Research Article |
Corresponding author: Bernhard Seifert ( bernhard.seifert@senckenberg.de ) Academic editor: Brian Lee Fisher
© 2015 Bernhard Seifert, Sandor Csosz.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Seifert B, Csösz S (2015) Temnothorax crasecundus sp. n. – a cryptic Eurocaucasian ant species (Hymenoptera, Formicidae) discovered by Nest Centroid Clustering. ZooKeys 479: 37-64. https://doi.org/10.3897/zookeys.479.8510
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The paper integrates two independent studies of numeric morphology-based alpha-taxonomy of the cryptic ant species Temnothorax crassispinus (Karavajev, 1926) and T. crasecundus sp. n. conducted by different investigators, using different equipment, considering different character combinations and evaluating different samples. Samples investigated included 603 individual workers from 203 nests – thereof 104 nest samples measured by Seifert and 99 by Csösz. The material originated from Europe, Asia Minor and Caucasia. There was a very strong interspecific overlap in any of the 29 shape characters recorded and subjective expert determination failed in many cases. Primary classification hypotheses were formed by the exploratory data analysis Nest Centroid (NC) clustering and corrected to final species hypotheses by an iterative linear discriminant analysis algorithm. The evaluation of Seifert’s and Csösz’s data sets arrived at fully congruent conclusions. NC-Ward and NC-K-means clustering disagreed from the final species hypothesis in only 1.9 and 1.9% of the samples in Seifert’s data set and by 1.1 and 2.1% in Csösz’s data set which is a strong argument for heterospecificity. The type series of T. crassispinus and T. crasecundus sp. n. were allocated to different clusters with p = 0.9851 and p = 0.9912 respectively. The type series of the junior synonym T. slavonicus (Seifert, 1995) was allocated to the T. crassispinus cluster with p = 0.9927. T. crasecundus sp. n. and T. crassispinus are parapatric species with a long contact zone stretching from the Peloponnisos peninsula across Bulgaria northeast to the southern Ukraine. There is no indication for occurrence of interspecifically mixed nests or intraspecific polymorphism. However, a significant reduction of interspecific morphological distance at sites with syntopic occurrence of both species indicates local hybridization. The results are discussed within the context of the Pragmatic Species Concept of
Numeric morphology-based alpha-taxonomy, Pragmatic Species Concept, parapatric species, hybridization, intraspecific dimorphism
The small Formicoxenine ants Temnothorax nylanderi (Förster, 1850) and T. crassispinus (Karavajev, 1926) are dominant elements of the forest floor fauna of European temperate woodland biomes. They have repeatedly stood in the focus of thorough studies after
According to mtDNA data (
Frequent hybridization of T. nylanderi and T. crassipinus along the front-line in Germany has been shown (
Very early it became obvious to the senior author (Seifert) that there existed a third cryptic entity of the T. nylanderi complex in the Balkans, Asia Minor and Caucasus that may occur syntopic with T. crassispinus. These preliminary investigations, however, came to a complete standstill after three rejections of research funding proposals in the years 2007–2009. More recently, the junior author (Csösz) could investigate further material from the Balkans and Asia Minor. Csösz’s data considerably improved the knowledge about the relations of the two species in the contact zone. We fuse in this paper the independent approaches of two investigators to a broad basis of evidence for the existence of the new cryptic species T. crasecundus sp. n. which is a sibling species of T. crassispinus. The contact situation is comparable to that found in T. nylanderi and T. crassispinus. T. crasecundus sp. n. and T. crassispinus are parapatric – the front line in Greece and Bulgaria is known and predicted to run diagonally through Romania and Moldova.
T. crasecundus sp. n. and T. crassispinus are truly cryptic species. Each of the 29 investigated shape characters and absolute size show a considerable interspecific overlap even in nest sample means and much more so on individual level. Furthermore, there is no clear signal provided by pigmentation and overall phenotypic impression. Both authors, having a long experience in identification of Temnothorax ants by simple eye inspection, are able to make a fair subjective guess on the species identity in a good number of samples but fail in many others. Both species thus fully fit the definition of cryptic species by
Morphology is essential to establish the link between species delimitation and Linnean nomenclature (
In this paper, we firstly present our argumentation why T. crasecundus sp. n. has a separate species status and why local hybridization is no argument against heterospecificity. Then we give the formal description of the new species.
A total of 203 nest samples with 603 individual workers of both species was investigated – 104 samples by Seifert and 99 by Csösz. Only one nest series, the type series of T. crassispinus (Karavajev, 1926), was investigated by both Csösz and Seifert. Though genetically representing the same nest population we treated these two samples as operationally different because of the deviating character systems of the investigators and their different individual selection of workers from the type series.
A total of 119 nest samples originated from the following countries: Austria 13, Bosnia 2, Bulgaria 7, Czechia 3, Germany 39, Greece 24, Hungary 8, Italy 2, Macedonia 3, Moldova 2, Poland 5, Serbia 1, Slovenia 4 and Ukraine 7 samples. A detailed account of the samples is given in the following under the sequence site, date in the format yyyy.mm.dd, sample No [latitude in decimal format, longitude in decimal format, altitude].
AUSTRIA: Arnoldstein, 1994.06.03, [46.550 N, 13.704 E, 570 m]; Bad Vöslau, pre 1980 [47.967 N, 16.222 E, 250 m]; Einöd-0.5 km N, 1994.08.05, No 40 [48.318 N, 15.732 E, 250 m]; Göttlesbrunn, 1955.08.25, [48.058 N, 16.738 E, 170 m]; Innsbruck, Kalvarienberg, 1994.08.05, No H2 [47.283 N, 11.435 E, 630 m]; Innsbruck, 1944.04.02 [ 47.266 N, 11.408 E, 580 m]; Kärnten: Gösselsdorfer See, 1994.08 [46.566 N, 14.618 E, 680 m]; Paudorf-1 km N, 1994.07.13, No 41, g82 [48.358 N, 15.618 E, 350 m]; Roppen-1 km E, 1994.07.04, No 207 [47.223 N, 10.831 E, 730 m]; Seebenstein: Türkensturz, pre 1970 [47.679 N, 16.136 E, 540 m]; Wachau: Spitz-5 km NW, 1990.07.01, No 2774 [48.392 N, 15.358 E, 400 m]; Wellersdorf-2 km NW, 1994.07.11 [46.567 N, 14.169 E, 770 m]. BOSNIA: Golubic-10 km N, 1989.09.15, No 51, 56 [44.768 N, 15.929 E, 400 m]. BULGARIA: Kokalyane-3 km E, 2009.06.06, No 080, 1033, 1035 [42.579 N, 23.434 E, 650 m]; Vitosha: Vladaya district, 2004, No 590, 856 [42.630 N, 23.205 E, 930 m]; Vitosha: Zheleznitsa- 1 km N, 2009.06.06, No 1009 [42.546 N, 23.365 E, 1000 m]; Vratehansica Planina: Vratsa-5 km S, 2009.06.07, No 1074 [43.137 N, 23.591 E, 1000 m]. CZECHIA: Masovice-3.7 km S, 1997.09.18, No 227 [48.824 N, 15.977 E, 285 m]; Strelna-0.5 km E, 1993.09.28, No 02, 03 [50.667 N, 13.755 E, 350 m]. GERMANY: Abensberg-10 km E, 2000.09.05, No AB1, AB2 [48.812 N, 11.969 E, 425 m]; Adlersberg- 0.5 km W, 2000.09.06, No AD/M6, AD/M7 [49.042 N, 12.004 E, 453 m]; Altenhain-1 km NW, 1994.06, No 07, 08 [51.299 N, 12.685 E, 15 m]; Berching, 2001.09, No 1, 3 [49.115 N, 11.444 E, 410 m]; Berzdorf, 1981.04.07 [51.055 N, 14.886 E, 274 m]; Berzdorf, 1993.03.19, No g31 [51.055 N, 14.886 E, 274 m]; Genthin-3.6 km S, 1994.06, No 6, 8, 04 [52.375 N, 12.150 E, 39 m]; Glesien-1.6 km W, 1994.06, No 3, 5 [51.445 N, 12.205 E, 122 m]; Hohburg-1 km S, 1993.06.19, No g77 [51.405 N, 12.799 E, 170 m]; Koldenhof, 2000.11.02, No-000074 [53.330 N, 13.340 E, 119 m]; Kühren-1 km W, 1993, No 94 [51.875 N, 11.978 E, 58 m]; Löbauer Berg, 1983.07.13 [51.089 N, 14.692 E, 392 m]; Meissen, Bosel, 1982.06.09, No 303 [51.138 N, 13.514 E, 170 m]; Mertitz, 2001.06.25 [51.177 N, 13.321 E, 138 m]; Obergruna-1 km S, 1993.04.12, No 59, 145 [51.005 N, 13.316 E, 285 m]; Ponholz-2 km SE, 2000.09.06, No P10, P11 [49.151 N, 12.119 E, 399 m]; Quolsdorf-2.8 km NE, 1992.04.06 [51.402 N, 14.872 E, 115 m]; Serrahn, 2000.05.04, No 000036, 17 [53.670 N, 12.350 E, 62 m]; Steffenhagener Heide, 2007.04.27 [54.110 N, 13.295 E, 7 m]; Technitz-1.2 km WNW, 1994.06, No 1, 6, 8 [51.130 N, 13.051 W, 165 m]; Trebsen-2.1 km SW, 1994, No 02, 03, 08, 09, 44 [51.272 N, 12.697 E, 165 m]; Uchtspringe-0.5 km S, 1995, No 10 [52.534 N, 11.605 E, 75 m]. GREECE: Chelmos, 1994.06.04, No 1394 [37.987 N, 22.198 E, 2000 m]; Kalamata-20 km E, 1994.06.01, No 1348, 1349 [37.080 N, 22.280 E, 1250 m]; Kastanitsa-4 km SW; 2000.04.22, No 20 [37.280 N, 22.670 E, 1300 m]; Konitsa-5 km N, 1996.05.23, No 355 [40.108 N, 20.764 E, 550 m]; Levidi-10 km S, 2000.04.27, No 118, 119, 137 [37.630 N, 22.280 E, 1700 m]; Meliana vic., 1996.05.21, No 024 [39.361 N, 20.787 E, 700 m]; Parnon Mts., 2011.04.26, No 117, 145 [37.270 N, 22.610 E, 1650 m]; Sitena vic., 2000.04.05, No 56 [37.300 N, 22.650 E, 1000 m]; Sitena-3 km W, 2000.04.25-66, 071,079 [37.300 N, 22.600 E, 1700 m]; Sparti-20 km SW, 2000.04.29, No 150b, 150c [36.970 N, 22.350 E, 1950 m]; Taigetos Oros, 2011.04.30, No 305 [36.948 N, 22.377 E, 1500 m]; Tripolis-15 km NNW, 2011.04.29, No 214, 221 [37.629 N, 22.302 E, 1000 m]; Tripolis-22 km NNW, 2011.04.29, No 234, 238 [37.654 N, 22.268 E, 1600 m]; Vamvakou-3 km SE, 2011.04.26, No 162 [37.22 N, 22.59 E, 1300 m]; Vitina-5 km NE, 2011.04.29, No 261 [37.681 N, 22.208 E, 1000 m]. HUNGARY: Budapest, 1909.05.21 [47.543 N, 18.966 E, 250 m]; Csillebérc, 2004.05.30, No 397 [47.500 N, 18.959 E, 400 m]; Hatvan, 2011.03.12, No 432, 433 [47.673 N, 19.647 E, 200 m]; Isaszeg, 2011.04.02, No 437 [47.535 N, 19.397 E, 200 m]; Pécs (leg. Kaufmann), pre 1945 [46.104 N, 18.245 E, 200 m]; Vérteskozma, 2009.04.30, No 411 [46.459 N, 18.432 E, 350 m]. ITALY: Villa Santina, 1989.09.18, No 03, 04 [46.403 N, 12.844 E, 460 m]. MACEDONIA: Jacupitsa, 2009.06.19, No 001 [41.418 N, 21.416 E, 1300 m]; Konopishte, 2009.06.19, No 003 [41.248 N, 22.079 E, 672 m]; Lukovo, 2009.06.16, No 004 [41.366 N, 20.606 E, 590 m]. MOLDOVA: Vall du Berlad (Barlad Valley), pre 1930, No 1, 2 [47 N, 28 E, 70 m]. POLAND: Kielce: Pongrac [50.83 N, 20.66 E, 300 m]; Osiecznica-3 km W, 1994.04.09, No 85, 104 [51.344 N, 15.379 E, 187 m]; Wolin: Wapnica-3.5 km E, 2006, No 46, 79 [53.883 N, 14.485 E, 35]. SERBIA: Sremska Kamenica, 1971.05.02 [45.22 N, 18.44 E, 300 m]. SLOVENIA: Knezac, 1989.09.17, No 10, 11 [45.619 N, 14.250 E, 615 m]; Postoijna-12 km W, 1997.05.29, No 401 [45.778 N, 14.062 E, 900 m]; Novo Mesto, 2007.05.29, No WL16 [45.811 N, 15.169 E, 200 m]. UKRAINE: Cherkassy: Kanev Nat. Res. [49.711 N, 31.477 E, 220 m]; Donetzk, S riv. Donetz, 1982.07.16 [47.9 N, 37.8, 120 m]; Golosseyev, pre 1926 [50.5 N, 30.5 E, 140 m]; Kiev vic., 1988.07.08 [50.5 N, 30.5 E, 140 m]; Pervomaijsk, 1998.06.10 [48.044 N, 30.859 E, 100 m]; Pischa, 2004.06.10 [51.071 N, 21.003 E, 170 m].
A total of 84 nest samples originated from the following countries: Armenia 1, Bulgaria 31, Georgia 6, Greece 15, Romania 4, Russia 2, Turkey 21 and Ukraine 4 samples. A detailed account of the samples is given in the following.
ARMENIA: Armenia: without site, 1986.06.11 [40 N, 45 E, 1600 m]. BULGARIA: Arkutino, 1978.08.01 [42.33 N, 27.77 N, 15 m]; Bistrits-1 km N, 2009.06.06, No 1004 [42.594 N, 23.363 E, 400 m]; Dobrostan, 1982.09.12-55 [41.93 N, 24.88 E, 1500 m]; Dospat-2SW, 2009.0610, No 1203 [41.634 N, 24.149 E, 1300 m]; German Monastery, 2004.05, No 467, 470 [42.602 N, 23.434 E, 800 m]; Harsovo-1 km SE, 2009.06.09, No 1130, 1130/1, 1131 [41.458 N, 23.390 E, 200 m]; Kiten, 2011.04.29, No 373 [42.238 N, 27.772 E, 27 m]; Kokalyane-3 km E, 2009.0606, No 079 [42.579 N, 23.434 E, 650 m]; Malko Tarnovo: Propada, 2009.07.26, No 370 [41.982 N, 27.492 E, 385 m]; Malko Tarnovo: Brashlyan 2009.08.22, No 374 [42.044 N, 27.427 E, 340 m]; Mladezhko, 2009.08.21, No 366 [42.152 N, 27.362 E, 220 m]; Novakovo-2 km SE, 2009.06.12, No 1299 [41.887 N, 25.099 E, 400 m]; Obsor, 1981.08.01 [42.82 N, 27.86 E, 50 m]; Pasarel-1 km NW, 2009.06.06, No 070, 078 [42.594 N, 23.362 E, 770 m]; Peshtera, 2008.05.25, No 369 [42.297 N, 24.299 E, 580 m]; Pirin Mts.: Rozen-8 km N, 1982.09.05, No 0, 396 [41.60 N, 23.45 E, 1400 m]; Plovdiv, 1977.05.27 [42.14 N, 24.72 E, 200 m]; Sofia: Borisova Park 1, 2004, No 841 [42.680 N, 23.342 E, 596]; Sofia: Borisova Park 2, 2004, No 509, 558 [42.678 N, 23.351 E, 586]; Sofia: Lozenetz distr., 2004, No 709 [42.666 N, 23.312 E, 620 m]; Tsaparevo-2 km S, 2009.0609, No 1168 [41.612 N, 23.097 E, 800 m]; Vitosha: Vladaya distr., 2004, No 596, 854 [42.630 N, 23.205 E, 930 m]; Zeleznitza-1 km N, 2009.06.06, No 043, 047 [42.539 N, 23.367 E, 1000 m]. GEORGIA: Abastumani-1.4 km W, 2013.09.17, No GEOII-70, GEOII-71, GEOII-72 [41.758 N, 42.817 E, 1532 m]; Daba-0.26 km E, 2010.08 [41.811 N, 43.452 E, 1030 m]; Pizunda, 1984.08.11 [43.153 N, 40.342 E, 17 m]; Sedaseni-Kloster, 2004.07.28 [41.871 N, 44.768 E, 1150 m]. GREECE: Karitza-6 km W, 1998.04.04 [39.840 N, 22.710 E, 750 m]; Kastanitza-6 km W, 1998.04.04 [37.28 N, 22.67 E, 1300 m]; Kosmas-2 km SW, 2000.04.26, No 93, 96, 098 [37.080 N, 22.730 E, 1100 m]; Levidi-10 km S, 2000.04.27, No 132 [37.630 N, 22.280 E, 1700 m]; Litohoro-3 km W, 1996.05.13 [40.080 N, 22.450 E, 1200 m]; Litohoro-7 km W, 1996.05.13, No 187 [40.112 N, 22.480 E, 600 m]; Parnon Mts., 2011.04.26, No 152 [37.27 N, 22.61 E, 1650 m]; Sitena-3 km W, 2000.04.25, No 62, 71a [37.300 N, 22.600 E, 1700 m]; Sparti-20 km SW, 2000.04.25, No 149 [36.97 N, 22.35 E, 1950 m]; Stagira, 2011.04.09, No 027 [40.531 N, 23.720 E, 585 m]; Taigetos Oros, 2011.04.30, No 302, 311 [36.948 N, 22.377 E, 1500 m]. ROMANIA: Comana: Vlasca, pre 1935, No 1, 2 [43.90 N, 28.31 E, 130 m]; Dobrogea: Babadag, 2005.06.02, No 105, 106 [44.857 N, 28.691 E, 100 m]. RUSSIA: Gelendzhik-5 km SSE, 2006.06.04, No 249 [44.48 N, 38.145 E, 150 m]; Obilnoje, 2006.06.08, No 219 [44.207 N, 43.538 E, 240 m]. TURKEY: Aydogdu-5 km SW, No 1147, 1157, 1158 [40.714 N, 42.495 E, 1500 m]; Cat-2 km S, 2012.07.02, No 046 [39.418 N, 35.957 E, 1550 m]; Demirköy, 2009.07.06, No 474, 476 [41.818 N, 27.814 E, 170 m]; Erzincan-25 km SE, 2012.07.13, No 179 [39.661 N, 39.734 E, 1200 m]; Eskishir-Sögüt, 2003.05.10, No 162 [39.550 N, 30.130 E, 966 m]; Ispir-10 km NW, 1993.07.01, No 1197 [40.585 N, 40.852 E, 1700 m]; Mahya hill, 2005.05.30, No 472 [41.771 N, 27.638 E, 820 m]; Mezraa vic., 2012.07.11, No 153, 159 [39.378 N, 39.805 E, 1300 m]; Ordu-25 km NW, 2012.07.21, No 396 [41.064 N, 37.711 E, 400 m]; Ordu-40 km WSW, 2012.07.21, No 402 [40.719 N, 37.622 E, 1000 m]; Posof-3 km E, 2012.07.17, No 303 [41.414 N, 42.762 E, 1500 m]; Pülümür vic., 2012.07.11, No 159 [39.482 N, 39.890 E, 1600 m]; Seydiler-7 km N, 2012.07.23, No 495 [41.694 N, 33.718 E, 1200 m]; Sögüt vic., 2003.05.10, No 166 [39.570 N, 30.13 E, 1050 m]; Tortum-15 km E: Kirecli Gecidi, 2012.07.13, No 205 [40.353 N, 41.704 E, 2400 m]; Tortum-45 km NNE, 2012.07.11, No 217 [40.325 N, 41.572 E, 1600 m]; Ulu Dag: Sogukpinar, 1993.07.05, No 1230 [40.055 N, 29.120 E, 750 m]. UKRAINE: Agarmis, 1980.09.29 [45.252 N, 35.025 E, 600 m]; Armiansk, 1985.05.04 [46.107 N, 33.693 E, 15 m]; Krasnolesye, 1980.09.12 [44.952 N, 34.102 E, 100 m]; Simferopol, 1995.08.13, No 826 [44.938 N, 34.099 E, 300 m].
Leptothorax nylanderi var. crassispina Karavajev, 1926
The type series, stored in Shmalhausen Institute of Zoology Kiev and certainly representing a nest sample, is labeled “Kiev: Golosejev (Karavaijev No 3057)”. Three syntype workers were investigated by Seifert and seven syntype workers by Csösz.
Leptothorax nylanderi slavonicus Seifert, 1995
The paratypes, seven workers on the same pin and originating from the nest that contained the queen holotype, are labeled “Kr. Görlitz, 19.3.1993, Schönau-Berzdorf, Hutberg, g31”, „Leptothorax n. slavonicus Seifert”, “Paratypes”. Four paratypes of this sample were investigated by Seifert. Material is stored in Senckenberg Museum of Natural History Görlitz. Csösz investigated four worker paratypes of another nest series from the type locality, labelled “Germany, Kr. Görlitz, Hutberg Schönau-Berzdorf, 19.03.1993 Seifert”.
Temnothorax crasecundus sp. n.
The worker holotype is labelled “BUL: 42.6785°N, 23.3508°E Sofia, 586 m, Borisova gradina Park, Part 2 V.Antonova 2004.05-509” and “Holotype Temnothorax crasecundus Seifert & Csösz”. Four worker paratypes, two males and two gynes from the holotype nest are mounted on two other pins and carry the same collecting data label and “Paratypes Temnothorax crasecundus Seifert & Csösz”. A second series with five worker paratypes on two pins is labeled “BUL: 42.6797°N, 23.3417°E Sofia, 596 m, Borisova gradina Park, Part 1 V.Antonova 2004.05-841” and “Paratypes Temnothorax crasecundus Seifert & Csösz”. These two type series are stored in Senckenberg Museum of Natural History Görlitz. A third paratype series containing three workers on the same pin, is labelled “Bulgaria_28: East Rhodopes, 2 km SE Novakovo 25 km SE. Asenovgrad, 1299, 400mH, 41°53'12"N, 25°5'55"E, 12.06.2009, Leg A Schulz”, “ANTWEB CASENT 0906045“ and is stored in the Hungarian Museum of Natural History Budapest.
The senior and junior author performed two independent investigations of worker ant morphology, considering different character combinations and using different microscopic equipment. Seifert recorded 18 and Csösz 22 primary morphometric characters. In bilaterally developed characters, arithmetic means of both body sides were calculated. All measurements were made on mounted and fully dried specimens. Measurements of body parts always refer to real cuticular surface and not to the diffuse pubescence surface.
Seifert used for spatial adjustment of specimens a pin-holding stage, permitting full rotations around X, Y, and Z axes and a Leica M165C high-performance stereomicroscope equipped with a 2.0 planapochromatic objective (resolution 1050 lines/mm) at magnifications of 120–384×. The mean relative measuring error over all magnifications was 0.3%. A Schott KL 1500 cold-light source equipped with two flexible, focally mounted light-cables, providing 30°-inclined light from variable directions, allowed sufficient illumination over the full magnification range and a clear visualization of silhouette lines. A Schott KL 2500 LCD cold-light source in combination with a Leica coaxial polarized-light illuminator provided optimal resolution of tiny structures and microsculpture at highest magnifications. Simultaneous or alternative use of the cold-light sources depending upon the required illumination regime was quickly provided by regulating voltage up and down. A Leica cross-scaled ocular micrometer with 120 graduation marks ranging over 52% of the visual field was used. To avoid the parallax error, its measuring line was constantly kept vertical within the visual field.
Equipment and measurement procedures of Csösz
Measurements were made using a pin-holding stage, permitting rotations around X, Y, and Z axes. An Olympus SZX9 stereomicroscope was used at a magnification of 150× for each character, allowing a precision of ± 2 µm.
29 morphometric characters were investigated – ten of these by both authors. We give the character definitions in alphabetic order. In square brackets is indicated who investigated a character.
CL [Csösz & Seifert]: maximum cephalic length in median line; the head must be carefully tilted to the position with the true maximum. Excavations of hind vertex and/or clypeus reduce CL.
CS [Seifert]: cephalic size; the arithmetic mean of CL and CW, used as a less variable indicator of body size.
CSb [Csösz]: cephalic size; the arithmetic mean of CL and CWb.
CW [Seifert]: maximum cephalic width; the maximum is found in Temnothorax and Leptothorax usually across and including the eyes, exceptionally posterior of the eyes.
CWb [Csösz]: maximum width of head capsule, measured just posterior of the eyes.
EL [Csösz]: maximum diameter of the eye.
EYE [Seifert]: eye-size index: the arithmetic mean of the large (EL) and small diameter (EW) of the elliptic compound eye is divided by CS, i.e. EYE=(EL+EW)/(CL+CW). All structurally visible ommatidia are considered.
FRS [Csösz & Seifert]: distance of the frontal carinae immediately caudal of the posterior intersection points between frontal carinae and the lamellae dorsal of the torulus (arrows in Fig.
MGr [Seifert]: depth of metanotal groove or depression, measured from the tangent connecting the dorsalmost points of promesonotum and propodeum; here given as per cent ratio of CS.
MH [Seifert]: in workers: with mesosoma in lateral view and measured orthogonal to “longitudinal mesosomal axis”, MH is the longest measurable section line of mesosoma at mesopleural level (not height above all). “Longitudinal mesosomal axis” in lateral view is defined as straight line from the centre of propodeal lobe (centre of circus in Fig.
ML [Csösz & Seifert]: in workers: mesosoma length from caudalmost point of propodeal lobe to transition point between anterior pronotal slope and anterior propodeal shield (preferentially measured in lateral view; if the transition point is not well defined, use dorsal view and take the centre of the dark-shaded borderline between pronotal slope and pronotal shield as anterior reference point). In gynes: length from caudalmost point of propodeal lobe to the most distant point of steep anterior pronotal face.
MW [Csösz & Seifert]: maximum mesosoma width; this is in workers pronotal width, in gynes it is measured anteriorly of the tegulae.
NOdL [Csösz]: Anterior length of petiole measured in dorsal view. Distance from the (centre of anteriormost seta pit on the petiolar node to the level of the constriction of articulation condyle with propodeum (Fig.
NOH [Csösz]: Maximum height of the petiolar node, measured in lateral view from the uppermost point of the petiolar node perpendicular to a reference line set from the petiolar spiracle to the imaginary midpoint of the transition between the caudal slope and the caudal cylinder of the petiole (Fig.
NOL [Csösz]: Length of the petiolar node, measured in lateral view from petiolar spiracle to dorso-caudal corner of caudal cylinder. Do not erroneously take as reference point the dorso-caudal corner of the helcium, which is sometimes visible.
PEH [Csösz & Seifert]: maximum petiole height. The chord (dashed line in Fig.
PEL [Seifert]: diagonal petiolar length in lateral view; measured from anterior corner of subpetiolar process to dorsocaudal corner of caudal cylinder.
PEW [Csösz & Seifert]: maximum width of petiole.
PL [Csösz]: Total petiole length measured in dorsal view; distance between the dorsalmost point of caudal petiolar margin and the dorsalmost point of anterior petiolar peduncle at the transversal level of its strongest constriction. Positioning of petiole as in NOdL (Fig.
PoOc [Csösz & Seifert]: postocular distance. Use a cross-scaled ocular micrometer and adjust the head to the measuring position of CL. Caudal measuring point: median occipital margin; frontal measuring point: median head at the level of the posterior eye margin. Note that many heads are asymmetric and average the left and right postocular distance (Fig.
PPH [Csösz]: Maximum height of the postpetiole in lateral view measured perpendicularly to a line defined by the linear section of the segment border between dorsal and ventral petiolar sclerite.
PPL [Csösz]: Maximum length of the postpetiole measured in lateral view perpendicular to the straight section of lateral postpetiolar margin (Fig.
PPW [Csösz & Seifert]: maximum width of postpetiole.
SL [Csösz & Seifert]: maximum straight line scape length excluding the articular condyle as arithmetic mean of both scapes.
SP [Seifert]: maximum length of propodeal spines; measured in dorsofrontal view along the long axis of the spine, from spine tip to a line, orthogonal to the long axis, that touches the bottom of the interspinal meniscus (Fig.
SPL [Csösz]: Minimum distance between the center of propodeal spiracle and the margin of subspinal excavation measured with both end points positioned in the same focal level (Fig.
SPBA [Csösz & Seifert]: the smallest distance of the lateral margins of the spines at their base. This should be measured in dorsofrontal view, since the wider parts of the ventral propodeum do not interfere with the measurement in this position. If the lateral margins of spines diverge continuously from the tip to the base, a smallest distance at base is not defined. In this case, SPBA is measured at the level of the bottom of the interspinal meniscus.
SPST [Csösz & Seifert]: distance between the centre of propodeal stigma and spine tip. The stigma centre refers to the midpoint defined by the outer cuticular ring but not to the centre of real stigma opening that may be positioned eccentrically.
SPTI [Csösz & Seifert]: the distance of spine tips in dorsal view; if spine tips are rounded or truncated, the centres of spine tips are taken as reference points.
SPWI [Csösz]: Maximum distance between outer margins of spines; measured in same position as SPBA.
The delimitation of the cryptic species was done by an interaction of Nest-Centroid Clustering (NC clustering) and a confirmative linear discriminant analysis (LDA). NC Clustering was run both as hierarchical NC-Ward clustering and non-hierarchical NC-K-means clustering. These methods were described in more detail by
The same mathematical procedures were applied for the data sets of Seifert and Csösz. NC-Ward clustering was run first to indicate the putative number of K main clusters. In the second step NC-K-Means was performed with the setting of K classes suggested by NC-Ward. Classifications being coincident between the hierarchical and non-hierarchical clustering formed the hypothesis for the controlling LDA that was subsequently run. Samples with classifications disagreeing between NC-Ward and NC-K-means were run in this LDA as wild-cards. The final classification (“final species hypothesis”) was established by the LDA in the iterative procedure described by
The W Palaearctic species of the Temnothorax nylanderi species complex – T. nylanderi, T. crassispinus and T. crasecundus sp. n. – can be separated from other species of the region by the following character combination.
Head short, mean index CL/CW only 1.053–1.063.
Whole dorsum of vertex regularly and continuously longitudinaly carinulate, shining surface areas are absent or restricted to a narrow median stripe.
Metanotal depression always visible, at least suggested.
Antennal club and femora never with a blackish pigmentation
Petiole in lateral view rather high and with a weakly concave frontal face; the anterior profiles of node and peduncle form an angle of about 150–155° whereas anterior and dorsal profiles of node form an angle of 90–105°. Dorsal profile of node steeply sloping down to caudal cylinder. The profiles of this slope and of the caudal cylinder form an angle of about 140°.
Propodeal spines acute, deviating from longitudinal axis of mesosoma by 32–42° and moderately long, SP/CS 0.200–0.260, SPST/CS 0.253–0.356.
In the data set of Seifert and considering all 18 characters, NC Ward clustering provided a clear separation of the cryptic species in two main branches (Fig.
The results were similar in the data set of Csösz, considering all 22 characters and following the same procedure. The classifications of NC-Ward and NC-K-means differed in 11.1% of samples. NC-Ward disagreed from the final species hypothesis in 11.1% and NC-K-means in 3.1% of samples. A character reduction by a stepwise LDA, again performed iteratively, improved the classification result significantly. NC-Ward disagreed from the final species hypothesis in only 2.1% (Fig.
We consider the congruent results of two independent investigations and investigators and the low disagreement of 1.1–2.1% between the classifications of exploratory data analyses with the final species hypothesis as a strong argument for heterospecificity of T. crasecundus sp. n. and T. crassispinus. This interpretation is supported by the coincidence of phenotyping with a clear-cut parapatric distribution (Fig.
Temnothorax crasecundus sp. n. (grey branch) and T. crassispinus (black branch). NC-Ward clustering. Data set of Csösz: 99 nest samples and 10 characters considered. Arrows point to samples clustered in disagreement with the final species hypothesis. Figs
We achieved an agreement of 97.9–98.9% between the final species hypothesis and four different cluster analyses. This indicates a significant evolutionary divergence and could indicate that no or very few hybrids are present in our data set. Yet, the present problem is more difficult because extremely similar species are involved which prevents a reliable phenotypic identification of a particular hybrid sample. A broader statistic approach is needed. Accordingly, we compare the positions of samples along the interspecific LDA vector (i) from the potential contact zone, (ii) from the allopatric zones and (iii) from sites with established syntopic occurrence of both species. Reduced interspecific distances in contact zones are an indication that hybridization could have occurred. We consider the territories of Greece, Bulgaria, Romania and Moldova to form the potential contact zone (Fig.
There is only a very weak, insignificant reduction of the distance from the zero point of the discriminant vector in samples from the potential contact zone compared to those from the allopatric zones (Tab.
Distance of the sample means of Temnothorax crassispinus and T. crasecundus sp. n. from the zero point of the interspecific discriminant vector. The ANOVA data are placed in the line between the compared data sets.
Distance from zero | |
---|---|
Potential contact zone (total 84, crassispinus 34, crasecundus 50) | 1.6173 ± 1.0000 [0.1186.3.8912] |
ANOVA [F, p] | 0.370, 0.543 |
Allopatric zones (total 119, crassispinus 85, crasecundus 34) | 1.6941 ± 0.7906 [0.0326,3.4717] |
ANOVA [F, p] | 11.267, 0.001 |
Syntopic sites (total 31, crassispinus 18, crasecundus 13) | 1.1465 ± 0.8740 [0.1186,3.4894] |
ANOVA [F, p] | 12.383, 0.001 |
Potential contact zone without syntopic sites (total 53, crassispinus 16, crasecundus 37) | 1.8927 ± 0.9727 [0.1258,3.8912] |
We conclude that T. crassispinus and T. crasecundus sp. n. are a pair of cryptic, parapatric species showing hybridization in the contact zone. We follow here the Pragmatic Species Concept of
Seifert proposed as operational criterion for the discipline “multivariate investigation of ant worker morphology” and as a remedy against over-splitting that at least 97% of the classifications by exploratory data analyses should agree with the classifications by linear discriminant analyses that form the final species hypothesis. However, 4% disagreement or 96% confirmation seem to be a more reasonable threshold when considering the performance of the two most powerful methods of NC-clustering, NC-K-Means and NC-Ward, in the separation of 74 cryptic ant species (Table 1 in
The consequences of threshold decisions may be illustrated by an example. Over many years we studied the situation in the Mediterranean Temnothorax lichtensteini complex and intended to accept three species: the largely western entity T. lichtensteini (Bondroit, 1918), a second, largely eastern, semipatric entity (provisionally designated by
We want to emphasize in this context that hybridization and reticulate evolution is a matter of fact in the living nature around us (reviewed, e.g., in Abbott et al. 2013). Exemplary groups in animals are ducks (
Finally we address the question if our two entities might represent an intraspecific dimorphism instead of representing different species by considering geographic distribution and the frequency and distribution of supposedly mixed nests in the sympatric and allopatric zones. Frequencies of discrete morphs provided by the gene pool of a single species may show steep geographic gradients - remind alone of the famous text book example of the Peppered Moth Biston betularia (e.g.,
In the provisional internal naming system of the senior author the new species had been designated over the years as “Temnothorax crassispinus sp. 2”. The taxonomic name, composed of “cra” (first syllable of crassispinus) and “secundus” (= the second), intends to indicate both this history and the close relationship.
See above under “Material”.
All morphometric data given in the following verbal description are arithmetic means of 256 worker individuals calculated by fusing Seifert’s and Csösz’s data sets. Harmonization of the different data sets has been performed by the function CW = 1.0791 * CWb.
Worker (Tables
Data set of Seifert; nest sample means of morphometric data of the workers of the cryptic species T. crasecundus sp. n. and T. crassispinus (Karavajev, 1926). Arrangement of data: arithmetic mean ± standard deviation [minimum, maximum]. F values and significance levels p are from an univariate ANOVA; the F values of the most separating characters are given in heavy type.
T. crasecundus sp. n. (n=29) | ANOVA F, p |
T. crassispinus (n=75) | |
---|---|---|---|
CS | 641 ± 36 [594,713] | 0.02, n.s. | 645 ± 30 [556,713] |
CL/CW | 1.065 ± 0.014 [1.038,1.087] | 16.90, 0.000 | 1.053 ± 0.013 [1.013,1.081] |
SL/CS | 0.767 ± 0.011 [0.741,0.786] | 1.38, n.s. | 0.764 ± 0.013 [0.734,0.790] |
PoOc/CL | 0.395 ± 0.007 [0.380,0.406] | 3.63, n.s. | 0.392 ± 0.007 [0.373,0.405] |
EYE/CS | 0.210 ± 0.005 [0.201,0.219] | 29.22, 0.000 | 0.216 ± 0.005 [0.207,0.233] |
FRS/CS | 0.361 ± 0.008 [0.344,0.376] | 2.48, n.s. | 0.364 ± 0.008 [0.345,0.386] |
SPBA/CS | 0.280 ± 0.010 [0.252,0.297] | 26.53, 0.000 | 0.294 ± 0.012 [0.268,0.322] |
SPTI/CS | 0.325 ± 0.014 [0.295,0.347] | 75.95, 0.000 | 0.350 ± 0.013 [0.324,0.377] |
SPST/CS | 0.285 ± 0.014 [0.250,0.308] | 179.20, 0.000 | 0.323 ± 0.014 [0.286,0.354] |
SP/CS | 0.218 ± 0.015 [0.188,0.246] | 202.63, 0.000 | 0.260 ± 0.012 [0.232,0.290] |
PEW/CS | 0.253 ± 0.007 [0.242,0.270] | 6.76, 0.011 | 0.258 ± 0.008 [0.239,0.275] |
PPW/CS | 0.353 ± 0.010 [0.335,0.375] | 24.77, 0.000 | 0.367 ± 0.013 [0.331,0.395] |
PEH/CS | 0.357 ± 0.008 [0.343,0.374] | 14.22, 0.000 | 0.364 ± 0.009 [0.343,0.391] |
PEL/CS | 0.472 ± 0.011 [0.452,0.497] | 7.41, 0.008 | 0.479 ± 0.013 [0.446,0.508] |
ML/CS | 1.187 ± 0.016 [1.158,1.221] | 0.08, n.s. | 1.188 ± 0.015 [1.156,1.239] |
MW/CS | 0.605 ± 0.010 [0.580,0.628] | 3.68, n.s. | 0.600 ± 0.011 [0.780,0.626] |
MH/CS | 0.524 ± 0.011 [0.500,0.547] | 11.93, 0.001 | 0.532 ± 0.012 [0.507,0.566] |
MPGR/CS | 2.10 ± 0.56 [1.31,3.41] | 0.25, n.s. | 2.04 ± 0.49 [1.07,3.23] |
Data set of Csösz; nest sample means of morphometric data of the workers of the cryptic species T. crasecundus sp. n. and T. crassispinus (Karavajev, 1926). Arrangement of data: arithmetic mean ± standard deviation [minimum, maximum]. F values and significance levels p are from an univariate ANOVA; the F values of the most separating characters are given in heavy type.
T. crasecundus sp. n. (n=55) | ANOVA F, p | T. crassispinus (n=44) | |
---|---|---|---|
CSb | 614 ± 37 [539,718] | 1.73, n.s. | 623 ± 36 [544,688] |
CL/CWb | 1.155 ± 0.018 [1.120,1.196] | 14.03, 0.000 | 1.140 ± 0.021 [1.082,1.180] |
SL/CSb | 0.791 ± 0.013 [0.763,0.836] | 6.40, 0.013 | 0.784 ± 0.013 [0.756,0.811] |
PoOc/CL | 0.390 ± 0.007 [0.379,0.405] | 0.00, n.s. | 0.390 ± 0.007 [0.370,0.410] |
EL/CSb | 0.255 ± 0.007 [0.236,0.279] | 0.71, n.s. | 0.256 ± 0.005 [0.247,0.268] |
FRS/CSb | 0.375 ± 0.008 [0.358,0.397] | 1.57, n.s. | 0.377 ± 0.008 [0.362,0.400] |
MW/CSb | 0.630 ± 0.010 [0.610,0.662] | 1.37, n.s. | 0.627 ± 0.017 [0.590,0.678] |
ML/CSb | 1.227 ± 0.017 [1.193,1.268] | 1.01, n.s. | 1.223 ± 0.023 [1.183,1.303] |
SPBA/CSb | 0.298 ± 0.012 [0.277,0.321] | 32.89, 0.000 | 0.312 ± 0.013 [0.282,0.339] |
SPTI/CSb | 0.333 ± 0.016 [0.300,0.384] | 96.86, 0.000 | 0.365 ± 0.016 [0.332,0.397] |
SPWI/CSb | 0.352 ± 0.018 [0.295,0.405] | 110.18, 0.000 | 0.388 ± 0.016 [0.355,0.422] |
SPST/CSb | 0.289 ± 0.015 [0.253,0.319] | 220.08, 0.000 | 0.330 ± 0.011 [0.298,0.356] |
SPL/CSb | 0.162 ± 0.006 [0.149,0.175] | 17.06, 0.000 | 0.157 ± 0.008 [0.137,0.175] |
PEW/CSb | 0.269 ± 0.008 [0.252,0.288] | 10.18, 0.002 | 0.275 ± 0.011 [0.255,0.317] |
PEH/CSb | 0.372 ± 0.009 [0.354,0.398] | 7.88, 0.006 | 0.377 ± 0.009 [0.361,0.400] |
NOH/CSb | 0.169 ± 0.007 [0.156,0.202] | 29.11, 0.000 | 0.177 ± 0.007 [0.164,0.189] |
NOL/CSb | 0.254 ± 0.010 [0.232,0.274] | 4.68, 0.033 | 0.258 ± 0.008 [0.237,0.275] |
NODL/CSb | 0.296 ± 0.014 [0.266,0.318] | 0.02, n.s. | 0.295 ± 0.017 [0.267,0.340] |
PPW/CSb | 0.368 ± 0.009 [0.348,0.387] | 19.17, 0.000 | 0.377 ± 0.012 [0.348,0.405] |
PPH/CSb | 0.350 ± 0.008 [0.335,0.368] | 7.74, 0.007 | 0.356 ± 0.011 [0.340,0.384] |
PPL/CSb | 0.255 ± 0.009 [0.234,0.273] | 1.04, n.s. | 0.257 ± 0.008 [0.236,0.272] |
PL/CSb [%] | 0.413 ± 0.011 [0.383,0.434] | 5.74, 0.019 | 0.419 ± 0.013 [0.393,0.454] |
Mesosoma moderately wide (MW/CS 0.608) and metanotal depression always developed (MpGr/CS 2.1%). Propodeal spines rather long and acute but distinctly shorter than in T. crassipinus (SPST/CS 0.283 vs. 0.322). Distance of their bases and tips rather large but significantly smaller than in T. crassipinus (SPBA/CS 0.286 vs. 0.300, SPTI/CS 0.324 vs. 0.353), spine tips slightly curving inwards. Direction of spines in lateral view deviating from longitudinal axis of mesosoma by 26–29°. Mesosoma irregularly microreticulate-rugulose with few superimposed longitudinal rugae on promesonotum. Metapleuron more regularly longitudinally carinate-rugose.
Petiole in lateral view rather high and with a weakly concave frontal face; the anterior profiles of node and peduncle form an angle of about 150–155° whereas anterior and dorsal profiles of node form an angle of 90–105°. Dorsal profile of node weakly convex or nearly straight and moderately long, steeply sloping down to caudal cylinder. The profiles of this slope and of the caudal cylinder form an angle of about 140°. Whole surface of petiolar and postpetiolar nodes microreticulate with a mesh width of 9–13 µm. Two longitudinal rugae typically demarcate the margin of dorsal petiolar plane while the sides of petiolar tergites are stabilized by one longitudinal carina/ruga on each side.
Overall body color dirty yellow to light brown with a strong yellowish component. Mesosoma, appendages, waist and basis of first gaster tergite usually lighter yellow to dirty yellow. Head dorsum and the posterior surfaces of gaster tergites usually darker, generally yellowish brown. Lighter heads occur.
There is considerable overlap in each of the 29 shape characters and absolute size (Tables
D(6) = 22.058*PoOc+17.640*SL-66.166*SPST+38.233*MW-28.926*PPW -35.873*SPTI-1.797
classified 203 nest samples with an error of 3.4%. Samples with an arithmetic mean of D(6) < 0 are determined as T. crassispinus and those with larger values as T. crasecundus sp. n. With PoOc, SL and SPST being recorded bilaterally, a trained investigator needs for the resulting nine measurements about 15 minutes per individual.
The most simple means for separation of T. crasecundus from T. nylanderi is geography: the shortest distance between a site of both species is 1000 km and a closing of this broad gap is prevented by habitat saturation of the omnipresent, highly competitive T. crassipinus. A rather simple phenotypical species delimitation is possible using three absolute measurements. With all measurements given in mm, the discriminant
D(3) = 129.53*SPBA–120.88*PPW+133.8*MpGR +4.446
classified 87 nest samples with an error of 3.4%. Samples with an arithmetic mean of D(3) < 0.64 are determined as T. nylanderi and those with larger values as T. crasecundus sp. n.
The present zoogeography (Fig.
We wish to thank Vera Antonova / Sofia and Andreas Schulz / Leverkusen for providing a lot of samples from the Balkans and Asia Minor, Alexandr Radchenko / Kiev for enabling a loan of type specimens of T. crassispinus and Michele Esposito / San Francisco and Roland Schultz / Görlitz for providing z-stack photos of type specimens of T. crassispinus and T. crasecundus.