Taxonomic review of Ceratozamia (Zamiaceae) in the Sierra Madre Oriental, Mexico

Abstract The genus Ceratozamia is revised for the Sierra Madre Oriental in Mexico. This region is one of the biogeographic areas with the greatest diversity of species in this genus. These species are highly variable morphologically and this variability has led to a complex taxonomic history with many synonyms, particularly with reference to C. mexicana. We present a comprehensive taxonomic revision with history of nomenclature and the morphology, relationships, distribution and use of these species. We also introduce a key for their identification, descriptions, full synonymy, nomenclatural notes, etymologies and neotypes as well as taxonomic comments describing relevant taxonomic changes. We recognise fourteen species in this biogeographic province: C. brevifrons, C. chamberlainii, C. decumbens, C. delucana, C. fuscoviridis, C. hildae, C. kuesteriana, C. latifolia, C. mexicana, C. morettii, C. sabatoi, C. tenuis, C. totonacorum and C. zaragozae. This study provides a foundation for future taxonomic work in Neotropical species of Ceratozamia.


Introduction
Ceratozamia Brongn. is one of the eight genera in the most diverse cycad family, Zamiaceae (order Cycadales). This genus is endemic to Mega-Mexico (sensu Rzedowski 1991), ranging from Mexico to Honduras. Mexico is considered the centre of diversity for Ceratozamia, given that 30 out of 31 recognised species occur in its territory (Vovides et al. 2004a;Nicolalde-Morejón et al. 2014), 90 percent of which are endemic (Calonje et al. 2013(Calonje et al. -2018. Diversity for this genus is particularly prevalent along the Sierra Madre Oriental and Southwest Mexico, especially in the political states of Veracruz, Oaxaca, Chiapas and Tabasco. Ceratozamia plants mainly inhabit high-elevation cloud forests ("bosque mesófilo de montaña"), pine-oak forests and relatively elevated sectors of evergreen forest in a narrow but continuous distribution (Moretti et al. 1980;Vovides et al. 2004b).
Ecologically, Ceratozamia requires conditions of abundant humidity to ensure proper development of immature embryos because the seeds are usually released from ovulate strobili approximately one year before germination (Norstog and Nicholls 1997). Considering the deforestation rates in Mexico and historical patterns of botanical collection, Ceratozamia is amongst the most threatened plant groups in the country and the genus is placed in the IUCN Red List (IUCN 2016), listed on CITES Appendix 1 and listed in the 'Norma Oficial Mexicana NOM-059-SEMAR- NAT-2010' (SEMARNAT 2010Donaldson 2003). The latter is an official document issued by Mexican authorities listing protection categories under which diverse taxonomic groups should be placed.
Species of Ceratozamia are similar in morphology, particularly with regards to vegetative characters' states, making taxonomic identification problematic. The most recent species-level taxonomic treatment of the genus is over eight decades old (Schuster 1932). Since then, researchers have focused on the study of the taxonomy of individual species and/or species complexes (Vovides et al. 2003;Osborne et al. 2006;Pérez-Farrera et al. 2009;Vovides et al. 2016).
In recent taxonomic works, quantitative morphological characters such as plant size and leaflet width have been commonly used as a basis for the identification and description of new species because they exhibit variability across the genus (Vovides et al. 2004a, c;Whitelock 2004). However, other contemporary studies have shown that these characters can be highly variable between and amongst populations, which limits species diagnosis (Martínez-Domínguez et al. 2016;2017a, b, c). Other characters commonly used are the direction, form and texture of leaflets and the formation of prickles (Pérez-Farrera et al. 2001;Vovides et al. 2003). However, analysis of morphological patterns for these characters along the distribution range of Ceratozamia indicates that leaflet shape is highly similar in many species ).
Ceratozamia was first described by Brongniart (1846) based on the sole species C. mexicana, which was described from wild specimens later cultivated at the Natural History Museum of Paris. Later, Miquel (1847Miquel ( , 1848 described five new species -namely, C. brevifrons, C. intermedia, C. latifolia, C. longifolia and C. robusta. In 1849, this Dutch botanist reduced C. intermedia as a variety of C. longifolia (var. minor Miq.). Later on, Regel (1857a, b) recognised only C. mexicana and C. robusta, along with a new species, C. kuesteriana Regel. All other Miquel names were placed in synonymy within C. mexicana. Miquel (1861) recognised all species listed in his 1849 work plus Ceratozamia kuesteriana and subsequently synonymised all of these names with C. mexicana (Miquel 1868(Miquel , 1869a. Regel (1876a, b) conserved his 1857 system, which recognised C. robusta. The two most recent taxonomic treatments of Ceratozamia, written by Thiselton-Dyer (1884) and Schuster (1932), are highly contrasting in the recognition of species and infraspecific categories. The first author recognises four species and the latter only two species, placing the rest of older names as varieties. Specimen scarcity and lack of nomenclatural types seem to have been largely responsible for the nomenclatural instability in Ceratozamia. This issue was clarified by .
Given the considerable increase in activities dedicated to the exploration of cycad diversity in Mexico (the target of the main taxonomic interest for the last 40 years) and the unstable taxonomy and nomenclature in Ceratozamia, we present a taxonomic clarification of the Ceratozamia species found along the Sierra Madre Oriental (SMO) from North Tamaulipas to North Oaxaca, which represent areas of endemism for this biogeographic province.

Materials and methods
Species-level circumscription is based on results of a phylogenetic analyses performed for Ceratozamia species that included quantitative and qualitative morphological data, molecular evidence, as a well as analyses of herbarium specimens (Martínez-Domínguez 2018). We present the taxonomic treatment for a part of the genus, corresponding to a well-delimited biogeographic region (Fig. 1). We examined herbarium specimens deposited in the following herbaria: CHAPA, CIB, ENCB, FCME, FTG, GH, HEM, IEB, K, LE, LSU, MB, MICH, MEXU, MO, NY, P, SERO, SLPM, U, UAT, US, XAL and XALU. This information was complemented by a review of the original taxonomic accounts for the relevant Ceratozamia species and additional data collected during fieldwork to evaluate diagnostic characters. The corresponding populations were monitored during four years (2014)(2015)(2016)(2017) to evaluate reproductive structure characters, as well as some ontogenetic stages in these characters. In all cases, nomenclatural types were reviewed by all authors or at least one of them.

Results
Vegetative morphology. Stems of the species here studied are epigeous and erect. They become decumbent with age and may develop two or more apices, with the exception of Ceratozamia latifolia, C. hildae, C. kuesteriana and C. zaragozae, which have semi-epigeous stems. Ceratozamia species can produce 4 to 15 leaves per year and each apex can carry up to 100 leaves. C. hildae and C. latifolia are exceptional in this respect because they only produce 1 to 3 leaves per year. Leaves of most species reach up to 2.80 m long. The longest leaves are present in C. mexicana, C. hildae and C. tenuis, whereas the shortest can be observed in C. latifolia and C. zaragozae. New leaf colour can vary from light green to yellowish, as in C. brevifrons, to dark green (blackish) as in C. tenuis or reddish-brown as in C. chamberlainii. Leaf colour can be persistent in some species and remain at the leaf base, margins, nerves and/or undersides of leaflets and the leaf rachis and/or petiole as in C. latifolia, C. chamberlainii, C. kuesteriana and C. fuscoviridis. In other species, leaf colouration can disappear at maturity as in C. totonacorum. At emergence, the leaf petiole and rachis have abundant trichomes of two types: brown for species distributed in Central or South Sierra Madre Oriental with the exception of C. decumbens and white for species distributed in North SMO (Fig. 2).
In most species, presence of prickles in the petiole and rachis is highly variable intraspecifically. Prickles can be long as in C. tenuis where they are up to 0.6 cm long or very short as in C. sabatoi of approximately 0.1 cm. However, the shape of prickles is a constant character within populations. Two forms have been identified: 1) thin and easily detached and 2) robust, wide and hard to separate from the plant.
Leaflet form can be lanceolate, oblong or linear. This character can be polymorphic within and between populations as in Ceratozamia delucana (Fig. 3). Leaflet width can vary from narrow, 0.4-0.7 cm in C. zaragozae (Fig. 4), to broad, up to 5 cm as in C. latifolia and C. decumbens. Other characters with taxonomic value are leaflet symmetry and texture. However, these characters can be polymorphic as in C. delucana. Leaflet texture is generally papyraceous as in C. latifolia or coriaceous as in C. brevifrons. In contrast, leaflets in C. hildae and C. zaragozae are membranaceous. Leaflet symmetry can be curved as in C. fuscoviridis or straight from the base to the apex as in C. latifolia. In terms of leaflet lamina shape, some species are abaxially curved, while others are planar, as C. fuscoviridis and C. decumbens, respectively.
Reproductive morphology. All qualitative characters of both the pollen strobili and the ovulate strobili are not very polymorphic within and between populations, with the exception of strobilar apices in ovulate plants for certain species, e.g. Ceratozamia brevifrons, which can have acuminate and apiculate apex. Several reproductive characters contribute to diagnosable characters separating closely related taxa.
Ovuliferous strobili provide most of the diagnostic/differential in Ceratozamia. The most relevant of these characters are the colour of the strobilus, the shape of the fertile part of the entire strobilus, horn form in the distal end of sporophylls and colour of the sarcotesta (Fig. 5). The colour of ovuliferous strobili at maturity vary from dark green, as in C. mexicana, to wine red, as in C. decumbens. The shape of the strobilus is cylindric in most species, with the exception of C. morettii and C. latifolia where it is globose. In C. delucana, this character is polymorphic as both shapes can occur.
Distribution and habitat. Endemic to Mexico and only known from the State of Veracruz, in the vicinity of the Sierra de Chiconquiaco, at the transition zone between cloud forest and oak forest, between 500 and 1,350 m of elevation (Fig. 8).

Ceratozamia chamberlainii
Distribution and habitat. Its range includes the states of San Luis Potosí, Querétaro and Hidalgo ( Fig. 9). Ceratozamia chamberlainii occurs in the cloud forest on rocky outcrops at 900-1,200 m.
Etymology. The specific epithet honours Charles Joseph Chamberlain in recognition of his outstanding contributions to knowledge of the biology of the cycads and his fieldwork on Mexican cycads.
Distribution and habitat. Endemic to a small mountain range in central Veracruz, 450-1,000 m elevation (Fig. 8). The vegetation type of the habitat is mountain tropical forest and cloud forest on karstic rocks.
Etymology. The epithet alludes to the decumbent nature of trunks in older mature plants.
Distribution and habitat. This species is known from the states of Veracruz and Puebla at 200-700 m in evergreen tropical forest (Fig. 8).
Etymology. The epithet is in honour of Dr. Paolo De Luca, Professor at University of Naples Federico II and researcher in the biology of Mexican cycads.

Ceratozamia fuscoviridis
Distribution and habitat. Endemic of south-central Carso Huasteco, from central Hidalgo mountain range, to southeast of Hidalgo, including the western portion of Veracruz at an elevation ranging between 1,800-1,900 m in cloud forest (Fig. 9).

Ceratozamia hildae
Distribution and habitat. Endemic to Mexico in San Luis Potosí (Fig. 9), in the evergreen tropical forests.
Distribution and habitat. Endemic to Mexico in Tamaulipas at 1,100-1,500 m ( Fig. 9), in pine-oak and cloud forests.
Etymology. The specific epithet is in honour of Baron K. von Kuester, 19 th century plant collector.

Ceratozamia latifolia
Distribution and habitat. This species is distributed widely in San Luis Potosí and southeast of Querétaro mountain region, between 600-1,100 m elevation (Fig. 9). The vegetation type of its habitat is pine-oak forest and cloud forest.

Ceratozamia mexicana
Distribution and habitat. Endemic to Mexico from the River La Antigua drainage system within Xico, Teocelo and Coatepec municipalities to south end of Sierra Madre Oriental in Veracruz state, between 500-1,300 m in cloud forest (Figure 8).
Etymology. The epithet is derived from the country of origin of the material for the description of the genus.
Distinguishing features. Ceratozamia mexicana is distinguished by its lanceolate, coriaceous and flat leaflets with a symmetric apex, ovulate strobilus dark green with black and grey trichomes at maturity, and a prominent distal face with an obtuse angle between horns (Fig. 11). Taxonomic comments. The taxonomy of Ceratozamia mexicana has undergone constant changes since its original publication in 1846. The identity of this binomial was ambiguous for several decades. This taxonomic uncertainty was due to the complexity of relationships with some of the other names published by Miquel during the immediately succeeding years (1847, 1848 and 1849), following the original publication of the genus -namely, C. brevifrons, C. robusta, C. intermedia, C. longifolia, and C. latifolia. Later on, some names were placed in synonymy and others were characterised as nomina nuda; in particular, C. brevifrons was listed as a synonym of C. mexicana under the assumption that it was in fact a juvenile form of the latter (see Miquel 1848Miquel , 1861. In further taxonomic treatments of Ceratozamia -specifically, in De Candolle (1864-1868) and including Miquel's Nouveaux matériaux pour servir à la connaissance des Cycadées (1868-1869a, b)-this nomenclatural status was maintained. In his mature work, Miquel considered the variation observed between juveniles and adults was due to the corresponding ontogenetic modifications and collapsed into synonymy with C. mexicana all five species previously described.
Half a century later, Schuster (1932) circumscribed the genus to include only two species: Ceratozamia kuesteriana and C. mexicana, with two varieties and two forms. However, in the first modern taxonomic treatment (Vovides et al. 1983) and continuing with the typification of names within the genus , the type species was circumscribed to the central region of Veracruz. In this context, Vovides et al. (1983) attributed a wide geographic distribution and high variation of morphological characters to C. mexicana while recognising three varieties: C. mexicana var. mexicana, C. mexicana var. latifolia (Miq.) Schuster and C. mexicana var. robusta (Miq.) Dyer.

Ceratozamia morettii
Distribution and habitat. Endemic to Veracruz, on karstic rocks and cliffs of the Sierra de Chiconquiaco, between 1,200-1,850 m (Fig. 8). The vegetation type where this species grows is cloud forest.
Etymology. The specific epithet honours Aldo Moretti, in recognition of his scientific contributions in the field of cycad biology. He is a researcher in the Orto Botanico and Istituto di Biologia Vegetale at the University of Naples Federico II, Italia.
Distinguishing features. Ceratozamia morettii is similar to C. delucana in leaf morphology; however, there are differences in the total size of plant and in reproductive structures. This species has leaves with up to 23 pairs of leaflets and the ovulate strobilus is green with blackish trichomes at maturity, 14-18 cm long, 8-9.5 cm in diameter and an apiculate apex. Moreover, C. morettii has minimal variation at the population level. Type. MEXICO. Querétaro: San Joaquín, 15 Apr. 1991, A. P. Vovides 1205.
Distribution and habitat. Endemic to Mexico and known from the states of Querétaro and Hidalgo, at 1,600-1,900 m in the Sierra Gorda mountain range of Querétaro, along the mountain range northwest of Hidalgo (Fig. 9). It inhabits the understorey herbaceous layer of the transition zone between oak forest and cloud forest.
Etymology. The specific epithet honours the late Sergio Sabato, distinguished professor at the University of Naples Federico II, Italia, for his contributions to knowledge of the biology and systematics of cycads, particularly in the Neotropics.
Distribution and habitat. Endemic to Mexico in the central Veracruz mountain region at 1,200-1,850 m elevation on volcanic soils with basaltic rocks (Fig. 8). The vegetation type of the habitat is cloud forest.
Distribution and habitat. Endemic to Mexico in the Sierra Norte of Puebla mountain region on rocky outcrops in exposed walls up to 80 m at 600−1,050 m (Fig. 8). It inhabits the transition zone between cloud forest and evergreen tropical forest.
Etymology. The specific epithet makes reference to the Totonaco ethnic group of Santiago Ecatlán in Sierra Norte of Puebla, whose residents use and manage this species in local cultural contexts related to rituals.
Distinguishing features. This species is distinguished by its petioles with abundant and long, thin prickles, brown leaves at emergence, but the colour disappears in the adult leaves; leaflets are oblong and papyraceous with asymmetric apex. Ovulate strobilus yellowish-green with brown trichomes.

Discussion
The taxonomic history of species names in the genus Ceratozamia, as presented here, allow us to conclude that infrageneric concepts in this gymnosperm taxon are increasingly stringent and that infraspecific ranks are no longer recognised (cf. Osborne and Walters 2004). Most of these changes elevate varieties through the inclusion of new species. In general, several taxonomic changes have taken place, including diverse transfers at the species level, and substantial taxonomic disagreements have occurred, such as those involving the synonymy of C. microstrobila. In particular, the case of C. mexicana has been controversial, as demonstrated by the series of relationships and taxonomic changes, which have been established around it. With respect to the recent publication of two independent works in which contrasting taxonomic scenarios were proposed with respect to C. mexicana (Medina-Villareal and González-Astorga 2016; Vovides et al. 2016), the present morphological evaluation validates the proposal of Vovides et al. (2016).
Further support can be raised in connection with our position on recent taxonomic stances for Ceratozamia mexicana. On the basis of their phenetic analysis, Medina-Villarreal and González-Astorga (2016) suggested that the type species of the genus, C. mexicana, should be geographically anchored to the locality of the C. brevifrons neotype, given its morphometric similarity to the C. mexicana holotype. Under this assumption, these authors consider that C. decumbens, C. morettii and C. brevifrons should all be synonyms of C. mexicana. On the other hand, according to Medina-Villarreal and González-Astorga (2016), populations from the Naolinco Valley, Veracruz, should be described as a new species. Vovides et al. (2016) disagree on this point and suggest that those populations correspond in turn to C. mexicana var. tenuis, whose taxonomic treatment would then be recognised at the species level as C. tenuis. Under this perspective, C. mexicana would be recircumscribed to populations in the southern extreme of Veracruz. With our recircumscription proposal and review of historical documents, we consider that the locality of the C. mexicana holotype corresponds to the proposal of Vovides et al. (2016).