Biodiversity of ecosystems in an arid setting: The late Albian plant communities and associated biota from eastern Iberia

Orthoptera gen. et sp. indet. Elcanidae? Orthoptera San Just Phytophagous Preempheria antiqua Empheriidae Psocodea San Just Detritivorous, decayed leaf-litter habitat Protoculicoides hispanicus Ceratopogonidae Diptera San Just Blood-feeder Protoculicoides sanjusti Ceratopogonidae Diptera San Just Blood-feeder Pseudoscorpiones gen. et sp. indet. Pseudoscorpiones San Just Predator, decayed leaf-litter habitat Psocodea gen. et sp. indet. Psocodea San Just and Arroyo de la Pascueta Detritivorous, decayed leaf-litter habitat Raphidioptera gen. et sp. indet. Raphidioptera Arroyo de la Pascueta Predator Serphites silban Serphitidae Hymenoptera San Just Parasitoid? Spinomegops aragonensis Lagonomegopidae Araneae San Just Predator Tenuelamellarea estefaniae Lamellareidae Acariformes San Just Detritivorous, decayed leaf-litter habitat Trhypochthonius lopezvallei Trhypochthoniidae Acariformes San Just Detritivorous, decayed leaf-litter habitat Utrillabracon electropteron Braconidae Hymenoptera San Just Parasitoid https://doi.org/10.1371/journal.pone.0282178.t002 PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 20 / 51

Principal Component Analysis (PCA) was performed to evaluate environmental gradients underlying the palynological dataset and to identify groups of taxa representing different types of vegetation. Only the most abundant or ecologically relevant taxa were analysed. PCA calcu- lations were performed using the SPSS and software (version 4.0.2) with FactoMineE and Rioja packages for PCA and Cluster Analysis, respectively [100, 101]. Mega and mesoflora Megaflora was initially investigated and documented by using an Olympus SZX 12 stereomi- croscope with DP 70 digital camera. Material with preserved morphological details was studied via Scanning Electron Microscopy (SEM). Specimens were mounted on cleaned aluminium stubs using nail polish, coated with gold and examined with a Hitachi S-3700N Environmental Scanning Electron Microscope at 2 kV, at the National Museum of Prague (Czechia), where the studied material is stored temporarily. Specimens from SJ and AP will be transferred in the future to the Museo Aragone´s de Paleontologı´a (Fundacio´n Conjunto Paleontolo´gico de Teruel-Dino´polis, Spain) as the definitive repository. Amber bioinclusions In the Maestrazgo Basin, amber pieces with bioinclusions have been collected for research pur- poses since 1998. They have been prepared using epoxy resin [102], to improve the visualisa- tion of the bioinclusions and their long-term preservation. The amber pieces from the San Just and Arroyo de la Pascueta outcrops are housed at the Museo Aragone´s de Paleontologı´a (Fun-

dacio´n Conjunto Paleontolo´gico de Teruel-Dino´polis, Teruel), and the ones from the La Hoya outcrop at the Museu de la Universitat de València d’Història Natural (Valencia). Bioinclu- sions have been photographed using a digital camera sCMEX-20 attached to a compound microscope Olympus CX41 with the software ImageFocusAlpha version 1.3.7.12967.20180920 (www.euromex.com) and processed with Photoshop CS6 version 13.0 (www.adobe.com). The list of identified taxa is included in Table 2 and S2 Appendix. Results Palaeobotanical aspects A total of 26 out of 92 samples processed for their palynological content were barren (1 in San Just section, 14 in Cortes de Arenoso section and 11 in the Arroyo de la Pascueta section). Pro- ductive samples showed an overall good recovery and well-preserved palynological content. Except on a few occasions, interpreted as reworking, miospores and dinocysts present a yellow to pale brown colour suggesting low thermal maturation [103]. A total of 367 palynomorph types (S2 Appendix) have been identified, mainly consisting of dinoflagellate cysts (dinocysts, 30 taxa; Fig 7C–7H), bryophyte, lycophyte and pteridophyte spores (186 taxa; Fig 8), gymno- sperms (53 taxa; Fig 7I–7T) and angiosperms (88 taxa; Fig 9). A small number of acantho- morph acritarchs, phycomes of prasinophytes, freshwater algae (Fig 7A and 7B) and linings of foraminifers have also been recorded (S1 Appendix). Unlike the upper Albian–lower Cenomanian successions of the Basque-Cantabrian and

Lusitanian basins [104, 105], dinocysts were only scarcely recorded in the studied assemblages. The most frequently occurring taxa are Cyclonephelium spp., Criboperidinium spp. (Fig 7C), Implestosphaeridium spp., Kiokansium unituberculatum (Fig 7H) and Oligosphaeridium com- plex (Fig 7G). The remaining of the identified marine and freshwater palynomorphs are not numerically significant (S1 Appendix). Terrestrial assemblages are characterised by abundant conifers with Classopollis (mainly Classopollis major) and Inaperturopollenites dubius (Fig 7K PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 21 / 51

Fig 7. Light photomicrographs of selected aquatic palynomorphs and gymnosperm pollen grains. (A) Botryococcus braunii, level AP-34.1. (B) Chomotriletes minor, level AP-34.1. (C) Cribroperidinium sp., level AP-10.1. (D) Ginginodinium cf. evittii, level SJ-2.1. (E) Tenua hystrix, level SJ-2.1. (F) Odontochitina rhakodes, level SJ-3.1. (G) Oligosphaeridium complex, level SJ-2.1. (H) Kiokansium unituberculatum, level AP-13-1. (I) Cycadopites sp., level SJ-3.7. (J) Afropollis jardinus, level CA-5.2. (K) Inaperturopollenites dubius, level SJ-2.1. (L) Uesuguipollenites callosus, level AP- 6. (M) Callialasporites segmentatus, level AP-6. (N) Callialasporites trilobatus, level AP-13.2. (O) Gnetaceaepollenites oreadis, level AP-16.2. (P) Exesipollenites tumulus, level SJ-2.1. (Q) Tetrad of Classopollis major, level SJ-2.3. (R) Parvisaccites radiatus, level AP-4.3. (S) Vitreisporites pallidus, level SJ-2.2. (T) Podocarpidites sp., level AP-25.6. Specimens A–C, H, L–O, R, T are from the Arroyo de la Pascueta section; specimens D–G, I, K, P–Q, S are from the San Just section; the specimen J was found in a sample from the Cortes de Arenoso section. Scale bar equals 20 μm except in S when it equals 10 μm. https://doi.org/10.1371/journal.pone.0282178.g007 PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 22 / 51

Fig 8. Light photomicrographs of selected spores of Bryophyta and Pteridophyta. (A) Reticulosporis gallicus, level CA-21.03. (B) Cyathidites australis, level SJ-2.1. (C) Polypodiisporonites cenomanianus, level CA-35.01. (D) Verrucosisporites sp., level AP-25.6. (E) Concavissimisporites cf. crassatus, level SJ-3.6. (F) Gregussisporites orientalis, level SJ-5.2. (G) Contignisporites cooksoniae, level AP-25.9. (H) Staplinisporites caminus, level AP-25.6. (I) Densoisporites velatus, level CA-2. (J) Foveosporites cf. parviretus, level AP-25.7. (K) Cibotiumspora juriensis, level CA-5.1. (L) Taurocusporites segmentatus, level SJ-3.1. (M–N) Cicatricosisporites proxiradiatus, level CA-21.04, (M) proximal side, (N) distal side. (O–P) Cicatricosisporites potomacensis, level AP-25.7, (O) proximal side, (P) distal side. (Q) Nodosisporites segmentus, sample AP-25.7. (R) Appendicisporites cf. crenimurus, level CA-10.01. (S) Appendicisporites tricornitatus, level AP-4.3. (T) Patellasporites tavaredensis, level SJ-5.2. Specimens A, C, I, K, M–N, R are from Cortes de Arenoso section; specimens B, E–F, L, T are from San Just section; specimens D, G, H, J, O–Q, S are from Arroyo de la Pascueta section. Scale bar equals 20 μm except in H when it equals 10 μm. https://doi.org/10.1371/journal.pone.0282178.g008 PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 23 / 51

Fig 9. Light photomicrographs of selected pollen grains of angiosperms. (A) Asteropollis asteroides, level CA-2. (B) Jusinghipollis cf. ticoensis, level AP-10.2. (C) Clavatipollenites tenellis, level AP-32.3. (D) Stellatopollis cf. barghoornii, level SJ-5.2. (E) Pennipollis reticulatus, level CA-4. (F) Liliacidites cf. clavatus, level CA-14.01. (G) Dichastopollenites sp, level SJ- 3.5. (H–I) Dichastopollenites dunveganensis, (H) level CA-7, (I) level SJ-3.8. (J) Doyleipollenites robbinsiae, level SJ-2.2. (K) Retimonocolpites dividuus, level AP-25.2. (L) Dichastopollenites cf. reticulatus, level AP-25.6. (M) Liliacidites tectatus, sample AP-25.7. (N) Tucanopollis crisopolensis, level CA-3.2. (O) Transitoripollis sp., level AP-25.6. (P) Rousea georgensis, level SJ-3.8. (Q) Phimopollenites pannosus, level SJ-3.7. (R) Tricolpites nemejci, level AP-10.2. (S–T) Dryadopollis vestalis, (S) specimen in polar view, level AP-10.2, (T) specimen in equatorial view, level CA-14-01. (U) Phimopollenites pseudocheros, level AP-34.2. (V–W) Tricolpites cf. maximus, (V) specimen in polar view, level CA-3.1, (W) specimen in equatorial view, level CA-3.1. (X) Hammenia fredericksburgensis, level SJ-3.4. (Y) Echinipollis cenomanensis, level CA- 35.01. (Z) Penetetrapites mollis, level CA-1. (AA) Hammenia fredericksburgensis, level CA-35.01. (AB) Artiopollis praecox, level AP-25.2. (AC) Senectotetradites sp., level AP-10.2. (AD) Senectotetradites grossus, level CA-35.01. Specimens A, E–F, H, N, V–W, Y–AA, AD are from Cortes de Arenoso section; specimens B–C, K–M, O, R–S, U, AB–AC are from Arroyo de

la Pascueta section; specimens D, G, I–J, P–Q, T, X are from San Just section. Scale bar equals 10 μm. https://doi.org/10.1371/journal.pone.0282178.g009 PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 24 / 51

and 7Q), constituting 80% of the total palynomorph sum at some levels. Other frequent gym- nosperms include Araucariacites australis, Exesipollenites tumulus (Fig 7P) and Monosulcites spp. Bisaccate grains (Fig 7R–7T) usually represent a minor component of the assemblages. Fern spores represented the most diversified group of miospores in all the studied sections and occasionally reached dominance, being Cyathidites australis and C. minor (Fig 8B) the most frequently occurring species. Angiosperms, mainly represented by the genera Clavatipollenites (Fig 9C) and Retimonocolpites (Fig 9K), were recorded in significant numbers at specific levels from the CA and AP sections. A low number of tricolpate grains were identified in the three successions (Fig 9P–9W). The PCA presents a variance of 33.8% in the first two dimensions (Fig 10). On the first axis (19.5% of variance), the taxa with negative coordinates were Classopollis spp., Exesipollenites tumulus, Araucariaceae, Patellasporites tavaredensis, Cicatricosisporites spp. and Crassipollis chaloneri. In contrast, Inaperturopollenites dubius, Eucommiidites troedsonii, Peromonolites allenensis, Monosulcites spp., Cyathidites/Deltoidospora, Gleicheniaceae, Clavatipollenites spp., Pennipollis spp., Tucanopollis spp. and the dinocysts presented positive coordinates in the PCA plot. The scattering of samples from AP and SJ along the first axis of the plot suggests the exis- tence of an ecological gradient defining two vegetation types at the extremes. The second axis

(14.3% of variance) discriminates spores in the negative extreme, from wind-transported gym- nosperm pollen grains plus marine protists, in the positive one. While samples from AP and Fig 10. Principal Component Analysis (PCA) performed with the most relevant identified taxa and levels. (A) Classopollis spp. (B) Exesipollenites tumulus. (C) Araucariaceae. (D) Patellasporites tavaredensis. (E) Cicatricosisporites spp. (F) Crassipollis chaloneri. (G) Clavatipollenites spp. (H) Plicifera spp. (I) Cyathidites/Deltoidospora. (J) Gleicheniidites senonicus. (K) Monosulcites spp. (L) Eucommiidites sp. (M) Pennipollis spp. (N) Peromonolites allenensis. (O) Tucanopollis spp. (P) Marine protists. (Q) Inaperturopollenites dubius. https://doi.org/10.1371/journal.pone.0282178.g010 PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 25 / 51

CA are mainly plotted against the negative side of the axis, levels from SJ are mainly restricted to the positive quadrant. Mega- and mesoremains were collected in the three studied sections. Generally, they exhibit good preservation and occur more abundantly in levels with marine influence (i.e., associated with palynological samples containing dinocysts) and without amber (e.g., SJ-2.1, CA-2, AP- 4.2; Figs 3–5). In sharp contrast with the palynofloras, the megafloral assemblages are poorly diverse and dominated by conifers, being Frenelopsis particularly abundant. Angiosperms rep- resent only a minor part of the mesofossil assemblages. The megaremain assemblage of AP has been interpreted as mostly allochthonous due to the range of disarticulation and fragmenta- tion of specimens of Frenelopsis turolensis [106]. The preservation of most of the fossil plants as compressions including well-preserved cuticles and the occurrence of ramified and articu- lated branches of Frenelopsis (Fig 11D), some of them presenting cones attached to their axes, would indicate, however, a parautochthonous production sensu [107]. One of the most abundant taxon is Frenelopsis, which presents in the Maestrazgo Basin a vegetative morphology markedly xeromorphic with highly reduced leaves, thick cuticles and deeply sunken stomatas [37, 106]. Leaves of Eretmophyllum, Mirovia and Dammarites, as well as twigs of Brachyphyllum and Widdringtonites, have been commonly found. San Just section (SJ). The lower palynological interval of this section (SJ-2.1–SJ-3.4; Fig

12 clusters I–II) is characterised by marine palynomorphs and high percentages of Inaperturo- pollenites dubius (Fig 7K) and Classopollis spp. (Fig 7Q). Its upper part (SJ-3.5–SJ-5.3; Fig 12 clusters III–V), corresponding with the amber outcrop, is devoid of dinocysts and reveals increasing abundances of fern spores (Cyathidites/Deltoidospora and Gleicheniaceae) and angiosperms (mainly Pennipollis). A renewed increase of I. dubius and Classopollis, this time accompanied by numerous Anemiaceae and spores of uncertain botanical affinities (Patellas- porites tavaredensis, Fig 8T), is characterising the uppermost interval of the succession (Fig 12, cluster V). The megaflora collected in SJ (Fig 3) is the most diversified among the studied outcrops from the Maestrazgo Basin. Frenelopsis justae is numerically prevailing (Fig 11E). This species shows robust axes consisting of nodes and internodes formed by three unified leaves [37]. In addition, scarce remains of Brachyphyllum cf. obesum (Fig 11I), Eretmophyllum sp., Dammar- ites sp. and Widdringtonites sp. (Fig 11J) have been found. Several small fruits of angiosperms were found as mesofossils. They mainly include fruits of Montsechia-type (Fig 11F) and seg- mented fruitlets of cf. Serialis sp. (Fig 11H). Cortes de Arenoso section (CA). The lower palynological interval of CA (CA-1–CA- 14.01; Fig 13, clusters I–IV) is characterised by fluctuating abundances of Classopollis and Ina- perturopollenites dubius. The levels with the highest numbers of Classopollis usually include

Fig 11. Selected plant macro and mesofossils from the El Maestrazgo Basin. (A) Leaf of Nehvizdya (= Eretmophyllum) penalveri, No. K 2788. (B) Shoot of Frenelopsis turolensis, No. K 2786. (C) Fragment of leaf of Dammarites cf. albens, No. K 2789. (D) Ramified shoot of Frenelopsis cf. turolensis, No. K 2797. (E) Shoot of Frenelopsis justae, No. K 2806. (F) Fruit fragment of Montsechia-type with orthotropous seed (Spermatites), No. K 2795. (G) Fragments of leaves of Mirovia gothanii, Nos. K 2807–2811. (H) Angiosperm fruit of cf. Serialis sp., SEM image, No. K 2803. (I) Shoot fragment of Brachyphyllum cf. obesum, SEM image, No. K 2804. (J) Shoot fragment of Widdringtonites sp., SEM image, No. K 2805. (K) Basal part of a leaf of Nehvizdya (= Eretmophyllum) penalveri, No. K 2794. Specimens A–C, G are from Arroyo de la Pascueta section; specimens D, F, K are from Cortes de Arenoso section; specimens E, H–J are from San Just section. Scale bars: A–D = 10 mm; E, G, K = 5 mm; F, H–J = 1 mm. https://doi.org/10.1371/journal.pone.0282178.g011 PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 27 / 51

Arroyo de la Pascueta outcrop (AP). In the palynological succession, Cyathidites/Deltoi- dospora and Classopollis exhibit antagonistic percentage fluctuations. Cheirolepidiaceae char- acterised the lower part of the succession (Fig 14, clusters I and III). Conversely, although defining a pattern slightly less pronounced, Cyathidites/Deltoidospora are more abundant in the upper part of the section (Fig 14, cluster V and cluster VI), and occur in assemblage devoid of marine palynomorphs. The two mentioned intervals are separated by a level with abundant angiosperms and common Anemiaceae (cluster IV). The amber-bearing stratum, typified by Cyathidites/Deltoidospora, is restricted to the uppermost part of the succession (cluster VI). Although not found in abundance, angiosperms are diverse in the amber-bearing bed and include various species associated with Chloranthaceae/Ceratophyllaceae, Laurales and eudi- cots (i.e., Fig 9U). Mega and mesofossil plants from AP show a balanced representation of remains with thick cuticles. The megaflora includes narrowly elongated leaves of Mirovia gothanii (Fig 11C), broadly obtuse entire-margined leaves of the ginkgophyte Nehvizdya (= Eretmophyllum) penalveri (Fig 11A), shoots of the Cheirolepidiaceae Frenelopsis turolensis (Fig 11B) and elon- gated leaves of Dammarites cf. albens (Fig 11C). While elongate seeds of the Spermatites type Fig 12. Detailed palynological chart for San Just section (SJ). Blue colour: marine palynomorphs; yellow colour: spores of ferns and allied; green colour: Gymnosperm pollen; red colour: Angiosperm pollen.

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are common, the anecdotic presence of megaspores (two specimens) is noteworthy as it may reflect occasional floodings or high-water table allowing the development of wetlands. Insects and amber The Cretaceous amber from the Maestrazgo Basin has provided an extensive arthropod record as bioinclusions from the San Just, Arroyo de la Pascueta and La Hoya outcrops (Table 2). San Just represents one of the most outstanding Cretaceous amber-bearing outcrops worldwide with diverse and abundant bioinclusions. Its entomological content has been the subject of extensive research over the last decades (i.e. [108, 109] and references therein). The arthropod assemblages from San Just include arachnids (Pseudoscorpions, Acari and Araneae; Fig 4C), non-insect hexapods (Collembola) and up to eleven orders of insects (Archaeognatha, Orthoptera, Blattodea, Mantodea, Psocodea, Thysanoptera, Hemiptera, Neu- roptera, Coleoptera [Fig 4D], Hymenoptera [Fig 4E and 4F] and Diptera), together with a high number of arthropod ichnofossils such as spiderwebs and coprolites. The site is also the type locality of 22 arthropod taxa to date, most of them insects (Table 2). Hymenoptera (mainly pla- tygastroids [Fig 4E]) and Diptera are the most abundant insects. Apart from arthropods, the amber from SJ also includes dinosaur feathers (Fig 4F), although the outcrop lacks faunal megaremains. Conversely, the bioinclusions from the Arroyo de la Pascueta and La Hoya

Fig 13. Detailed palynological chart for Cortes de Arenoso section (CA). Blue colour: marine palynomorphs; yellow colour: spores of ferns and allied; green colour: Gymnosperm pollen; red colour: Angiosperm pollen. 1. spike due to the abundance of Tucanopollis spp. 2. spike due to the abundance of “Liliacidites” minutus. https://doi.org/10.1371/journal.pone.0282178.g013 PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 29 / 51

outcrops are poorly known and only few amber pieces have been described so far. Addition- ally, new material collected from these two outcrops is currently being prepared and new find- ings are expected. Hymenoptera, including one new species [110], Hemiptera and Diptera have been reported from Arroyo de la Pascueta while Diptera and Blattodea have been recorded in La Hoya. Collectivelly, the amber from the three outcrops has delivered 40 arthro- pod taxa, presenting higher or similar level of diversity than other mid-Cretaceous amber- bearing sites of Iberia such as Peñacerrada and El Soplao (northern Spain). Discussion Palynomorph-based biostratigraphy In the AP section, the presence of Tricolporoidites (Fig 6), Artiopollis praecox (Fig 9AB), Drya- dopollis vestalis (Fig 9S), Gnetaceaepollenites oreadis (Fig 7O), Liliacidites tectatus (Fig 9M), Rousea miculipollis, Senectotetradites spp. (Fig 9AC) and Tricolpites nemejci (Fig 9R) indicates a late Albian age (Fig 15) since none of these taxa have been identified in older strata [78, 93, 104, 111–113]. This age is significantly younger than the one inferred by [29] based on a long- distance correlation with palaeontological data from the neighbouring Morella Sub-basin. The age of the SJ section can be independently dated as late Albian or younger based on the record of Odontochitina rhakodes (Fig 7F). This age is further supported by the presence of Fig 14. Detailed palynological chart for Arroyo de la Pascueta section (AP). Blue colour: marine palynomorphs; yellow colour: spores of ferns and allied;

green colour: Gymnosperm pollen; red colour: Angiosperm pollen. https://doi.org/10.1371/journal.pone.0282178.g014 PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 30 / 51

Tricolpites nemejci and Senectotetradites sp. [93, 104, 111, 114–116]. This age inference updates and refines the previously proposed middle to early late Albian age, which was based on a more limited palynological data set [32]. The presence of Cicatricosisporites apicanalis, Con- tignisporites cooksoniae (Fig 8G), Gregussisporites orientalis (Fig 8F) and Nevesisporites dailyi (Fig 15) in both SJ and AP, also allows defining an intra late Albian upper limit for the age interval as none of the taxa have been recorded in strata of different age in Europe [31, 113, 117]. In addition to the biostratigraphic markers already present in SJ and AP, the lower part of the CA section also includes Reticulosporis gallicus (Fig 8A), a taxon only recorded in France in strata independently dated as late Albian [90, 118], and Microfoveolatosporis pseudoreticula- tus and Cupuliferoidaepollenites parvulus (Fig 15), both taxa presenting the oldest occurrence in strata of that age in North America and Western Europe [105, 112, 119]. A slightly younger early Cenomanian could be considered for the uppermost part of the CA section that corre- sponds to the La Hoya amber outcrop (CA-35.1–CA-35.3; Fig 5) as Echinipollis cenomanensis (Fig 9Y), Polypodiisporonites cenomanianus (Fig 8C) and Senectotetradites grossus (Fig 9AB) recorded therein have not been reported from older strata [111, 120–122]. The CA record Fig 15. Composite range chart showing selected species of biostratigraphic interest identified in the Lower Cretaceous sections from the Maestrazgo Basin. L = lower; M = middle; U = upper.

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could equally represent a range extension down into the late Albian for these taxa, which remain not widely distributed (i.e., rare species). The updated new age for the sections has important implications for the local stratigraphy as it allows us to confidently attribute them to the Utrillas Group [61], while previous studies considered them as part of the Escucha Formation [28, 29, 31, 32, 43]. The new biostrati- graphic inferences are also supported by previous results obtained for the Utrillas Group in the Maestrazgo Basin based on indirect correlations [47, 50], palynological [123, 124] and faunal data (ostracods) [61]. From a palaeoenvironmental perspective, the biostratigraphic markers recorded in the three studied sections suggest that resin production/deposition in the Maestrazgo Basin was either coeval or close in time. In that regard, the amber levels might represent a biostrati- graphic event of regional significance. The marine carbonates of the Mosqueruela Formation [50], immediately overlying the amber-bearing strata at CA and AP, represent the regional expression of the flooding event described elsewhere in uppermost Albian strata (Mortoni- ceras rostratum/M. perinflatum ammonite zones) [125]. In eastern Iberia, this transgressive pulse led to the development of widespread marine platforms across the Mesozoic intraplate basins [48, 49]. Environments and vegetation The record of highly variable fossil remains and amber with biological inclusions suggest the presence of an ecosystem structurally framed by multi-tiered vegetation [14, 28, 34–36] sus-

taining complex arthropod communities (Table 2). Terrestrial ecosystems of such complexity might seem surprising for deposits associated with a desert system. However, there are no rea- sons to believe that arid and semi-arid ecosystems from the Cretaceous were less complex than their modern counterparts, such as the Namib and Kalahari deserts [126, 127]. The presence of palynological assemblages of varying composition in marine-influenced intervals is other- wise expected, given the taphonomic, palynological and palaeobotanical constraints [14, 128– 131], and indicates the existence of mosaic vegetation types in the area with conifer-forests along the Tethyan coasts (Fig 16). The vegetation of this region was strongly conditioned by both the influence of the sea and the desert and other factors such as palaeowildfires, as the abundant charcoal occurrence in the Spanish Cretaceous amber-bearing deposits indicates [40, 132]. A picture of the regional vegetation: Cheirolepidiaceae-Cupressaceae communities The significant representation of Classopollis and Inaperturopollenites dubius in the studied assemblages reflects the importance of the families Cheirolepidiaceae and Cupressaceae in the vegetation. The increased numbers of these two taxa observed in assemblages with marine palynomorphs (Figs 12–14) and the correlation between I. dubius and dinocysts in the PCA (Fig 10), representing the most distal settings indicate that their pollen-producers were wide- spread regionally, probably forming a significant part of the hinterland vegetation.

Cheirolepidiaceae was one of the most widespread conifer families among the Mesozoic vegetation. They colonised a wide range of environments ranging from low to high latitudes in both hemispheres [8, 133–135] and commonly assumed a dominant position in the vegetation from the Early Jurassic to the mid-Cretaceous [134, 136], especially in some low latitude set- tings associated with evaporites [8, 137]. While historically linked to arid environments and/or salty ones [79, 138–140], recent studies suggest broader ecological preferences for, at least, some representatives of the family [134, 141, 142]. PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 32 / 51

Inaperturopollenites dubius-producers are thought to belong to Cupressaceae, a conifer family with species widely distributed in present-day ecosystems of the Northern Hemisphere [143]. While some modern forms, such as the bald cypress and the Chinese swamp cypress, mostly belonging to the Glyptostrobus–Taxodium–Cryptomeria clade (Taxodioideae), exhibit a marked ecological affinity toward wet environments [144], a significant number of Cupressa- ceae species prefer well-drained habitats [143]. Distinguishing moisture-loving forms from Fig 16. Schematic model of the late Albian fore-erg setting, including the types of vegetation and their distribution in the different sub-environments. https://doi.org/10.1371/journal.pone.0282178.g016 PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 33 / 51

plants preferring well-drained substrates based solely on palynological evidence is complex due to the similarity of the pollen produced by both groups of plants. By analogy with modern pollen of Taxodium, fossil grains presenting a papilla such as Taxodiaceaepollenites are assumed to have been produced by trees preferring saturated soils and wetlands subjected to periodic floodings. Taxodioideae pollen is frequent in Cenozoic deposits and has also been found in high numbers in Cretaceous deposits from high latitude locations [135, 145]. These pollen grains have also been recorded in mid-latitude assemblages associated with spores pro- duced by water ferns [90]. However, they are scarcely represented in the successions from the Maestrazgo Basin (S1 Appendix). Perinopollenites halonatus, pollen with two distinct exine layers related to Taxodioideae [77], neither reaches a significant representation in the studied sections (S1 Appendix). Therefore, most Cupressaceae pollen recovered is assumed to have been produced by trees or shrubs that preferred well-drained substrates. The presence of Widdringtonites sp. in SJ (Fig 11J) would support the presence of Cupressa- ceae linked to well-drained substrates as the specimens assigned to this taxon present well- developed Florin rings in their stomata, reflecting xerophytic conditions. Interestingly, the genus Widdringtonites has been related to the extant Widdringtonia, which presents species adapted to very hot, dry summers and recurring wildfires in South Africa [146].

If an arid/semi-arid environment can be inferred from the large number of Cupressaceae pollen and Classopollis as well as regional sedimentology, the presence of localised thickets of Taxodioideae colonising the wettest parts of the landscape (i.e., wet interdunes, see below), while unlikely, cannot be discarded. The palynofloral data presented here and indicating the prevalence of Cupressaceae in local vegetation confirm the results obtained from the fossil wood dataset for the Albian (T2 time bin in [5]). Cheirolepidiaceae and Cupressaceae are often the co-dominant families in the studied paly- nofloras. Fluctuating frequencies of these taxa could, however, reflect drastic changes in vege- tation which are difficult to ascertain. While Cheirolepidiaceae usually dominate in AP and CA (Figs 13 and 14), Cupressaceae remains are more abundant in SJ palynofloras and the amber-bearing levels of AP and CA (Figs 12–14). These two taxa have previously recorded similar dominance of the palynofloras in numerous mid-latitude Jurassic and Cretaceous deposits [25, 90, 104, 136, 142]. Present-day growth habits of Cupressaceae, as well as whole plant reconstructions of fossil representatives of Cheirolepidiaceae, suggest that these plants were dominantly, if not exclusively, arborescent during the Cretaceous. The corresponding shrubs and trees might have represented the sub-dominant and dominant components of open vegetation (savanna–woodlands; Fig 16) similar to those observed in modern arid to semi-arid climates [143]. This conifer-dominated plant community was very different from

the Jurassic and Cretaceous fern savanna biome characterised by the families Anemiaceae and Matoniaceae [75, 147–151] which might have also grown on dunes. The diversity of angiosperm pollen types and their varying abundance pattern (S2 Appen- dix) suggest that producers had distinct ecophysiologies allowing them to colonise not only moister habitats and, probably, water-logged, if not aquatic, environments [12, 35, 152–154] but also well-drained and/or dry edaphic settings. Originally associated with moist environ- ments [155], angiosperms may have already radiated into more arid settings by the late Albian, as their presence in deposits associated with the desert system could suggest [14]. It is likely that the representation of flowering plants in pre-Cenomanian ecosystems is underestimated owing to morpho-functional and taphonomic biases (i.e., low production and dispersal of entomophilous pollen [156]). While the exact phenology of early angiosperms is poorly under- stood, the existence of short life cycles allowing flash flowering [152, 157] could explain the localised high abundance of this pollen. PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 34 / 51

Diversified local vegetation. • Coastal settings with Araucariaceae Pollen of Araucariaceae is consistently recorded in the palynofloras of the three studied out- crops (S1 Appendix), albeit in low abundance (<5% of the total miospores). However, the presence and relative abundance of these conifers in the local vegetation may be underesti- mated since both modern [158, 159] and fossil Araucariaceae pollen [136] are believed to pres- ent low dispersal capabilities. Several researchers [158] noted that modern palynological assemblages collected from Kauri (Agathis australis) forests only include relatively low per- centages (<5–10%) of Araucariaceae pollen. Palaeobotanical and geochemical evidence suggests a clear Araucariacean affinity for the late Albian amber recovered in the Basque-Cantabrian Basin [90, 160]. However, a geochemi- cal analysis of the amber from the San Just outcrop seems to support an araucariacean-cheiro- lepidiacean affinity [161]. The ubiquity of Araucariaceae in palynological assemblages and the record of presumed specialised insect pollen feeders (nemonychid beetles) in amber inclusions from the San Just outcrop [162] would represent additional circumstantial evidence linking Araucariaceae to the resin production. The precise role of Araucariaceae in Cretaceous vegetation and the structure of the forest types they formed still need to be ascertained, given the characteristics of the habitats colonised by modern forms. Apart from isolated occurrences in savanna-like vegetation growing on

mafic-ultramafic soils of New Caledonia (i.e., Agathis ovata; [163]), modern araucarians are often part of evergreen rainforests where they act as emergents (Agathis microstachya in Aus- tralia, A. ovata and Araucaria schmidii in New Caledonia, Agathis australis in New Zealand), or constitute small thickets and discrete stands in low altitude forests of New Zealand (A. aus- tralis) and Australia (Araucaria heterophylla) [164, 165]. Species distributed in low-fertility soils, ridgetops, and steep slopes also reflect probable competitive exclusion by angiosperms, i.e., a derived, relictual distribution (a realised niche much more restricted than a potential niche sensu [166]). Some Recent Araucariaceae, such as Araucaria columnaris, A. luxurians and A. nemorosa are coastal trees able to withstand the influence of salt wind [74, 167], and this ecological habit was probably already exhibited by representatives of the family during the Mesozoic. The presence of Microphorites utrillensis (Dolichopodidae: Microphorinae) in amber from the SJ outcrop [168] would support a sandy coastal habitat for Araucariacites- producers, as living microphorines are known to inhabit this type of habitats [169] (Figs 16 and 17). It is interesting to mention the association of modern Araucariaceae with other conifers, mainly Podocarpaceae, but also Cupressaceae in the evergreen forests of New Zealand [165]. Associations among conifers, probably fortuitous within the context of modern vegetation dominated by angiosperms, should have been more frequent during most of the Mesozoic.

The association of Araucariaceae with Cheirolepidiaceae and Exesipollenites tumulus in PCA (Fig 10) would support the existence of mixed-gymnosperm communities in the area. The spa- tial proximity of conifers from distinct families is also suggested by the megaremain assem- blages where Brachyphyllum cf. obesum (Fig 11I), a species that indicates mesophytic to xerophytic conditions attributed to Araucariaceae, has been found together with the Cheirole- pidiaceae Frenelopsis justae (Fig 11E). Two types of plant formations involving Araucariaceae can be hypothesised: (i) thickets of limited lateral extent involving one or a few species of Araucariaceae as the dominant ele- ments. This hypothesis, still not supported by in situ palaeobotanical evidence, could explain the low dispersion of pollen (i.e., underrepresentation in palynological assemblages), the PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 35 / 51

resulting canopy acting as a barrier to pollen transport in a fashion analogous to modern stands of Araucaria columnaris in New Caledonia and A. angustifolia in Brazil [159] and a pre- dicted poor species richness for the stand [170]; (ii) mixed conifer communities with Cupres- saceae, Podocarpaceae or/and Cheirolepidiaceae conifers constituting single-tiered or two- tiered woodlands, the latter with emergent araucarians. It would be reasonable to assume the existence of a transition from woodlands to more open, savanna-like, vegetation structures constrained by local edaphic conditions and the presumed arid to semi-arid conditions at a regional scale. The presence of Araucariaceae in hinterland vegetation could also be regarded as plausible, but it is likely not to have produced the recovered palynological signal given the low transport capacity of the araucarian pollen. Conversely, the ubiquity of Cheirolepidiaceae in pollen assemblages suggests a more efficient dispersal and that their contribution to the mixed conifer association described above might not have been that significant. • Salt marshes and swamps Mega and mesofloral assemblages from the Maestrazgo Basin are characterised by abun- dant Frenelopsis. While the genus has been associated with brackish coastal marshes when found in the mid-Cretaceous of Europe [106, 171, 172], it seems likely that the constituting species exhibited more diverse palaeoenvironmental affinities. In the three outcrops, the genus is represented by F. turolensis (Fig 11B), and/or F. justae (Fig 11E). While F. turolensis has

Fig 17. Conceptual palaeoartistic reconstruction of the late Albian wet interdunes in the fore-erg from the Maestrazgo Basin (eastern Spain). This environment developed near the Tethys, where (A) coastal forests with Araucariaceae grew. The riparian places of the wet interdunes (oasis) (B) were covered by fern and angiosperm vegetation. (C) Fresh water/brackish aquatic angiosperms inhabited wet interdunes. (D–E) Xerophytic plant community involving Cheirolepidiaceae, Cupressaceae (D) and other gymnosperms groups such as Bennettitales, Cycadales (Monosulcites/Cycadopites-producers) and Gnetales (Equisetosporites-producers) (E) that integrated arid woodlands. https://doi.org/10.1371/journal.pone.0282178.g017 PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 36 / 51

been reconstructed as a small ligneous plant or shrub growing in coastal salt-influenced envi- ronments [106, 173], F. justae [37] suggests more mesic settings [174, 175]. The association of F. turolensis with Nehvizdya (= Eretmophyllum) penalveri [29] (Fig 11A and 11K), a ginkgoalean previously interpreted as halophytic [83, 176], supports colonisation of salt-influenced settings (Fig 16). This type of ginkgoalean constituted coastal vegetation associated with Cheirolepidiaceae [177]. These communities could thrive in supratidal coastal areas [174, 178]. A similar inference has also been proposed by [28, 29] based on specimens collected in the AP succession. If not taphonomically induced, the low diversity of Frenelopsis- Eretmophyllum could be interpreted as supporting evidence of stressful ecological conditions [179]. Similarly, the complex nature of the fruit of Montsechia-type with Spermatites seeds inside recorded in SJ and CA (Fig 11F) could reflect near-shore deposits as these seeds (Spermatites) have been recorded in physical association with Pseudoasterophyllites cretaceous, a Cenoma- nian angiosperm described in tidally-influenced deposits from Bohemia [98, 180, 181]. While still debated, Montsechia has been considered by some authors as a halophytic plant growing in periodically desiccated coastal marshes [182]. Irrespective of the specific ecological require- ments of Montsechia, the existence of halophytic/xerophytic vegetation formation, including angiosperms is supported by the presence of Tucanopollis. While this pollen has been recorded

in European mid-Cretaceous deposits [183–185], it constitutes an essential element of assem- blages characterising arid settings of central [186, 187] and NE Gondwana [16] and has been observed in stamens of P. cretaceous [98]. The high proportion of Tucanopollis in the lower part of the CA succession (representing the bulk of the ‘other angiosperms’ in level CA-3.2; Fig 13, cluster II; S1 Appendix) is assumed to reflect the proximity of a halophytic/xerophytic plant community also involving Cupressaceae (Inaperturopollenites dubius-producers), Ertma- nithecales and Cheirolepidiaceae (F. turolensis) and probably Monosulcites-producers of uncertain botanical affinities (Fig 17; S1 Appendix). This plant association would characterise coastal, salt-influenced, episodically (but not periodically) flooded areas. The existence of man- groves (i.e., vegetation colonising habitats under periodic marine water influence) is possible, although currently not supported by anatomical evidence. • Wet interdunes and wetlands The presence of freshwater bodies in the study area during the late Albian to early Cenoma- nian has been inferred from sedimentological and palaeobotanical evidence [188, 189]. The existence of habitats with fluctuating humidity or with high moisture is also indicated in San Just by amber inclusions, including the hemipteran Iberofoveopsis miguelesi [44], as well as the presence of edaphic fauna linked to the litter degradation and soils [190–192] (Table 2). The high proportions of fern spores in palynological assemblages devoid of marine palyno-

morphs and characterised by low numbers of gymnosperms (see Fig 12 clusters III and IV; Fig 13 cluster IV; Fig 14 clusters II and VI) also indicate that freshwater was available, at least dur- ing part of the year, as spore producers need free water to complete their biological cycle. A series of fern spikes, mainly consisting of abundant spores with smooth exine attributed to Cyathidites and Deltoidospora, are observed in the three sections. The palynological signal corresponding to these spikes is challeging to interpret. Similar spikes have been recorded after the end-Permian, Triassic, and Cretaceous events [193–195] and interpreted as fern-dominated pioneer vegetation characterising the early stage of recov- ery after the disruption of some terrestrial ecosystems. If the existence of a global adverse palaeoenvironmental event during the late Albian can be discarded on available evidence, the fern spikes observed in the three sections could still be interpreted as riparian early succes- sional developments [196] in the present context. Modern arid ecosystems are fragile and PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 37 / 51

prone to significant vegetation shifts when disturbances such as draughts occur [197, 198], and similar disruptions of vegetation could well have happened in similar past environments. A recovery vegetation reduced to pioneer ferns, and conifers and other resilient floral elements remains a plausible ecological scenario. It is interesting to highlight that palynofloras charac- terised by a high proportion of Cyathidites and Deltoidospora (94% of the total miospore con- tent) have already been recorded in other deposits associated with the Iberian Aptian–Albian desert system in SW Spain (Oliete Sub-basin) [2, 199]. If the palaeoenvironmental interpreta- tion presented here is valid, the recurring fern spikes suggest that episodes of accentuated draught were a common feature during the mid-Cretaceous at mid-latitude locations and that the vegetation responded to palaeoenvironmental disturbance in a similar way. The high con- tent in spores of palynofloras reflecting arid and disturbed vegetation may appear counter- intuitive, but a significant number of modern spore-producers can withstand extended periods of drought, present a high tolerance to environmental stress or simply thrive in edaphically low-nutrient habitats [200–202]. This adaptability has also been inferred for several Palaeozoic and Mesozoic spore-producers (e.g. Pennsylvanian sphenophytes, Jurassic Gleicheniaceae, Late Cretaceous Matoniaceae [73, 151]), which are often presented as stress-tolerators. Peri- odic ecological disturbances are also thought to have been favoured by early angiosperms

[155]. Besides Tucanopollis-producers, angiosperms colonising the Iberian Desert System included representatives of the ANITA grade, Chloranthales of diverse affinities (Clavatipolle- nites, Transitoripollis, etc.), probable monocots (Monocolpopollenites, Retimonocolpites) and eudicots (Table 1). These angiosperms were probably well integrated into the landscape, form- ing part of multiple vegetation types, including terrestrial wetlands (wet interdunes, oases sensu [189]) and freshwater aquatic ecosystems as helophytes and/or floating elements [4, 34, 36] (Fig 17). The occurrence of a high abundance of “Liliacidites” minutus in CA (Fig 13 clusters IV) could also reflect advanced stages of ecological succession, involving colonisation of settings dominated by ferns. This pollen type, currently related to Laurales [203], could have been pro- duced by angiosperms of broad ecological tolerances similar to the one exhibited by Eucalypto- laurus deprei, a representative of the family described from the Cenomanian of France [176, 204]. The high number of spores attributed to Anemiaceae (i.e., Cicatricosisporites) in the studied assemblages indicates that ferns were probably still the dominant floral elements in wetlands and maybe in some arid places. Early Cretaceous representatives of the family have been previ- ously associated with swamps, lacustrine, and savanna environments [75, 150, 205], and a sim- ilar environment can be assumed for the Iberian Anemiaceae, along with another spore (Patellasporites tavaredensis) and pollen (Crassipollis chaloneri) producers (Fig 10). The

inferred vegetation should have been of low stature and have surrounded ephemerous or semi-permanent water bodies developed in interdunes, maybe as part of the understory of open woodlands [75, 188, 206]. • Hinterland vegetation The presence of bisaccate grains in assemblages otherwise characterised by marine palyno- morphs suggests that a fourth type of vegetation involving conifers could have constituted a more distant vegetation type. Bisaccate pollen grains present a high dispersal capability, and their higher representation in distal, marine settings is often interpreted to reflect a tapho- nomic (i.e., biostratinomic) bias [207, 208], known as Neves Effect in Palaeozoic floras [209]. In the present context, the higher abundance of bisaccate grains in marine assemblages could represent hinterland vegetation involving Pinaceae and/or Podocarpaceae that would have col- onised topographically higher settings of the Iberian Massif [14]. PLOS ONE Late Albian desert biota from Iberia PLOS ONE | https://doi.org/10.1371/journal.pone.0282178 March 2, 2023 38 / 51