spur 10 2.4 - 4.0 4.1 - 6.8 3.3 5.7 0.5 0.9 ? ?
Internal branches length ratio 16 80% - 102% - - - 93% - 7% - ? -
Claw 4 lengths
Anterior primary branch 12 12.6 - 18.4 23.7 - 28.9 15.2 25.8 1.6 1.9 15.8 25.9
Anterior base + secondary branch 12 11.2 - 17.4 22.6 - 27.5 14.7 25.0 1.8 1.6 16.5 27.1
Anterior spur 7 2.7 - 5.2 5.6 - 8.2 3.7 6.3 0.8 0.9 4.2 6.9
Anterior branches length ratio 11 85% - 104% - - - 97% - 6% - 104% -
Posterior primary branch 12 11.7 - 20.0 23.7 - 31.4 16.0 27.3 2.2 2.0 17.5 28.7
Posterior base + secondary branch 11 12.1 - 18.5 24.0 - 28.9 15.6 26.5 2.2 1.8 17.5 28.7
Posterior spur 7 2.9 - 5.2 5.3 - 8.2 3.8 6.7 0.9 1.0 4.4 7.2
Posterior branches length ratio 10 92% - 103% - - - 98% - 4% - 100% -
Claws of the Milnesium type, stout ( Figs 8 - 11 ). Primary branches on all legs with small, but distinct accessory points detaching from the branch at its greatest curvature ( Fig. 10 , empty arrowhead). Secondary branches of claws similar in length to primary branches and sometimes even longer. Secondary branches with rounded basal thickenings ( Figs 9-11 ). Secondary branches on legs I-III with three points, secondary claws of anterior and posterior claws IV with four points (claw configuration: [3-3]-[4-4]). The fourth point on secondary branches is always very small and located near the base of the claw ( Figs 10-11 , arrowheads). Single, long transverse, cuticular bars present under claws I-III ( Fig. 9 , arrow). Figures 9-11. Milnesium wrightae sp. nov. 9 Claws I (paratype), arrow indicates bar under claw 10 claws IV (holotype), empty arrowhead indicates small accessory point, filled arrowheads indicate the fourth points on secondary branches near the base of the claw 11 focus on the fourth points on secondary branches near the base of the claw IV (holotype, filled arrowheads). All in PCM. Males unknown. Eggs oval, smooth and deposited in the exuvium as in all other known Milnesium species. DNA sequences. We obtained good quality sequences for the applied molecular markers: 28S rRNA sequence (GenBank: MN191504), 638 bp long; COI sequence (GenBank: MN187057), 638 bp long; ITS-2 sequence (GenBank: MN239907), 392 bp long. Remarks. The fourth points on secondary branches of posterior claws can be barely visible or not visible at all in some positions of the specimens. Type locality. Madagascar, 22°37'04.5"S , 46°43'14.1"E , ca. 1198 m asl, Fianarantsoa Province, Ivohibory forest. Etymology. This species is named after Patricia Chapple Wright, an American primatologist and conservationist, best known for her studies on lemurs. She contributed to the establishment of the Ranomafana National Park in Madagascar. She also organized and led the expedition to the Ivohibory forest, during which several new species of tardigrades were found, including this species. Type depositories. The holotype and 23 paratypes (slides: MAD109/1, MAD109/3, MAD109/4, MAD109/5, MAD109/7) are deposited at the Department of Animal Taxonomy and Ecology, Adam Mickiewicz University in Poznan , Uniwersytetu Poznanskiego 6, Poznan , Poland, five paratypes (slides: MAD109/2) are deposited at the Institute of Zoology and Biomedical Research, Jagiellonian University, Gronostajowa 9,30-387, Krakow , Poland. Morphological differential diagnosis. The new species, by the presence of four points on secondary branches of claws IV, is most similar to Mil. quadrifidum Nederstroem , 1919, which is the only valid Milnesium species with four points on secondary branches of all claws. However, Mil. wrightae sp. nov. differs from Mil. quadrifidum not only by claw configuration ([4-4]-[4-4] in Mil. quadrifidum vs. [3-3]-[4-4] in Mil. wrightae sp. nov.), but also by the position of fourth points on secondary branches of claws IV (located near the base of the claw in the new species vs. near the top of the claw in Mil. quadrifidum ). Additionally, all secondary branch points have similar length in Mil. quadrifidum , whereas the fourth points are very clearly smaller than the others in Mil. wrightae sp. nov. Genotypic differential diagnosis. The ranges of uncorrected genetic p-distances between the Mil. wrightae sp. nov. and species of the genus Milnesium , for which molecular marker sequences are available from GenBank (see Table 6 for details), are as follows: 1. 28S rRNA: 5.7-8.0% (6.7% on average), with the most similar being Milnesium sp. from North America (JX888585.1, JX888586.1, JX888587.1) (unpublished) and the least similar being Mil. t. tardigradum from Poland (KC138808.1 and KC138809.1) (unpublished); 2. COI: 17.7-38.4% (22.0 % on average), with the most similar being Mil. variefidum from UK (KT951663.1) ( Morek et al. 2016 ) and the least similar being Mil. t. tardigradum from Spain (FJ435810.1) ( Guil and Giribet 2012 ); 3. ITS-2: 25.6-36.3% (31.5% on average), with the most similar being Mil. matheusi sp. nov. (present studies) and the least similar being Mil. cf. granulatum from USA (MK681879.1) ( Jackson and Meyer 2019 ).