Planktonic Protistan Communities in Lakes Along a Large-Scale Environmental Gradient Maryia Khomich, Håvard Kauserud, Ramiro Logares, Serena Rasconi, Tom Andersen

Planktonic protistan communities in lakes along a large-scale environmental gradient Maryia Khomich, Håvard Kauserud, Ramiro Logares, Serena Rasconi, Tom Andersen

Supplementary Information

Fig. S1. Scatterplot matrix of spatial variables (latitude, longitude, altitude) against distance- weighted regionally averaged TP (TPreg) within a 100, 250, 350, and 500 km radius of each lake, calculated using the same data and the same algorithm as in Ptacnik et al. (2010) . All Pearson correlations between longitude and TPreg are between 0.93 and 0.97.

Fig. S2. Scatterplot matrix of spatial (longitude, latitude and altitude) and environmental (TOC, TP and conductivity, all log transformed) predictors.

Fig. S3. NMDS ordination of Hellinger transformed relative abundance data for 10 samples that were sequenced twice. Each replicate is connected by a line with a respective sample (dark blue is actual sample; light blue is its replicate).

Fig. S4. NMDS ordination plot of protistan functional groups. Each point represents an OTU, and point color and shape reflect a functional group (autotroph, heterotroph, parasite and unclassified protists). Arrows represent spatial (longitude, latitude and altitude) and environmental (TOC, TP and conductivity, all log transformed) predictors. Stress = 0.195. Putative autotrophs: Bacillariophyceae, Charophyta, Chlorophyta, Chrysophyceae, Cryptophyta, Dictyochophyceae, Dinoflagellata, Haptophyta, Raphidophyceae, Synurophyceae. Putative heterotrophs: Bicosoecida, Centroheliozoa, Choanozoa, Ciliophora, Fungi. Putative parasites: Cercozoa, Chytri diomycota, Cryptomycota, Ichthyosporea, Perkinsea. Group ‘other ’ included Amoebozoa, Excavata, Katablepharida and Oomycota.

Fig. S5. Scatterplot matrix of alpha diversity measures (rarefied OTU richness, Shannon H and Simpson D diversity) and NMDS site scores (k=2). Fig. S6. Scatterplot matrix of climatic variables (annual mean temperature (Tmean), maximum temperature (Tmax), minimum temperature (Tmin) and precipitation (Prec)) from the Worldclim data set and spatial predictors (latitude, longitude, altitude).

Table S1. Location and environmental characteristics of sampling sites.

Table S2. Relative abundance of different eukaryotic supergroup/group sequences and OTUs and occurrence of groups in n lakes of the whole set of Scandinavian lakes ( n=74).

Table S3. Summary of diversity indices.

Table S4. Kendall's Tau correlation tests between DCA and NMDS axes for NMDS ordination run in two (k=2) and three (k=3) dimensions. Significant correlation between axes (p < 0.05) is indicated by bold font.

Table S5. Permutation-based significance probabilities (p.perm) between NMDS axes scores and rarefied richnesses (S.rar) of the most abundant taxonomic groups.

Table S6. Results of the RDA variation partitioning for protistan community data. Fig. S1. Scatterplot matrix of spatial variables (latitude, longitude, altitude) against distance- weighted regionally averaged TP (TPreg) within a 100, 250, 350, and 500 km radius of each lake, calculated using the same data and the same algorithm as in Ptacnik et al. (2010). All Pearson correlations between longitude and TPreg are between 0.93 and 0.97. Fig. S2. Scatterplot matrix of spatial (longitude, latitude and altitude) and environmental (TOC, TP and conductivity, all log transformed) predictors. Fig. S3. NMDS ordination of Hellinger transformed relative abundance data for 10 samples that were sequenced twice. Each replicate is connected by a line with a respective sample (dark blue is actual sample; light blue is its replicate). Fig. S4. NMDS ordination plot of protistan functional groups. Each point represents an OTU, and point color and shape reflect a functional group (autotroph, heterotroph, parasite and unclassified protists). Arrows represent spatial (longitude, latitude and altitude) and environmental (TOC, TP and conductivity, all log transformed) predictors. Stress = 0.195. Putative autotrophs: Bacillariophyceae, Charophyta, Chlorophyta, Chrysophyceae, Cryptophyta, Dictyochophyceae, Dinoflagellata, Haptophyta, Raphidophyceae, Synurophyceae. Putative heterotrophs: Bicosoecida, Centroheliozoa, Choanozoa, Ciliophora, Fungi. Putative parasites: Cercozoa, Chytridiomycota, Cryptomycota, Ichthyosporea, Perkinsea. Group ‘other ’ included Amoebozoa, Excavata, Katablepharida and Oomycota. Fig. S5. Scatterplot matrix of alpha diversity measures (rarefied OTU richness, Shannon H and Simpson D diversity) and NMDS site scores (k=2). Fig. S6. Scatterplot matrix of climatic variables (annual mean temperature (Tmean), maximum temperature (Tmax), minimum temperature (Tmin) and precipitation (Prec)) from the Worldclim data set and spatial predictors (latitude, longitude, altitude). Table S1. Location and environmental characteristics of sampling sites.

SampleID Lake ID Lake Country Date Time Latitude Longitude Altitude TOC TP Cond Area.km 2 170 170 Gjersjøen N 08.08.2011 14:00 59,7897 10,7749 40 6,898 9,80 22,6 2,64 180 180 Øgderen N 29.07.2011 16:38 59,7139 11,4130 133 6,944 19,20 5,82 12,66 189 189 Krøderen N 28.07.2011 16:20 60,1349 9,7586 133 4,395 3,55 1,98 43,91 191 191 Rødbyvatnet N 09.08.2011 15:00 59,5818 10,4872 118 6,652 9,35 8,48 1,16 194 194 Sperillen N 28.07.2011 15:15 60,5040 10,0584 150 3,500 4,00 2,08 37,28 214 214 Gjesåssjøen N 30.07.2011 11:26 60,6817 11,9924 176 9,613 15,10 4,52 3,98 233 233 Osensjøen N 27.07.2011 16:50 61,1758 11,8877 438 7,689 3,05 1,7 43,64 236 236 Rokossjøen N 30.07.2011 13:10 60,7869 11,4407 215 11,810 8,25 2,8 3,77 242 242 Sør Mesna N 28.07.2011 09:45 61,0764 10,8000 522 7,514 6,70 2,25 6,93 252 252 Vermundsjøen N 30.07.2011 15:32 60,6951 12,3870 215 11,945 9,25 1,98 3,34 258 258 Gjønvatnet N 22.07.2011 10:50 60,2697 5,8410 40 1,300 2,00 1,67 2,89 261 261 Kalandsvatnet N 22.07.2011 14:30 60,2709 5,4022 53 2,854 3,80 5,64 3,37 264 264 Myrkdalsvatnet N 22.07.2011 13:00 60,8117 6,4709 229 0,954 1,00 0,67 1,56 277 277 Engsetdalsvatnet N 20.07.2011 11:49 62,5333 6,6328 43 2,656 1,15 3,05 4,40 285 285 Rotevatnet N 23.07.2011 13:40 62,1405 6,1185 47 3,427 2,55 2,33 1,40 288 288 Vatnevatnet N 20.07.2011 16:56 62,1531 6,2286 90 1,314 4,70 1,98 2,05 326 326 Einavatnet N 10.08.2011 10:00 60,5381 10,6530 398 5,572 3,00 8,7 13,74 328 328 Randsfjorden N 28.07.2011 11:55 60,7230 10,2677 135 4,514 2,65 3,55 140,09 339 339 Ringsjøen N 28.07.2011 10:55 60,8824 10,3547 378 8,022 5,55 6,03 1,22 340 340 Sæbufjorden N 16.08.2011 12:40 61,0126 9,2151 379 3,659 3,10 3,01 1,53 344 344 Strondafjorden N 16.08.2011 14:15 60,9775 9,2486 355 2,833 2,00 2,9 13,36 345 345 Trevatna N 28.07.2011 14:05 60,6388 10,4353 384 9,680 4,30 2,06 4,80 349 349 Bogstadvannet N 11.08.2011 10:25 59,9705 10,6185 145 7,026 6,60 3,31 1,09 SampleID Lake ID Lake Country Date Time Latitude Longitude Altitude TOC TP Cond Area.km 2 353 353 Aspern N 29.07.2011 11:53 59,1471 11,6844 106 8,539 9,95 5,56 6,59 361 361 Rødenessjøen N 29.07.2011 14:20 59,5320 11,6260 118 9,115 12,20 5,49 15,95 363 363 Rømsjøen N 29.07.2011 15:32 59,7011 11,8438 138 6,695 1,20 3,18 13,66 374 374 Edlandsvatnet N 21.07.2011 11:08 58,7626 5,8705 104 2,631 3,85 5,28 2,11 378 378 Hetlandsvatn N 21.07.2011 15:23 59,1752 6,1092 48 2,370 2,35 6,99 2,06 380 380 Lutsivatn N 21.07.2011 12:45 58,8601 5,8478 25 3,622 6,00 8,1 2,83 394 394 Vatsvatnet N 21.07.2011 17:18 59,5267 5,7042 15 4,092 18,90 4,68 2,17 395 395 Vostervatnet N 21.07.2011 14:02 59,0958 5,9754 54 3,122 3,60 5,87 2,66 404 404 Jølstravatnet N 23.07.2011 16:45 61,5579 6,4005 207 0,665 0,50 1,41 39,25 405 405 Oppstrynvatnet N 23.07.2011 11:55 61,9146 7,0767 29 0,344 1,70 1,39 22,97 433 433 Bandak N 27.07.2011 09:52 59,4016 8,2300 72 2,588 1,20 1,53 26,67 436 436 Grungevatnet N 27.07.2011 12:49 59,7071 7,7592 540 2,542 1,55 1,44 1,62 453 453 Vinjevatn N 27.07.2011 11:30 59,6123 7,8522 466 3,490 1,05 1,31 3,23 481 481 Åsrumvatnet N 26.07.2011 12:30 59,1633 10,0578 7 7,111 20,20 7,4 1,14 482 482 Bergsvannet N 09.08.2011 14:00 59,5884 10,0527 36 7,070 17,85 9,55 3,01 486 486 Goksjø N 26.07.2011 14:05 59,1725 10,1650 28 6,576 27,45 9,91 3,40 487 487 Hallevatnet N 26.07.2011 10:47 59,0253 9,9092 48 5,013 2,00 7,12 3,66 498 498 Dagarn S 05.08.2011 19:30 59,9037 15,7029 130 6,217 3,00 5,4 1,67 519 519 Langen S 02.08.2011 13:05 59,5911 18,5215 28 6,392 4,90 12,2 1,38 2252 2252 Rotnessjøen N 30.07.2011 09:59 60,4969 12,3412 260 11,175 4,55 2,23 1,09 2268 2268 Mylla N 10.08.2011 12:15 60,2443 10,5911 496 4,114 2,45 5,56 1,69 2312 2312 Femsjøen N 29.07.2011 10:55 59,1332 11,4707 79 8,420 7,45 5,32 10,66 2374 2374 Klämmingen S 02.08.2011 11:33 59,0920 17,2983 9 8,832 11,45 10,7 9,99 2678 2678 Torrsjøn S 29.07.2011 09:33 58,9783 12,1139 102 7,619 2,35 4,98 1,76 2870 2870 Visten S 03.08.2011 11:05 59,6299 13,3968 63 4,859 6,00 3,71 32,03 SampleID Lake ID Lake Country Date Time Latitude Longitude Altitude TOC TP Cond Area.km 2 2872 2872 Stora Le S 29.07.2011 13:00 59,3222 11,7941 103 5,253 1,90 4,49 84,88 2875 2875 Näsrämmen S 03.08.2011 14:40 60,0339 14,1374 239 7,602 5,85 3,11 2,73 2878 2878 Rangsjön S 31.07.2011 09:56 60,8236 12,5075 490 7,365 3,20 2,22 2,74 2887 2887 Tisjön S 31.07.2011 10:59 60,9188 12,9657 431 6,525 4,55 1,78 27,05 2888 2888 Halsjøen S 30.07.2011 17:23 60,8640 12,3111 299 12,275 5,80 1,28 5,18 2899 2899 Jangen S 03.08.2011 12:25 60,1463 13,2715 212 9,409 5,25 2,97 4,51 3017 3017 Sör -älgen S 04.08.2011 09:45 59,7243 14,6065 181 7,219 5,85 3,11 15,54 3019 3019 Möckeln S 03.08.2011 09:48 59,3039 14,5383 88 7,538 12,60 5,34 17,99 3020 3020 Ljusnaren S 05.08.2011 17:45 59,8792 14,9342 163 8,522 6,75 3,59 9,60 3025 3025 Halvarsnoren S 04.08.2011 10:53 59,6318 14,5964 165 7,441 5,40 3,54 16,88 3027 3027 Nätsjön S 03.08.2011 15:40 59,8913 14,4809 206 4,993 3,45 2,84 2,92 3029 3029 Örlingen S 04.08.2011 12:50 59,8790 14,4200 190 6,836 6,70 2,74 1,40 3031 3031 Saxen S 04.08.2011 11:50 59,7738 14,4097 178 5,612 4,85 4,53 7,02 3106 3106 Långbjörken S 02.08.2011 15:55 59,7680 15,9521 92 9,503 9,65 3,75 1,67 3160 3160 Skattungen S 31.07.2011 15:39 61,1941 14,9452 200 7,553 4,25 2,81 19,50 3165 3165 Bäsingen S 02.08.2011 17:15 60,1606 16,3895 67 6,107 8,50 4,13 12,74 3167 3167 Tisken S 01.08.2011 17:05 60,5309 15,6727 107 7,063 8,00 5,82 63,50 3185 3185 Stora Almsjön S 31.07.2011 11:58 60,8784 13,2081 463 12,545 5,55 2,54 2,04 3189 3189 Dragsjön S 31.07.2011 13:27 60,9927 13,5987 451 10,780 7,25 1,88 1,44 3201 3201 Milsjön S 03.08.2011 13:40 60,1946 14,0086 291 9,118 3,75 2,37 3,29 3220 3220 Stora Korslängen S 05.08.2011 18:40 59,9829 15,2304 262 5,943 2,05 2,2 3,45 3384 3384 Hinsen S 01.08.2011 16:10 60,6821 16,0743 191 5,492 3,55 3,32 11,90 3397 3397 Storsjön S 01.08.2011 13:37 61,6539 15,7592 355 3,768 3,15 2,9 1,89 3399 3399 Grycken S 31.07.2011 16:55 61,5760 15,4245 240 6,299 3,30 2,24 3,17 3516 3516 Holmsjön S 01.08.2011 11:51 62,4103 15,3132 244 5,856 3,10 3,69 50,73 SampleID Lake ID Lake Country Date Time Latitude Longitude Altitude TOC TP Cond Area.km 2 3541 3541 Stornaggen S 01.08.2011 10:44 62,2666 15,9710 357 7,174 4,70 2,7 3,04 5000 5000 Forsjösjön S 02.08.2011 10:00 58,9535 16,2635 27 12,895 16,30 13,2 1,87 10000 10000 Hurdalsjøen N 12.08.2011 11:05 60,3765 11,0408 176 3,711 1,60 2,77 32,81 10001 10001 Harestuvatnet N 12.08.2011 13:10 60,1932 10,7121 234 4,060 4,35 6,27 1,98

Basic characteristics of lakes sampled Lake ID Project lake identifier Lake Lake name Country Country of lake's location (N = Norway, S = Sweden) Date Sampling date (DD.MM.YYYY) Time Sampling time of day Latitude Latitude of sampling site (decimal degrees, WGS84, from airplane GPS) Longitude Longitude of sampling site (decimal degrees, WGS84, from airplane GPS) Altitude Lake altitude (m) TOC Total Organic Carbon (mg/L; NIVA/UiO), Shimadzu TOC analyzer TP Total Phosphorus (µg/L: below detection limit (= <1) coded as -1; NIVA/UiO), molybdate -blue method (auto -analyzer) on persulfate di gested sample Cond Specific conductivity (mS/m) Area.km 2 Lake surface area (km²) Table S2. Relative abundance of different eukaryotic supergroup/group sequences and OTUs and occurrence of groups in n lakes of the whole set of Scandinavian lakes ( n = 74).

Supergroup Group % of total OTUs % of total reads Occurrence, n lakes Amoebozoa Amoebozoa 0.21 < 0.1 6 Archaeplastida Charophyta 0.53 0.19 62 Chlorophyta 7.12 3.26 74 other Archaeplastida < 0.1 < 0.1 8 Centroheliozoa Centroheliozoa 3.35 4.00 70 Cryptophyta Cryptophyta 7.12 26.60 74 Excavata Excavata 0.32 < 0.1 16 Haptophyta Haptophyta 0.43 0.20 45 Katablepharida Katablepharida 0.58 1.28 74 Opisthokonta Fungi 6.64 1.59 74 Choanozoa 1.65 1.29 71 Ichthyosporea 0.53 0.10 33 other Opisthokonta 0.11 < 0.1 24 Alveolata Ciliophora 11.85 10.95 74 Dinoflagellata 20.40 24.02 74 Perkinsea 5.37 2.97 74 other Alveolata 2.66 0.96 74 Rhizaria Cercozoa 7.49 3.81 74 Stramenopiles Bicosoecida 1.54 0.20 63 Chrysophyceae 12.86 11.10 74 Bacillariophyceae 1.22 1.23 67 Dictyochophyceae 1.38 0.52 73 MAST 0.74 0.22 67 Oomycota 0.48 0.24 62 Raphidophyceae 0.53 1.05 42 Synurophyceae 2.02 3.05 73 Eustigmatales 0.32 < 0.1 45 Hyphochytriales 0.21 < 0.1 16 Labyrinthulida 0.16 < 0.1 26 other Stramenopiles 1.65 0.57 73 Telonemia Telonemia 0.21 0.31 46 Incertae sedis Apusomonadida < 0.1 < 0.1 3 unassigned unassigned 0.21 < 0.1 25 Table S3. Summary of diversity indices.

Shannon, Simpson, inv.Simpson Non -rarefied species richness Rarefied species richness LakeID Shannon (H) Simpson (D) inv.Simpson S.obs S.chao1 se.chao1 S.ACE se.ACE S.obs S.chao1 se.chao1 S.ACE se.ACE 170 3,63 0,93 14,57 342 463,11 29,59 468,77 11,80 162 243,62 25,14 260,78 9,37 180 4,10 0,96 25,35 309 453,84 34,09 473,95 12,69 201 349,78 40,08 361,95 11,58 189 3,75 0,91 11,56 345 683,04 70,27 730,56 17,69 217 500,16 72,22 479,58 12,65 191 4,04 0,96 22,66 412 531,89 26,72 560,68 12,01 193 395,54 56,24 394,41 12,57 194 4,49 0,98 40,37 392 612,68 44,35 669,86 15,88 237 464,53 55,99 485,73 13,46 214 4,02 0,94 16,24 407 598,95 42,29 578,77 12,79 204 349,92 39,44 370,05 11,71 233 4,38 0,97 32,68 370 565,00 42,19 590,01 13,51 212 376,16 45,13 369,58 11,50 236 3,81 0,92 11,92 470 730,01 50,86 713,60 15,14 199 325,98 33,37 398,08 13,64 242 4,74 0,98 57,88 356 571,33 47,73 546,82 12,26 245 384,42 35,98 384,22 11,47 252 4,60 0,96 24,68 531 770,91 43,82 810,24 16,20 283 585,08 70,48 524,13 14,45 258 4,72 0,98 58,00 419 603,08 39,89 600,64 12,95 241 359,15 30,49 377,74 10,84 261 1,88 0,71 3,40 145 233,44 29,37 249,41 9,08 65 131,60 30,95 158,93 8,73 264 3,84 0,96 22,70 296 451,22 38,44 449,27 11,68 151 284,00 42,79 297,42 10,94 277 2,32 0,75 3,93 221 328,33 30,03 340,69 10,47 111 162,11 18,39 196,54 9,20 285 2,84 0,86 7,02 202 339,57 35,85 394,52 12,32 124 282,05 51,97 294,72 9,98 288 3,62 0,92 12,57 289 424,13 34,37 424,34 11,07 173 296,48 36,06 301,87 11,15 326 4,61 0,98 41,06 406 625,49 45,33 624,34 13,90 255 434,14 42,35 481,04 13,40 328 3,04 0,86 7,35 278 447,53 39,99 483,44 13,05 143 313,15 54,38 317,94 11,42 339 3,10 0,88 8,43 313 460,87 33,65 498,59 12,46 160 319,68 44,56 386,18 12,88 340 4,37 0,97 31,01 560 871,95 52,78 935,55 18,12 232 443,69 52,27 494,18 13,91 344 3,68 0,94 17,04 432 641,82 41,13 689,60 15,53 170 356,24 54,44 368,43 12,51 345 4,13 0,96 27,88 382 603,80 47,06 599,55 13,72 197 444,73 68,48 447,69 13,73 Shannon, Simpson, inv.Simpson Non -rarefied species richness Rarefied species richness LakeID Shannon (H) Simpson (D) inv.Simpson S.obs S.chao1 se.chao1 S.ACE se.ACE S.obs S.chao1 se.chao1 S.ACE se.ACE 349 4,71 0,98 54,99 494 728,12 44,42 763,58 15,50 249 464,00 52,94 465,03 12,79 353 4,38 0,96 25,67 536 816,86 53,93 770,60 14,79 249 445,63 46,61 485,14 14,00 361 3,72 0,91 11,72 376 645,63 60,56 587,04 12,95 189 296,33 30,03 321,09 11,38 363 4,83 0,99 68,68 519 761,17 45,97 753,99 14,50 264 471,02 48,30 526,38 14,50 374 3,91 0,94 17,87 332 487,77 36,92 487,13 11,76 195 374,52 49,12 390,60 12,59 378 4,38 0,97 33,49 419 609,84 42,54 589,48 12,66 205 455,14 74,44 365,37 11,03 380 3,63 0,92 12,74 358 489,21 29,61 519,30 12,29 181 357,79 50,16 351,43 11,08 394 3,99 0,97 30,83 283 420,33 36,62 407,90 10,70 149 303,59 52,96 274,18 9,50 395 4,24 0,97 29,48 363 531,81 39,62 527,15 12,19 198 319,42 33,96 332,73 10,66 405 4,26 0,97 33,32 321 519,72 49,59 495,37 12,31 182 292,70 33,33 296,59 9,87 433 4,71 0,98 60,56 451 622,59 37,36 623,30 13,42 238 421,08 46,34 444,23 13,42 436 4,79 0,98 59,64 395 542,09 32,51 560,37 12,83 259 422,29 39,28 442,05 12,22 453 4,64 0,98 51,53 461 791,08 62,13 814,52 16,88 255 454,81 46,60 493,53 13,85 481 3,37 0,88 8,54 357 569,11 46,35 569,10 13,13 167 346,08 51,97 388,52 13,30 486 3,19 0,89 8,80 214 362,53 41,55 356,70 11,21 139 267,62 42,25 272,80 9,63 487 4,85 0,97 39,21 670 938,60 46,55 938,95 16,14 313 480,19 34,86 571,17 15,69 498 4,95 0,99 79,73 376 635,50 54,38 647,28 15,06 281 520,70 54,90 554,74 15,30 519 4,67 0,98 47,03 460 645,01 35,80 710,62 15,06 254 403,04 35,84 454,30 13,22 2252 4,61 0,97 39,35 422 603,25 36,37 638,01 14,36 253 498,92 59,24 525,33 14,76 2268 4,64 0,98 40,71 429 666,54 46,87 674,76 14,77 264 432,80 38,74 497,93 14,64 2312 4,43 0,96 27,16 411 533,82 26,78 570,20 13,03 233 366,22 34,21 386,58 11,33 2374 4,43 0,96 26,08 414 590,23 36,90 599,81 13,34 256 469,06 48,80 532,22 15,34 2678 4,24 0,96 26,23 234 376,50 35,87 398,85 12,05 219 359,14 36,39 381,68 12,10 2870 4,74 0,98 48,09 449 787,35 67,03 740,92 15,67 265 466,07 48,52 438,80 11,44 Shannon, Simpson, inv.Simpson Non -rarefied species richness Rarefied species richness LakeID Shannon (H) Simpson (D) inv.Simpson S.obs S.chao1 se.chao1 S.ACE se.ACE S.obs S.chao1 se.chao1 S.ACE se.ACE 2872 4,57 0,97 35,51 514 709,10 36,46 759,02 15,03 251 392,22 33,94 445,29 12,97 2875 4,87 0,98 54,22 430 689,11 50,50 712,97 15,68 279 481,71 47,15 494,00 13,88 2878 4,46 0,98 44,44 332 515,96 43,84 508,18 12,27 202 343,21 41,12 330,99 10,38 2887 4,65 0,98 47,99 454 592,06 28,20 634,64 14,04 245 393,75 36,66 428,70 12,44 2899 4,17 0,95 19,80 522 765,11 44,71 771,59 15,11 235 477,38 58,53 507,87 14,02 3017 4,99 0,99 80,66 574 859,89 49,36 900,69 16,75 284 430,08 33,64 477,29 13,73 3019 4,75 0,98 48,93 431 701,88 52,59 706,74 15,12 275 516,73 55,69 529,26 14,74 3020 4,73 0,98 63,21 361 544,10 38,19 607,28 15,02 243 452,46 52,71 470,24 13,68 3025 4,91 0,99 68,71 523 761,86 44,23 779,56 15,15 286 499,29 48,23 501,30 13,64 3027 4,99 0,99 69,93 420 656,97 45,85 701,31 15,91 304 465,28 34,40 575,53 15,93 3029 4,97 0,99 75,31 681 1040,49 57,85 1018,57 17,44 303 601,53 63,72 667,55 16,85 3031 4,87 0,98 61,90 462 687,36 44,21 718,79 15,36 270 465,07 46,64 467,42 12,57 3106 5,10 0,98 54,61 610 851,41 42,08 876,15 15,72 347 674,22 65,55 673,72 15,91 3160 4,84 0,99 71,08 542 782,34 45,05 787,27 15,26 258 459,37 49,33 455,19 12,69 3165 5,14 0,99 93,28 771 1180,04 62,06 1184,73 18,68 326 728,78 85,05 744,29 17,55 3167 4,21 0,96 27,28 335 499,34 35,79 560,41 14,36 207 459,12 69,52 439,75 13,72 3185 5,15 0,99 92,56 596 933,57 58,67 890,52 16,34 319 488,90 35,99 562,06 15,30 3189 5,12 0,99 83,89 462 808,89 64,45 790,40 16,08 326 624,33 62,49 633,78 15,97 3201 4,65 0,98 53,36 515 813,77 55,78 801,95 16,40 247 551,94 76,96 514,23 14,62 3220 4,92 0,98 65,60 431 707,54 54,72 694,57 14,92 291 567,52 61,86 577,17 14,66 3384 5,02 0,99 80,30 549 742,61 35,67 782,90 14,92 290 479,81 43,57 474,85 12,41 3397 4,65 0,98 54,17 430 676,67 49,38 694,30 15,27 245 515,00 67,16 504,43 14,02 3399 4,74 0,98 55,71 353 533,09 38,49 558,58 13,65 257 434,40 42,28 487,50 14,01 3516 5,04 0,99 79,39 598 857,88 45,47 872,01 16,08 319 689,31 77,45 737,25 17,25 Shannon, Simpson, inv.Simpson Non -rarefied species richness Rarefied species richness LakeID Shannon (H) Simpson (D) inv.Simpson S.obs S.chao1 se.chao1 S.ACE se.ACE S.obs S.chao1 se.chao1 S.ACE se.ACE 3541 4,83 0,98 58,89 531 792,38 49,44 782,34 15,03 276 544,93 62,80 550,48 15,10 5000 4,03 0,94 17,74 411 649,98 50,82 619,38 13,20 203 353,41 40,09 375,07 11,28 10000 4,90 0,98 63,95 600 769,98 32,16 779,16 14,11 292 517,51 49,28 570,17 15,68 10001 4,63 0,98 52,32 458 654,78 40,45 659,43 13,92 247 488,58 58,87 502,35 14,64 Table S4. Kendall's Tau correlation tests between DCA and NMDS axes for NMDS ordination run in two (k = 2) and three (k = 3) dimensions. Significant correlation between axes (p < 0.05) is indicated by bold font.

DCA1 DCA2 Kendall's tau p-value Kendall's tau p-value k = 2 NMDS1 -0.8430 < 0.0001 0.0218 0.7831 NMDS2 -0.0433 0.5851 -0.8282 < 0.0001 k = 3 NMDS1 -0.8334 < 0.0001 0.0863 0.2769 NMDS2 -0.1411 0.0754 0.8134 < 0.0001 Table S5. Permutation-based significance probabilities (p.perm) between NMDS axes scores and rarefied richnesses (S.rar) of the most abundant taxonomic groups.

S.rar by Intercept NMDS1 NMDS2 R2.dev p.perm taxonomic group Total 5,4303 0,4104 -0,1332 0,471 0,001 Amoebozoa -2,4786 -0,5228 -1,0381 0,048 0,593 Bacillariophyceae 1,2654 0,4243 -0,4230 0,074 0,037 Bicosoecida 0,3180 0,9240 -0,4705 0,101 0,050 Centroheliozoa 1,9885 0,4264 -0,3475 0,095 0,021 Cercozoa 2,7079 0,3635 -0,4980 0,234 0,001 Charophyta -0,1384 -0,3097 -1,0095 0,151 0,004 Chlorophyta 2,5172 -0,0967 -0,3156 0,061 0,096 Choanozoa 1,3396 0,3491 -0,4028 0,091 0,018 Chrysophyceae 3,4102 0,9073 0,1614 0,570 0,001 Chytridiomycota 1,5375 0,5150 -0,6801 0,222 0,001 Ciliophora 3,5356 0,3039 -0,2632 0,208 0,001 Cryptomycota 0,3648 0,4781 -0,1984 0,037 0,270 Cryptophyta 3,1990 -0,1442 0,0281 0,068 0,075 Dictyochophyceae 1,2097 0,7483 0,3232 0,174 0,001 Dinoflagellata 3,9141 0,5775 0,0416 0,406 0,001 Excavata -1,4423 -0,3888 -1,5176 0,168 0,011 Fungi -0,5564 1,3955 -0,9351 0,184 0,003 Haptophyta -0,4017 1,0187 -0,5081 0,110 0,015 Ichthyosporea -1,0029 0,8888 -0,0426 0,038 0,423 Katablepharida 1,1600 0,0573 -0,3958 0,070 0,041 Oomycota -0,1684 0,2131 -1,1422 0,164 0,003 Perkinsea 2,1376 0,3335 0,3512 0,090 0,035 Raphidophyceae -0,0526 0,7801 -1,5045 0,269 0,001 Synurophyceae 1,7743 0,2874 -0,4394 0,109 0,018 Table S6. Results of the RDA variation partitioning for protistan community data.

Notes: Values in the table are adjusted R 2 with associated p-values. [E | S] is the sole effect of environmental factors, [S | E] is the sole effect of spatial predictors, and [E+S] is the shared effect between environmental and spatial gradients. RDA is variation partitioning for Hellinger transformed OTU table, S.rar is rarefied OTU richness, H is Shannon diversity, D is Simpson diversity, NMDS1 and NMDS2 are site scores for axes 1 and 2, respectively. References: Ptacnik R, Andersen T, Brettum P, Lepistö L & Willén E (2010) Regional species pools control community saturation in lake phytoplankton. Proceedings of the Royal Society B: Biological Sciences 277 : 3755 -3764.