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).CharacterNRangeMeanSDHolotypeµmptµmptµmptµmptBody length17329-553---448-60-515-Peribuccal papillae length126.8-10.413.2-16.69.014.91.11.09.315.3Lateral papillae length85.1-8.410.0-13.06.711.31.00.96.110.0Buccal tubeLength1744.8-65.6---58.4-6.5-60.9-Stylet support insertion point1531.2-45.869.3-73.240.870.64.71.143.972.1Anterior width1614.0-23.029.5-37.919.032.72.52.420.133.0Standard width1413.0-20.727.5-36.417.731.02.32.619.832.5Posterior width1412.7-20.126.6-33.516.929.62.22.318.931.0Standard width/length ratio1428%-36%---31%-3%-33%-Posterior/anterior width ratio1488%-97%---91%-3%-94%-Claw 1 lengthsExternal primary branch1611.0-15.219.5-24.613.021.91.21.613.822.7External base + secondary branch159.6-14.919.5-23.412.621.21.41.013.822.7External spur72.8-3.74.7-5.63.25.20.30.3??External branches length ratio1487%-103%---97%-5%-100%-Internal primary branch1610.9-14.019.4-24.612.420.90.91.412.720.9Internal base + secondary branch169.0-14.018.9-21.712.120.41.40.812.821.0Internal spur132.8-3.64.7-6.53.15.20.30.53.25.3Internal branches length ratio1583%-103%---98%-7%-101%-Claw 2 lengthsExternal primary branch1510.6-15.320.5-26.013.322.41.21.514.624.0External base + secondary branch149.3-13.718.8-21.512.220.51.40.712.520.5External spur83.1-4.14.9-6.73.45.50.30.64.16.7External branches length ratio1378%-103%---92%-7%-86%-Internal primary branch1410.9-15.019.2-24.312.521.31.11.713.522.2Internal base + secondary branch159.0-14.218.0-22.312.120.21.51.012.921.2Internal spur122.6-4.64.3-6.93.45.70.60.73.76.1Internal branches length ratio1382%-103%---95%-8%-96%-Claw 3 lengthsExternal primary branch1710.8-15.219.0-26.513.222.61.41.7??External base + secondary branch169.5-15.719.1-24.712.020.71.61.4??External spur73.0-4.04.9-6.83.35.50.40.7??External branches length ratio1679%-103%---92%-6%-?-Internal primary branch1710.7-14.119.2-25.012.421.31.11.7??Internal base + secondary branch169.0-14.117.8-21.811.519.71.51.1??Internal spur102.4-4.04.1-6.83.35.70.50.9??Internal branches length ratio1680%-102%---93%-7%-?-Claw 4 lengthsAnterior primary branch1212.6-18.423.7-28.915.225.81.61.915.825.9Anterior base + secondary branch1211.2-17.422.6-27.514.725.01.81.616.527.1Anterior spur72.7-5.25.6-8.23.76.30.80.94.26.9Anterior branches length ratio1185%-104%---97%-6%-104%-Posterior primary branch1211.7-20.023.7-31.416.027.32.22.017.528.7Posterior base + secondary branch1112.1-18.524.0-28.915.626.52.21.817.528.7Posterior spur72.9-5.25.3-8.23.86.70.91.04.47.2Posterior branches length ratio1092%-103%---98%-4%-100%-

Table 6. Sequences of 28S rRNA, COI and ITS-2 of Milnesium taxa available in GenBank and used in differential diagnosis.DNA markerTaxonAccession numberSource28S rRNAMilnesium sp.JX888585.1Adams et. al. unpublishedJX888586.1Adams et. al. unpublishedJX888587.1Adams et. al. unpublishedMilnesium tardigradumJX888541.1Adams et. al. unpublishedJX888540.1Adams et. al. unpublishedKC138808.1Zawierucha unpublishedKC138809.1Zawierucha unpublishedMilnesium sp.AY210826.1Mallatt et. al. 2004Milnesium tardigradumFJ435780.1Guil and Giribet 2012FJ435779.1Guil and Giribet 2012Milnesium berladnicorumKT951661.1Morek et. al. 2016Milnesium variefidumKT951665.1Morek et. al. 2016COIMilnesium sp.KX306950.1Fox et al. unpublishedMilnesium tardigradumEU244603.1Schill unpublishedEU244604Schill unpublishedFJ435810.1Guil and Giribet 2012Milnesium t. tardigradumJN664950.1Michalczyk et al. 2012Milnesium cf. tardigradumJX683824.1Vicente et al. 2013JX683823.1Vicente et al. 2013JX683822.1Vicente et al. 2013Milnesium sp.KJ857002.1Velasco-Castrillón et al. 2015KJ857001.1Velasco-Castrillón et al. 2015Milnesium cf. alpigenumKU513422.1Kosztyła et al. 2016Milnesium variefidumKT951663.1Morek et al. 2016Milnesium berladnicorumKT951659.1Morek et al. 2016Milnesium sp.EF632553.1Sands et. al unpublishedMilnesium cf. granulatumMH751517.1Jackson and Meyer 2019Milnesium lagniappeMH751518.1Jackson and Meyer 2019Milnesium tardigradumMG923558.1Morek et al. 2019MG923559.1Morek et al. 2019MG923560.1Morek et al. 2019MG923561.1Morek et al. 2019MG923562.1Morek et al. 2019MG923563.1Morek et al. 2019MG923564.1Morek et al. 2019MG923565.1Morek et al. 2019Milnesium dornensisMG923566.1Morek et al. 2019ITS-2Milnesium alpigenumMH000382.1Morek et al. unpublishedMilnesium sp.MH000386.1Morek et al. unpublishedMH000387.1Morek et al. unpublishedMilnesium tardigradumHM150648.2Wełnicz et. al. 2010GQ403682.1Schill et al. 2010GQ403681.1Schill et al. 2010Milnesium t. tardigradumJF951049Michalczyk et al. 2012Milnesium variefidumKT951667.1Morek et. al. 2016KT951666.1Morek et. al. 2016Milnesium berladnicorumKT951662.1Morek et. al. 2016Milnesium cf. granulatumMK681875.1Jackson and Meyer 2019MK681876.1Jackson and Meyer 2019MK681877.1Jackson and Meyer 2019MK681878.1Jackson and Meyer 2019MK681879.1Jackson and Meyer 2019MK681880.1Jackson and Meyer 2019MK681881.1Jackson and Meyer 2019MK681882.1Jackson and Meyer 2019MK681883.1Jackson and Meyer 2019MK681884.1Jackson and Meyer 2019MK681885.1Jackson and Meyer 2019MK681886.1Jackson and Meyer 2019

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).CharacterNRangeMeanSDHolotypeµmptµmptµmptµmptBody length17329-553---448-60-515-Peribuccal papillae length126.8-10.413.2-16.69.014.91.11.09.315.3Lateral papillae length85.1-8.410.0-13.06.711.31.00.96.110.0Buccal tubeLength1744.8-65.6---58.4-6.5-60.9-Stylet support insertion point1531.2-45.869.3-73.240.870.64.71.143.972.1Anterior width1614.0-23.029.5-37.919.032.72.52.420.133.0Standard width1413.0-20.727.5-36.417.731.02.32.619.832.5Posterior width1412.7-20.126.6-33.516.929.62.22.318.931.0Standard width/length ratio1428%-36%---31%-3%-33%-Posterior/anterior width ratio1488%-97%---91%-3%-94%-Claw 1 lengthsExternal primary branch1611.0-15.219.5-24.613.021.91.21.613.822.7External base + secondary branch159.6-14.919.5-23.412.621.21.41.013.822.7External spur72.8-3.74.7-5.63.25.20.30.3??External branches length ratio1487%-103%---97%-5%-100%-Internal primary branch1610.9-14.019.4-24.612.420.90.91.412.720.9Internal base + secondary branch169.0-14.018.9-21.712.120.41.40.812.821.0Internal spur132.8-3.64.7-6.53.15.20.30.53.25.3Internal branches length ratio1583%-103%---98%-7%-101%-Claw 2 lengthsExternal primary branch1510.6-15.320.5-26.013.322.41.21.514.624.0External base + secondary branch149.3-13.718.8-21.512.220.51.40.712.520.5External spur83.1-4.14.9-6.73.45.50.30.64.16.7External branches length ratio1378%-103%---92%-7%-86%-Internal primary branch1410.9-15.019.2-24.312.521.31.11.713.522.2Internal base + secondary branch159.0-14.218.0-22.312.120.21.51.012.921.2Internal spur122.6-4.64.3-6.93.45.70.60.73.76.1Internal branches length ratio1382%-103%---95%-8%-96%-Claw 3 lengthsExternal primary branch1710.8-15.219.0-26.513.222.61.41.7??External base + secondary branch169.5-15.719.1-24.712.020.71.61.4??External spur73.0-4.04.9-6.83.35.50.40.7??External branches length ratio1679%-103%---92%-6%-?-Internal primary branch1710.7-14.119.2-25.012.421.31.11.7??Internal base + secondary branch169.0-14.117.8-21.811.519.71.51.1??Internal spur102.4-4.04.1-6.83.35.70.50.9??Internal branches length ratio1680%-102%---93%-7%-?-Claw 4 lengthsAnterior primary branch1212.6-18.423.7-28.915.225.81.61.915.825.9Anterior base + secondary branch1211.2-17.422.6-27.514.725.01.81.616.527.1Anterior spur72.7-5.25.6-8.23.76.30.80.94.26.9Anterior branches length ratio1185%-104%---97%-6%-104%-Posterior primary branch1211.7-20.023.7-31.416.027.32.22.017.528.7Posterior base + secondary branch1112.1-18.524.0-28.915.626.52.21.817.528.7Posterior spur72.9-5.25.3-8.23.86.70.91.04.47.2Posterior branches length ratio1092%-103%---98%-4%-100%-

Table 1. Primers used for amplification and sequencing of DNA fragments.DNA fragmentDirectionCodeSequence (5 ’-3’)ReferenceCOIForvardbcdF01CATTTTCHACTAAYCATAARGATATTGGDabert et al. 2010ReversebcdR04TATAAACYTCDGGATGNCCAAAAAADabert et al. 2008ITS-2ForvardITS2_Eutar_FfCGTAACGTGAATTGCAGGACStec et al. 2018ReverseITS2_Eutar_RrTGATATGCTTAAGTTCAGCGG28S rRNAForvard28SF0001ACCCVCYNAATTTAAGCATATMironov et al. 2012Reverse28SR0990CCTTGGTCCGTGTTTCAAGAC

Table 1. Primers used for amplification and sequencing of DNA fragments.DNA fragmentDirectionCodeSequence (5 ’-3’)ReferenceCOIForvardbcdF01CATTTTCHACTAAYCATAARGATATTGGDabert et al. 2010ReversebcdR04TATAAACYTCDGGATGNCCAAAAAADabert et al. 2008ITS-2ForvardITS2_Eutar_FfCGTAACGTGAATTGCAGGACStec et al. 2018ReverseITS2_Eutar_RrTGATATGCTTAAGTTCAGCGG28S rRNAForvard28SF0001ACCCVCYNAATTTAAGCATATMironov et al. 2012Reverse28SR0990CCTTGGTCCGTGTTTCAAGAC

Table 2. PCR cocktails used for the amplification of DNA fragments.ComponentConcentrationAdditional noteH2O-sterile MQbuffer1 ×5X Phusion HF Buffer; Thermo ScientificdNTPs200 µMdNTP Mix; Thermo Scientificforward primer0.5 µM-reverse primer0.5 µM-polymerase0.02 U/µlPhusion High-Fidelity DNA Polymerase; Thermo ScientificDNA--

Table 2. PCR cocktails used for the amplification of DNA fragments.ComponentConcentrationAdditional noteH2O-sterile MQbuffer1 ×5X Phusion HF Buffer; Thermo ScientificdNTPs200 µMdNTP Mix; Thermo Scientificforward primer0.5 µM-reverse primer0.5 µM-polymerase0.02 U/µlPhusion High-Fidelity DNA Polymerase; Thermo ScientificDNA--

Table 3. PCR programmes used for the amplification of COI, ITS-2 and 28S rRNA.StepCOIITS-2 and 28S rRNACyclesTime [min.:sec.]Temp. [°C]CyclesTime [min:sec]Temp. [°C]initial denaturation-05:0098-05:0098denaturation500:3098---annealing00:3045---extension01:0072---denaturation3000:30983500:3098annealing00:305000:3050extension01:007201:0072final extension-07:0072-07:0072

Table 4. Measurements and pt values of selected morphological structures of adult females of Milnesium matheusi 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).CharacterNRangeMeanSDHolotypeµmptµmptµmptµmptBody length6630-766---691-45-766-Peribuccal papillae length510.0-12.018.6-22.111.019.90.81.511.818.9Lateral papillae length79.4-10.716.5-19.710.018.10.41.210.316.5Buccal tubeLength951.3-62.5---56.6-3.8-62.5-Stylet support insertion point934.5-42.366.1-69.438.467.82.41.341.566.4Anterior width925.2-35.947.6-57.928.951.03.23.131.450.2Standard width923.1-31.142.4-50.826.346.52.73.029.447.0Posterior width923.0-30.241.1-50.325.745.32.63.128.946.2Standard width/length ratio942%-51%---46%-3%-47%-Posterior/anterior width ratio984%-94%---89%-4%-92%-Claw 1 lengthsExternal primary branch917.2-21.830.2-35.218.933.31.51.621.834.9External base + secondary branch913.3-16.723.5-27.915.026.51.21.516.626.6External spur73.5-5.36.4-9.64.47.80.71.3??External branches length ratio976%-82%---80%-2%-76%-Internal primary branch916.0-21.130.2-34.518.332.31.61.621.133.8Internal base + secondary branch913.3-16.624.5-27.314.826.21.11.016.326.1Internal spur93.3-5.56.1-10.54.47.70.81.45.58.8Internal branches length ratio977%-88%---81%-4%-77%-Claw 2 lengthsExternal primary branch817.4-21.232.9-36.519.534.91.41.421.233.9External base + secondary branch713.7-17.024.5-27.515.026.71.11.217.027.2External spur33.9-4.97.2-7.84.47.60.50.44.97.8External branches length ratio772%-81%---77%-3%-80%-Internal primary branch816.8-20.531.1-35.718.733.31.31.520.232.3Internal base + secondary branch913.0-16.325.0-27.914.726.01.10.916.326.1Internal spur93.4-5.86.1-10.34.47.80.81.54.77.5Internal branches length ratio874%-81%---78%-3%-81%-Claw 3 lengthsExternal primary branch519.7-21.032.3-38.320.535.70.62.5??External base + secondary branch614.2-16.324.5-28.415.427.10.71.5??External spur53.5-5.26.4-9.34.47.70.71.1??External branches length ratio572%-82%---75%-4%-?-Internal primary branch518.9-20.431.3-36.519.734.30.62.0??Internal base + secondary branch613.7-16.023.7-28.214.926.20.81.9??Internal spur53.8-5.67.0-9.74.88.30.71.1??Internal branches length ratio570%-79%---75%-4%-?-Claw 4 lengthsAnterior primary branch719.6-23.035.1-39.820.937.21.31.523.036.8Anterior base + secondary branch714.6-17.226.3-29.415.828.20.91.117.227.5Anterior spur64.1-6.37.5-11.55.49.70.91.66.09.6Anterior branches length ratio771%-80%---76%-4%-75%-Posterior primary branch720.5-24.038.1-41.321.838.91.11.124.038.4Posterior base + secondary branch715.2-17.726.9-29.616.128.60.81.017.728.3Posterior spur74.4-5.87.6-10.35.29.30.61.15.58.8Posterior branches length ratio770%-76%---74%-2%-74%-

Table 5. Measurements and pt values of selected morphological structures of adult males (with modified claws I) of Milnesium matheusi 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).CharacterNRangeMeanSDµmptµmptµmptBody length2409-428---419-13-Peribuccal papillae length33.0-3.98.9-11.33.510.20.51.2Lateral papillae length35.6-6.016.2-17.85.917.10.20.8Buccal tubeLength333.8-34.5---34.2-0.4-Stylet support insertion point221.2-22.362.7-64.621.863.70.81.4Anterior width39.4-11.227.8-32.610.530.81.02.6Standard width39.1-9.826.9-28.59.527.80.40.8Posterior width39.4-10.227.8-29.69.828.70.40.9Standard width/length ratio327%-28%---28%-1%-Posterior/anterior width ratio388%-100%---94%-6%-Claw 1 lengthsExternal primary branch215.8-16.345.9-48.216.147.10.41.6External base + secondary branch314.1-15.041.7-43.514.642.70.50.9External spur23.2-3.49.3-9.93.39.60.10.4External branches length ratio287%-94%---90%-5%-Internal primary branch314.9-15.743.2-46.415.445.10.51.7Internal base + secondary branch314.0-14.540.6-42.914.241.40.31.3Internal spur33.0-3.78.9-10.73.49.90.40.9Internal branches length ratio389%-94%---92%-2%-Claw 2 lengthsExternal primary branch216.9-17.949.0-53.017.451.00.72.8External base + secondary branch113.2-13.239.1-39.113.239.1??External spur13.5-3.510.4-10.43.510.4??External branches length ratio174%-74%---74%-?-Internal primary branch316.4-16.947.5-50.016.748.80.31.2Internal base + secondary branch212.7-12.837.2-37.612.837.40.10.3Internal spur23.5-5.010.4-14.54.312.41.13.0Internal branches length ratio275%-76%---76%-1%-Claw 3 lengthsExternal primary branch316.2-17.447.1-51.516.849.00.62.3External base + secondary branch212.1-12.835.2-37.912.536.50.51.9External spur13.9-3.911.3-11.33.911.3??External branches length ratio274%-75%---74%-1%-Internal primary branch314.8-17.043.0-50.316.046.71.13.6Internal base + secondary branch212.7-13.037.6-37.812.937.70.20.2Internal spur22.9-4.08.4-11.83.510.10.82.4Internal branches length ratio275%-88%---81%-9%-Claw 4 lengthsAnterior primary branch316.3-17.047.4-49.316.648.50.41.0Anterior base + secondary branch212.4-12.936.7-37.512.737.10.40.6Anterior spur13.8-3.811.0-11.03.811.0??Anterior branches length ratio275%-79%---77%-3%-Posterior primary branch317.7-18.851.3-54.718.353.40.61.8Posterior base + secondary branch312.7-13.737.1-39.813.138.20.61.5Posterior spur23.0-4.18.7-11.93.610.30.82.3Posterior branches length ratio369%-73%---72%-2%-