Pinuxylon alonissianum Mantzouka & Sakala 2019

DESCRIPTION Macroscopic description The sample belongs to a xylitic remnant of dark brown colour with the following dimensions: 6 × 5 × 4 cm (Fig. 3 D). Microscopic description Transverse section. Growth ring boundaries distinct, transition from early- to latewood gradual (Fig. 5 A), latewood tracheids thin walled (Fig. 5 A-D), large resin canals (axial and radial distributed but mainly at the latewood) with thin-walled epithelial cells (Fig. 5 - C), bordered pits in radial walls with well-defined disc-shaped torus (Fig. 5 D), no axial parenchyma observed. Tangential section. Exclusively uniseriate rays (fusiform because they contain radial intercellular canals and have a specific shape), average height of rays 4 - 12 cells (mostly “ medium ” sized but they can be up to 20 cells). Resin canals in rays with thin-walled epithelial cells (Fig. 5 L-O). Radial section. Tracheid pits uniseriate (Fig. 5 E, F). Ray tracheids present with up to four rows of cells (Fig. 5 G, H, K), cell walls of ray tracheids smooth (Fig. 5 G, K) and occasionally slightly dentate (Fig. 5 G-I), ray tracheid pit borders with dentate thickenings (Fig. 5 I). End walls of ray parenchyma cells smooth (Fig. 5 J, K). Horizontal walls of ray parenchyma cells smooth (Fig. 5 J, K), no indentures were observed, cross-field pitting pinoid in 1 - 2 rows of usually 1 - 2 pits per cross-field but up to 6 (Fig. 5 E, G, H, J, K).

Pinuxylon halepensoides van der Burgh (van der Burgh 1973) differs from our wood in having 7 - 8 pits in tangential tracheid walls, pits of piceoid, cupressoid and pinoid type and up to two pits per cross-field.

Pinuxylon vateri (Platen) Rössler (Rössler 1937) differs from our wood in having taxodioid and pinoid pits per cross-field. Pinuxylon ponderosoides van der Burgh (van der Burgh 1973) is very close to our sample but it has only up to four pinoid pits per cross field. As resumed from the aforementioned comparison none of the above-mentioned species was identical to the new finding from Alonissos. Consequently the fossil pine wood from Alonissos Island is declared as a new species, named according the ICBN rules (McNeill et al. 2006: art. 23; art. 60, ICBN – Vienna Code) after the place where it was found: Pinuxylon alonissianum Mantzouka & Sakala, sp. nov. BOTANICAL AFFINITIES The distinction between the sections Pinus (Diploxylon or hard pines) and Strobus (Haploxylon or soft pines) has been discussed more than a hundred years ago (Shaw 1914) and along with the history of the fossil findings of the genus is well documented in Xu et al. (2015). This divergence took place probably during the mid-Cretaceous (Keeley 2012). Following the identification key by van der Burgh (1973) we come to the conclusion that our fossil modern relative belongs to one of the following: section Pinaster, or section Sula (P. leucodermis), or section Leiophylla or section Lumholtzii. Following the classification by Price et al. (1998) and the detailed work of the wood anatomical characteristics of the conifers by Esteban et al. (2004) we examined the modern species belonging to Diploxylon pines (and especially the species of the sections that van der Burgh’s identification key had revealed) in order to find the botanical affinities of ours: 1) Section Pinus (Subsection Pinus – because some of the species revealed following van der Burgh’s identification key are now belonging to this subsection). Pinus densiflora Siebold & Zuccarini, P. mugo Turra, P. mugo subsp. uncinata (Ramond) Domin, P. nigra J. F. Arnold, P. resinosa Aiton, P. sylvestris L. and P. tabuliformis Carrière are excluded because they have 1 - 2 - fenestroid pits per cross-field. Pinus heldreichii H. Christ has rays very high of up to 30 cells, ray parenchyma with pitted horizontal walls and 1 - 2 pinoid pits per cross-field. Pinus kesiya Royle ex Gordon has axial parenchyma. Pinus massoniana Lambert, P. lumholtzii B. L. Robinson & Fernald and P. pinaster Aiton have biseriate bordered pits on the radial walls of the axial tracheids. Pinus merkusii Junghuhn & de Vriese ex de Vriese and P. thunbergii Parlatore have biseriate bordered pits on the radial walls of the axial tracheids and fenestroid cross-field pits. 2) Section Pinus (Subsection Canarienses – Canary Islands, Himalayas). Pinus canariensis C. Smith ex Buch has axial parenchyma and biseriate bordered pits on the radial walls of the axial tracheids. Pinus roxburghii Sargent (former named as P. longifolia) has biseriate bordered pits on the radial walls of the axial tracheids (Groom & Rushton 1913). 3) Section Pinus (Subsection Halepenses – S. Europe, W. Asia, N. Africa). Pinus brutia Tenore, is excluded because it has ray parenchyma with nodular end walls and with pitted horizontal walls. Pinus halepensis Miller is excluded because it has biseriate bordered pits on the radial walls of the axial tracheids and ray parenchyma with nodular end walls and with pitted horizontal walls. 4) Section Pinus (Subsection Pineae – S. Europe). Pinus pinea L. is excluded because it has biseriate bordered pits on the radial walls of the axial tracheids and ray parenchyma with nodular end walls and with pitted horizontal walls. 5) Section Pinus (Subsection Leiophyllae – Mexico and adjacent SW USA). Pinus leiophylla Schiede ex Schlechtendal & Chamisso has up to two pinoid pits per cross-field. P. leiophylla var. chihuahuana (Engelm.) Shaw has up to four pits per cross-field. Therefore we believe that this species could be the modern analogue of our fossil. 6) Section Pinaster. Pinus banksiana Lambert, P. caribaea Morelet, P. clausa (Chapman ex Engelmann) Vasey ex Sargent, P. coulteri D. Don, P. echinata Miller, P. glabra Walter, P. hartwegii Lindley, P. lawsonii Roezl ex Gordon & Glendinning, P. montezumae Lambert, P. occidentalis Swartz, P. palustris Miller, P. patula Schiede ex Schlectendahl & Chamisso, P. pseudostrobus Lindley, P. pungens Lambert, P. radiata D. Don, P. rigida Miller, P. sabiniana Douglas ex D. Don, P. serotina Michaux, P. taeda L. are excluded because they have biseriate bordered pits on the radial walls of the axial tracheids. Pinus attenuata Lemmon, P. elliottii Engelmann, P. muricata D. Don, P. ponderosa Douglas ex Lawson, P. teocote Schiede ex Schlechtendal & Chamisso, P. torreyana Parry ex Carrière, P. virginiana Miller are excluded for having ray parenchyma with nodular end walls and / or with pitted horizontal walls. Pinus contorta Douglas ex Loudon is excluded for having ray parenchyma with nodular end walls and fenestroid cross-field pits. From this section it seems that Pinus arizonica Engelmann ex Rothrock, P. arizonica var. cooperi (C. E. Blanco) Farjon, P. engelmannii Carrière, P. jeffreyi Greville & Balfour could also resemble the modern representatives of our fossil.

DISCUSSION ON THE WOOD ANATOMICAL CHARACTERISTICS The occurrence of thin walled resin canals (axial and radial), the absence of parenchyma cells, the uniseriate tracheid pitting, the pinoid cross-field pitting, the existence of ray tracheids, the smooth horizontal and end walls of ray parenchyma indicate that our fossil belongs to Pinaceae and more precisely to the genus Pinus L. The cell walls of ray tracheids of our sample have been described as “ smooth and occasionally slightly dentate ” following the terminology by Ickert-Bond (2001: 361) and in Ray tracheids with up to four rows of cells, cell walls of ray tracheids slightly dentate (white arrow), cross-field pitting pinoid in 2 horizontal rows of usually 1 - 2 pits per cross-field but up to 6; I, cell walls of ray tracheids slightly dentate (white arrows), ray tracheid pit borders with dentate thickenings (black arrow); J, end walls of ray parenchyma cells smooth, horizontal walls of ray parenchyma cells smooth, cross-field pitting pinoid in 1 - 2 rows of usually 1 - 2 pits per cross-field but up to 6; K, tracheid pits uniseriate. Ray tracheids with up to four rows of cells, cell walls of ray tracheids smooth, end walls of ray parenchyma cells smooth, horizontal walls of ray parenchyma cells smooth, cross-field pitting pinoid in 1 - 2 rows of usually 1 - 2 pits per cross-field but up to 6; L-O, exclusively uniseriate rays (fusiform because they contain radial intercellular canals and have a specific shape), average height of rays 4 - 12 cells (mostly “ medium ” sized but they can be up to 20 cells). Resin canals in rays with thin-walled epithelial cells. A - D, transversal section; E - K, radial longitudinal sections; L - O, tangential longitudinal sections. Scale bars: A, 400 μm; B, C, E, N, 50 μm; D, F-K, 25 µm; L, 200 μm; M, O, 100 μm. accordance with the “ smooth as well as slightly dentate ray tracheids ” as discussed in van der Burgh (1964: 254 - 259; 1973: 104 - 106) and in Dolezych et al. (2011: 53). The same kind of identification of the cell walls of ray tracheids has been pointed out as “ inconspicuously dentate ” in Phillips (1941: 265, feature 17, pl. 15, fig. 30, judging from the “ thickest parts ”) and as “ less prominent (and with outer tracheid walls typically sinuous) dentate ” (cause the term “ smooth ” is used only for generally thin walls with no ornamentation at all) discussed in IAWA Committee (2004: 43, feature 82). In Greece a lot of fossil pine remnants have been found (e. g. Velitzelos et al. 2014; Zidianakis et al. 2016) but only a few pine fossils in the form of wood have been identified until now. Moreover, the fossil pine cones and seeds from Greece have been related to Pinus roxburghii Sarg. and P. canariensis C. Sm. (Mai & Velitzelos 2002, 2007; Boyd 2009; Kvaček et al. 2014). Our specimen has been compared with the fossil pine woods from Greece and has been found to have a lot of differences with them: Pinoxylon parenchymatosum Süss & Velitzelos (Süss & Velitzelos 1993) from Lemnos Island is characterized by the occurrence of axial parenchyma, a feature not in accordance with our fossil. Pinoxylon paradoxum Süss & Velitzelos (Süss & Velitzelos 1994) from Lesbos Island is characterized by the existence of spirals and P. pseudoparadoxum (Süss & Velitzelos 1994, Lesbos Island) has high rays – up to 50 – cells. Our specimen differs also from two more species from Lesbos Island identified by Süss & Velitzelos (2009): from Lesbosoxylon (= Pinoxylon) diversiradiatum Süss & Velitzelos because the latter one has bi-seriate rays and from L. (= Pinoxylon) graciliradiatum Süss & Velitzelos which has large rays with “ idioblast-like ” cells. There are also differences with Lesbosoxylon (= Pinoxylon) ventricosuradiatum Süss & Velitzelos (e. g. rays with “ idioblast-like ” cells and pits in the cross-field and ray tracheids not ascertainable) (Süss & Velitzelos 2010). Our specimen has also been compared with Pinuxylon sp. cf. Pinus sylvestris L. (Iamandei et al. 2011, 2016) and found different because the fossil from the Romanian Carpathians has axial parenchyma and 2 - 3 - seriate rays. It has been also compared with Pinoxylon yabei Shimakura (Shimakura 1936) which is completely different from our fossil having axial parenchyma, bordered pits arranged in three rows, thick-walled epithelial cells of resin canals, occurrence of traumatic canals and 1 - 3 seriate rays with up to 70 cells height. Our fossil has dentate horizontal walls of ray tracheids, therefore it is assigned to Diploxylon pines. So, we immediately reject from comparison the fossil pine woods belonging to Haploxylon type. Wang et al. (2017) have made a list including the majority of the fossil pine woods described in literature and their characteristics. This study has divided their list in two categories: a) Haploxylon; and b) Diploxylon pines. Haploxylon The fossil Haploxylon pines (with smooth horizontal walls of ray tracheids as also seen in Wang et al. 2017: table 1) which are rejected from being correlated with our fossil are the following: Pinus uniseriata H. B. Wang, A. A. Oskolski & Z. K. Zhou (Wang et al. 2017), P. albicauloides S. K. Choi & K. Kim (Choi et al. 2010), Pinuxylon chemrylensis Blokhina (Blokhina 1995), Pinoxylon dakotense Knowlton emend. Read (Nishida & Nishida 1995), Pinuxylon cembraeforme Rössler (Rössler 1937), P. microporosum Ogura (Nishida & Nishida 1995) or P. microporum Ogura (Ogura 1944), which has no ray tracheids (and as also discussed in Blokhina & Bondarenko [2016] shouldn’t be assigned to Pinuxylon), P. similkameenensis Miller (Miller 1973), Pinuxylon sp. (Tao et al. 1994; Zhang et al. 2008 as discussed in Wang et al. 2017), P. woolardii Tidwell, Parker & Folkman (Tidwell et al. 1986), P. zobelianum (Göppert) Kräusel (van der Burgh 1964). Our research has also revealed some more Haploxylon fossil pine woods not in accordance with our fossil described by: 1) Greguss (1967), Petrescu & Bican-Brişan (2004) & Iamandei et al. (2016): Pinuxylon haploxyloides Greguss (only with a single pinoid cross field pit; holotype still available in Budapest, see in Sakala et al. 2018), P. albicauloides Greguss (has only fenestroid cross field pits), P. tarnocziense (Tuzśon) Greguss and Pinuxylon sp.; 2) Dolezych et al. (2011): Pinuxylon succiniferum (Göppert) Kräusel emend. Dolezych; 3) Vozenin-Serra (1971): Pinuxylon nightigalense Vozenin-Serra (as discussed in Iamandei et al. 2016); 4) van der Burgh (1964): Pinuxylon parryoides (Gothan) Kräusel emend. van der Burgh; and 5) Iamandei & Iamandei (2000): Pinuxylon marinasii Iamandei & Iamandei. Diploxylon The selected Diploxylon fossil pines of the literature for the purposes of the work by Wang et al. (2017) which have similarities with our fossil wood are: Pinus nanfengensis Wang, Oskolski & Zhou (Wang et al. 2017), Pinus cf. armandii Franchet (Yi et al. 2005), Pinus hatamuraenase Jeong & Kim (Jeong et al. 2012), Pinus henanensis J. J. Yang (Yang et al. 1990, 1996; Qi et al. 2005, as discussed in Wang et al. 2017), Pinuxylon arjuzanxianum Huard (Huard 1966; van der Burgh 1973), Pinuxylon eschweilerence van der Burgh (van der Burgh 1973), Pinuxylon halepensoides van der Burgh (van der Burgh 1973), Pinuxylon parryoides (Kräusel) van der Burgh (van der Burgh 1964, 1973; Iamandei 2000), Pinuxylon paxii Kräusel (Rössler 1937; van der Burgh 1973), Pinuxylon pinastroides (Kraus) Stockmans & Willière (van der Burgh 1973), Pinuxylon ponderosoides van der Burgh (van der Burgh 1973), Pinuxylon taedioides Kräusel (van der Burgh 1964, 1973; former Pinuxylon landensis Huard, 1966), Pinuxylon tarnocziense (Tuzśon) Greguss (van der Burgh 1973), Pinuxylon vateri (Platen) Rössler (Rössler 1937). P. nanfengensis (Wang et al. 2017) from the late Miocene of the Xianfeng Basin (central Yunnan, southwestern China) is different from our sample in terms of having partially biseriate rays (and not only uniseriate), smooth to slightly pitted ray tracheids and different cross-field pitting (pinoid and taxodioid). Other similarities There are also similarities with the fossil pines from China, i. e., Pinus armandii Franchet (Yi et al. 2002) and P. cf. armandii (Yi et al. 2005) which belong to Diploxylon pines of Cembra section but they have different cross-field pitting (fenestroid in 1 - 2 rows with 1 - 4 pits instead of pinoid in 1 - 2 rows and 1 - 6 pits in our sample). Our fossil has similarities with Pinus hatamuraense Jeong & Kim (Jeong et al. 2012) from the Akita Prefecture but it differs at the height of rays (P. hatamuraense has very long rays up to 32 cells), the fact that our fossil does not have parenchyma but P. hatamuraense does and the cross field pitting which, in the case of P. hatamuraense, includes also taxodioid pits. Pinus henanensis J. J. Yang (as described in Wang et al. 2017 because the original sources are published in Chinese only) is also different from our sample having very high rays consisting of up to 33 cells and pinoid and taxodioid cross-field pitting.

ETYMOLOGY. — The epithet, alonissianum, is due to the origin of the described material (Alonissos Island, Greece).

HOLOTYPE. — Designated here. Specimen DMALNS 6 (Repository: Museum of Geology and Palaeontology in the National and Kapodistrian University of Athens).

TYPE HORIZON. — Lignite horizon. AGE. — Early Miocene. TYPE LOCALITY. — Votsi, Central Alonissos Island, Greece. DIAGNOSE. — Coniferous wood with distinct growth ring boundaries, gradual transition from early to latewood, thin walled latewood tracheids, axial and radial resin canals (mainly at the latewood), thin-walled epithelial cells, bordered pits in radial walls with welldefined disc-shaped torus, no axial parenchyma observed, rays: exclusively uniseriate, fusiform rays with average height of rays 4 - 12 (- 20) cells, tracheid pits uniseriate, ray tracheids present with up to four rows of cells, cell walls of ray tracheids smooth and occasionally slightly dentate, ray tracheid pit borders with dentate thickenings, horizontal and end walls of ray parenchyma cells smooth, no indentures observed, cross-field pitting pinoid in 1 - 2 rows of 1 - 2 (- 6) pits per cross-field.