Phylogenetic and geographic diversification/differentiation as an evolutionary avenue in the genus Cephalocereus (Cactaceae) Evolutionary Avenue in Cephalocereus

Phylogenetic and geographic diversification/differentiation as an evolutionary avenue in the genus Cephalocereus (Cactaceae) Evolutionary Avenue in Cephalocereus | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Phylogenetic and geographic diversification/differentiation as an evolutionary avenue in the genus Cephalocereus (Cactaceae) Evolutionary Avenue in Cephalocereus Héctor J. Tapia, Salvador Arias, Juan J. Morrone, Patricia Dávila This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-2481800/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Phylogenetic and geographic distances were estimated to produce a combined Distancing Index as a measure of historical reproductive isolation in the genus Cephalocereus . Geographic and climatic barriers were inferred from potential and observed distributions. Distances were extracted from a Bayesian tree for seven chloroplast regions and 26 terminals, and from the geolocation dataset for the exact same sampling. Distance matrices were relativized, and a Mantel test was applied to identify deviations between phylogenetic and geographic distances. Heatmaps and scatterplots were implemented to visualize combined trends. Two basal clades show patterns of differentiation or complete isolation; the first includes C. scoparius , C. apicicephalium , C. nizandensis , and C. totolapensis , and the second includes C. parvispinus , C. polylophus , and C. euphorbioides . The species C. fulviceps , and C. sanchezmejoradae appear in a differentiated grade as sisters of a well-defined clade that includes C. mezcalaensis , C. macrocephalus , C. tetetzo , C. senilis , C. columna-trajani , C. multiareolatus , and C. nudus , where geographic or phylogenetic distances lie below the mean, indicating a diversification process in absence of hard barriers. At the generic level, separation is related to climatic factors as temperature and moisture, while factors as the altitude could be determinants of separation at the species level. The steady accumulation of variants may lead to opposed evolutionary outcomes: differentiation or diversification, in isolated and non-isolated lineages, respectively. More studies are needed on how genetic variation is transferred or interchanged between and among lineages, and how morphological differentiation of diverging lineages account for reproductive isolation. divergence cladogenesis reproductive isolation syngenesis Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Introduction After Darwin ( 1859 ), geography has been acknowledged as a factor that affects the composition of biotas and drives to morphological change of living beings after uncountable generations in isolation have passed, as some of the most evident aspects of biological evolution. Currently, the emergence of new kinds of organisms are strongly related to spatial and reproductive isolation (allopatric speciation), which is considered an ordinary way for biological diversity to be produced and to be maintained functionally separated in distinguishable classes, in contrast to other alternative processes as sympatric speciation (Lande, 1980 , De Queiroz, 2007 ; Fitzpatrick et al., 2009 ). Must isolation be understood as an evident and sustained discontinuity in spatial or temporal distributional traits; isolation is equivalent to separation, regardless of the appearance of genetic or morphological differentiation. Then, isolation can account for genetic and morphological differentiation in a conditional manner, when genetic flow ceases and mutation and genetic drift produce noticeable among-populations changes (Slatkin, 1987 ). Given the independence between isolation and differentiation it is necessary to measure the actual distances in structural (morphologic or genetic) or functional (ecologic or physiologic) attributes, solely or in correlation with the spatial or temporal (time since) separation. To date, most of the evolutionary methods and related approaches have emphasized on an intrinsic inclination of living beings to drift apart from their relatives, as in population genetics, systematics, biogeography and ecology, among others (e.g. Karlsson-Green et al., 2014 ). The cladogenetic events depicted in a branching phylogenetic diagram or a phylogenetic tree are cases of this logic, even when other evolutionary processes would also be accounted for (Lande, 1980 ; Schnable et al., 1998 ; Soltis & Soltis, 2009 ). The ecological isolation and the isolation by distance are also examples of divergentist models, where spatial or ecological isolation and differentiation are expected to occur simultaneously. On the other hand, processes as hybridization and incomplete lineage sorting (ILS) reveal discrepancies in observed characters, either morphological, genetical or both (Soltis & Soltis, 2009 ; Murillo et al., 2022 ). Hybridization events in plants are responsible for the appearance of a noticeable number of new species, but in the overall results insufficient to be considered a main evolutionary process. The present study advocate for an alternative point of view, where convergence may indeed be as important as divergence is to the evolutionary processes, none of them being exclusive over the other. Convergence is referred here as a syngenetic or reproductive process and is opposed to the cladogenetic or reproductive isolation process, either if it is intrinsically or extrinsically driven. This alternative approach requires the development of new methodologies or new interpretations of current methods. For example, common distance methods estimate rates of evolutionary change or change accumulation through time, between and among populations and species, including geographic and genetic distances, and populational parameters as the flow of individuals and their genes (Nei, 1972 ; Slatkin, 1993 ). The present study follows an exploratory viewpoint and is intended to quantify the amount of evolutionary change accumulated between and among species and species groups, as well as to contrast the geographic distances in a structured and relativized framework, such that the different groupings (from populations to genera) can be located and compared in a simplified graphical and statistical manner. The family Cactaceae is an outstanding model for the study of geographic and climatic isolation because it includes species highly adapted to harsh environments in arid and semiarid regions of the Americas (Bravo-Hollis, 1978 , Gibson & Nobel, 1986 , Anderson, 2001 ). Several authors have related the ability to survive in these environments to structural and physiological features, as the presence of spines, reduced or absent leaves, succulent tissues, growth forms, and CAM metabolism, among others (Bravo-Hollis, 1978 ; Anderson, 2001 ; Hernández-Hernández et al, 2011 ). Extrinsic microhabitat features also play a key role in the establishment and survival of plantlets, as occur with rocky outcrops, cliffs, nurse plants and nurse rocks (Peters et al., 2008 , Bárcenas-Argüello et al., 2010 , Ocampo & Columbus, 2010 , Arakaki et al., 2011 ). Geographically, it has been recognized the influence of topographic and climatic factors limiting distributions and affecting the diversity of cactus species, as high elevations, or high precipitations, added to the seasonal and historical variation in climatic conditions (see Barthlott et al., 2015 ). Cephalocereus is endemic to Mexico and a common representative of the columnar cacti in the central and southern regions (Bravo-Hollis, 1978 ). Much of its species’ diversity occurs among the states of Jalisco, Colima, Michoacán, Guerrero, Oaxaca, Puebla, and Chiapas, within the Southeast Lowlands, Balsas Basin, and Sierra Madre del Sur biogeographic provinces. Some species are distributed northward, crossing the Transmexican Volcanic Belt, into the Sierra Madre Oriental and Veracruzan provinces, in the states of Hidalgo, Querétaro, San Luis Potosí, Guanajuato, Veracruz, and Tamaulipas. Provinces are referred following Morrone ( 2019 ; Morrone et al., 2022 ). Particularly, the Tehuacan-Cuicatlan valley region (Oaxaca and Puebla) has the higher species’ diversity with five out of sixteen species occurring there (Bravo-Hollis, 1978 , Gibson & Nobel, 1986 , Anderson, 2001 , Hunt et al., 2006). Our hypothesis is that geographic and climatic barriers have produced the isolation or separation of the Cephalocereus genus into phylogenetically differentiated species and species groups, while the absence of barriers in part of its distributional range has led to processes of phylogenetic diversification while developing less differentiated instead. Material And Methods Taxonomic and genetic sampling. Taxon sampling includes 26 terminals corresponding to the 16 recognized species in the genus Cephalocereus , and two closely related species as outgroup: Bergerocactus emoryi and Marshallocereus aragonii ssp. eichlamii (Tapia et al., 2017 ; Arias et al., 2019 ; Arias in Korotkova et al., 2021 ); Bergerocactus and Marshallocereus hereafter. Disjunct or widespread species’ distributions are represented with two and three terminals respectively. Collected species and distributions are detailed in Table 1 , and approximate collecting sites are shown in Fig. 1 . Table 1 Species, locations, and distribution types. Species Key Collect number Distribution type Cephalocereus C. apicicephalium SA1257 Restricted C. columna-trajani SA1377 Restricted C. euphorbioides (VER) HJTS001 Disjunct (TAMPS) Hamman (cult.) C. fulviceps HJTS029 Medium C. macrocephalus HJTS028 Restricted C. mezcalaensis (GRO) HJTS036 Widespread (MOR) HJTS042 (PUE) HJTS044 C. multiareolatus HJTS034 Restricted C. nizandensis TT633 Restricted C. nudus (JAL) HJTS030 Widespread (OAXa) HJTS011 (OAXb) HJTS024 C. parvispinus HJTS038 Single site C. polylophus HJTS007 Medium C. sanchezmejoradae HJTS039 Single site C. scoparius (OAX) HJTS010 Disjunct (VER) HJTS002 C. senilis TT529 Restricted C. tetetzo (OAXa) HJTS020 Widespread (OAXb) HJTS041 (PUE) SA1376 C. totolapensis TT628 Restricted Bergerocactus emoryi SA1307 Medium Marshallocereus eichlamii SA1363 Medium Genetic sampling includes 7 chloroplast loci: rpl16 , trnL-F , psbA-trnH , rpl32-trnL , trnQ-rps16 , petL-psbE , and ycf1 . All the sequences have been generated and deposited previously in the GenBank (see Table 2 ). PCR conditions are detailed in Tapia et al. ( 2017 ). DNA sequences were aligned and trimmed, producing a slightly different matrix length in base pairs from the previous analysis, due to the absence of some indels in the actual taxa sample. Primers, sizes, and basic statistics for each region are shown in Table 3 . Table 2 GenBank accessions. Terminal Accession numbers ( rpl16 , trnL-F , psbA-trnH , rpl32-trnL , trnQ-rps16 , petL-psbE , ycf1 ) C. apicicephalium DQ099996, DQ099927, KY624684, KY624717, KY624756, KY624787, KY624825 C. columna-trajani AY181599, AY181648, KY624685, KY624718, KY624757, KY624788, ND C. euphorbioides (VER) KY624637, KY624651, KY624666, KY624699, KY624738, KY624769, KY624808 C. euphorbioides (TAMPS) AY181595, AY181635, KY624667, KY624700, KY624739, KY624770, KY624809 C. fulviceps AY181602, AY181621, KY624683, KY624716, KY624755, KY624786, KY624824 C. mezcalaensis (PUE) KY624638, KY624652, KY624669, KY624702, KY624741, KY624772, KY624811 C. mezcalaensis (MOR) KY624639, KY624653, KY624670, KY624703, KY624742, KY624773, KY624812 C. mezcalaensis (GRO) AY181600, AY181645, KY624671, KY624704, KY624743, KY624774, KY624813 C. macrocephalus DQ100013, DQ099944, KY624668, KY624701, KY624740, KY624771, KY624810 C. multiareolatus AY181594, AY181636, KY624672, KY624705, KY624744, KY624775, KY624814 C. nizandensis DQ099997, DQ099928, KY624686, KY624719, ND, KY624789, ND C. nudus (JAL) KY624644, KY624658, KY624679, KY624712, KY624751, KY624782, KY624821 C. nudus (OAXa) KY624643, KY624657, KY624678, KY624711, KY624750, KY624781, KY624820 C. nudus (OAXb) KY624642, KY624656, KY624677, KY624710, KY624749, KY624780, KY624819 C. parvispinus MK165433, MK165434, MK165435, MK165436, MK165437, MK165438, MK165439 C. polylophus AY181597, AY181644, KY624673, KY624706, KY624745, KY624776, KY624815 C. sanchezmejoradae KY624640, KY624654, KY624674, KY624707, KY624746, KY624777, KY624816 C. scoparius (OAX) AY181596, AY181625, KY624675, KY624708, KY624747, KY624778, KY624817 C. scoparius (VER) KY624641, KY624655, KY624676, KY624709, KY624748, KY624779, KY624818 C. senilis AY181616, AY181638, KY624687, KY624720, KY624758, KY624790, KY624826 C. tetetzo (PUE) AY181592, AY181632, KY624680, KY624713, KY624752, KY624783, ND C. tetetzo (OAXa) KY624645, KY624659, KY624681, KY624714, KY624753, KY624784, KY624822 C. tetetzo (OAXb) KY624646, KY624660, KY624682, KY624715, KY624754, KY624785, KY624823 C. totolapensis KY624647, KY624661, KY624688, KY624721, KY624759, KY624791, KY624827 B. emoryi DQ099994, DQ099925, KF783478, KY624730, KF783697, KY624800, KY624834 M. eichlamii AY181610, AY181629, KY624690, KY624723, KY624760, KY624793, KY624828 Table 3 Plastid regions employed, and primer sequences. Region Sequence Expected length Reference rpl16 F(1F) GCT ATG CTT AGT GTG TGA CTC GTT 900–1,200 Hernández-Hernández et al. 2011 R(3R) CTT CTA TTT GTC TAG GCG TGA TCC trnL-F F€ CGA AAT CGG TAG ACG CTA CG 700–1,100 Taberlet et al. 1991 R(f) ATT TGA ACT GGT GAC ACG AG F-int€ GGT TCA AGT CCC TCT ATC CC R-int(d) GGG GAT AGA GGG ACT TGA AC psbA-trnH F(psbA) GTT ATG CAT GAA CGT AAT C 300–700 Sang et al. 1997 Tate and Simpson 2003 R(trnH2) CGC GCA TGG TGG ATT CAC A rpl32-trnL F(rpl32) CAG TTC CAA AAA AAC GTA CTT C 500–1,400 Shaw et al. 2007 R(trnL UAG ) CTG CTT CCT AAG AGC AGC GT F-int GTA ACT CTT GAA ATC ATT ATT TC Plume et al. 2013 R-int GTT ATC TTA GGT TTC AAC AAA CC trnQ-rps16 F(trnQ UUG ) GCG TGG CCA AGY GGT AAG GC 600–1,500 Shaw et al. 2007 R(rps16 XI ) GTT GCT TTY TAC CAC ATC GTT T petL-psbE F(petL) AGT AGA AAA CCG AAA TAA CTA GTT A 400–700 Shaw et al. 2007 R(psbE) TAT CGA ATA CTG GTA ATA ATA TCA GC ycf1 F(4182) AAA TAY RRA TAG AAA ATA TTT KGA TT 900–1,100 Franck et al. 2012 R(5248) GAA TTC TYA ATT CTC TAC GAC G Phylogenetic analysis. A Bayesian Inference analysis was conducted in MrBayes 3.2.6 (Huelsenbeck & Ronquist, 2001 ), using the GTR + I + G model, during 10’000’000 generations, without matrix partitions due to their same-organelle origin, it was applied a burnin fraction = 0.5. Statistics for trees and probabilities were summarized to visualize topologies, convergence of chains, branch lengths, and posterior probabilities. The consensus Bayesian tree with branch lengths and posterior probabilities appended was rooted using the outgroup, exported in newick format, and retained for further analyses. Distance analyses. A phylogenetic pairwise distance matrix (DGEN) was computed from the Bayesian consensus tree, imported, and analyzed in R environment (RStudio 3.6.3, 2020), using the ‘ape’ library. The resulting table (26x26) contains two symmetrical triangles and a diagonal with only-zero values that can be reduced to 325 single comparison values. This table was then transformed into relative values (positives from ~ 0 to 1), dividing each cell by the maximum value in the table. Geographic pairwise distance matrix (DGEO) was computed in Qgis software (2016; ver. 3.22), using geographic data (latitude and longitude) for the same 26 terminals included in the phylogenetic analysis. Linear Geographic distance was computed resulting in an identical data structure. These values were standardized and reduced in the same way as for phylogenetic distance matrix. Absolute and relative values for both datasets are provided as supplemental data. Spearman’s correlation and Mantel test. Phylogenetic distances were tested for spatial correlation against geographic distances, to evaluate the existence of deviations from null hypothesis. These analyses were implemented in Excel (tm) using the complement GenAlEx6.5 (Peakall & Smouse, 2012 ). Non-parametric correlation analysis was performed using Spearman’s coefficient and a Mantel test with the maximum allowed permutations (9’999). Climatic barriers. Geographic data from samples were integrated into a larger geolocation dataset. This matrix contains 423 records for species from the three genera, 407 of them corresponding to Cephalocereus . At the generic level, all the records were considered, but at species’ level some records were discarded from species with restricted distributions or species known from a single locality. The species not considered were C. apicicephalium , C. nizandensis , C. parvispinus , C. sanchezmejoradae , C. senilis , and Marshallocereus . Climatic models were performed using MaxEnt 3.4.4 (Phillips et al., 2006 ), generating potential distribution maps to identify climatic barriers and environmental factors limiting species distributions. Distancing index (d). Both datasets, phylogenetic and geographic, were verified as Euclidean values, then relativized, and identically pairwise organized. A combined distance measure was implemented using standardized phylogenetic and geographic distance matrices between pairs of terminals, as follows: $$d=\sqrt{{p}^{2}+{g}^{2}}$$ where p and g represent respectively the phylogenetic and geographic distances between the same pair of terminals. The limit of this index is \(\sqrt{2}\) (≈ 1.4142). The following assumptions were taken into account to interpret the distancing index : a) phylogenetic distance is indicative of the time since two lineages split up, b) distancing index can be translated as a probability of reproductive isolation between two lineages, c) differentiated lineages reach the higher distance values in both dimensions, d) non-differentiated lineages will have lower distance values in one dimension, phylogenetic or geographic, and could be interpreted as diversified groups, and e) existence of a geographic or climatic barrier is crucial to decide whether or not exists a differentiation or a diversification process. Given that assumptions, a decision matrix was constructed to evaluate the results as follows: Where higher distance values (+ +) indicate a complete differentiation process, combination of low and high distance values (- +, + -) indicate diversification processes, and both lower values (- -) indicate an identity process, or a continuous reproductive system. Distance and diversification/differentiation matrices were represented as heatmaps as a visual guide to identify multiple simultaneous relationships. Results Geographic and climatic barriers. Three main barriers were inferred from species’ distributions and climatic models, not only as unoccupied areas but also as areas with a factor accountable for impeding the establishment of Cephalocereus , Bergerocactus or Marshallocereus species. The first climatic barrier is in southeastern Mexico, as a stripe broader than 80 km running from southern Veracruz towards the Caribbean Sea, along the Gulf of México Lowlands province, and separating the Yucatán peninsula from the continent. The model for all the Cephalocereus species exhibits a pattern of avoidance related to temperature and moisture factors: temperature annual range (bio7), precipitation of coldest quarter (bio19), and mean temperature of warmest quarter (bio10). The second climatic barrier is in northwestern Mexico, between Baja California peninsula and the continent, separating Bergerocactus from all other species. The climatic variables are also related to temperature and moisture factors: mean temperature of warmest quarter (bio10), precipitation of coldest quarter (bio19) and precipitation of warmest quarter (bio18). Statistics for these potential distribution models are presented in Table 4 , following the same abbreviations as in text. Table 4 Climatic and geographic barriers, main variables, and contributions. Barrier Type Variable Percent contribution Permutation importance South Veracruzan Climatic bio10 29.4 0.4 bio19 27.1 10.2 bio18 21.1 1.8 bio17 11.8 76.7 Transmexican Volcanic Belt Greographic - - - Baja-californian Climatic bio7 56.4 19.8 bio19 12.8 11.8 bio15 11.7 18 The third barrier is located along the Transmexican Volcanic Belt and is explained geographically, possibly related to altitudinal or topographical constrains, given that modelled and observed distributions did not match in this region. Observed and modelled distributions for the genera Cephalocereus and Bergerocactus , as well as climatic and geographic barriers are shown in Fig. 2 . The species’ distribution models are mostly compatible with the generic models (data not shown). Tree topology. The Bayesian tree shows high support values in most of the internal nodes. This tree is represented as a phylogram to show the correspondence with the phylogenetic distance matrix (Fig. 3 ). Two clades derived basally: the first including the species C. scoparius , C. apicicephalium , C. nizandensis , and C. totolapensis , and the second including C. parvispinus , C. euphorbioides , and C. polylophus . Posterior probabilities for the first clade are low, although it is spatially congruent. Two species, C. fulviceps , and C. sanchezmejoradae , appear as sister taxa of a major clade, which includes the most variable and widespread species C. mezcalaensis , C. nudus , and C. tetetzo , along with the restricted species C. macrocephalus , C. multiareolatus , C. columna-trajani , and C. senilis . Species within this clade occupy an area extended from the Sierra Norte de Oaxaca to the Sierra Madre del Sur, the Balsas Basin, and the Pacific Lowlands, except for C. senilis , which inhabits the Sierra Madre Oriental, at the north of the Transmexican Volcanic Belt. Distancing Index and Differentiation. Spatial and temporal dimensions are presented in a relative scale (0–1, or 0–100%) and provide a framework to explain relationships from both phylogenetic and geographic variation. Samples at species or under species ranks exhibit relative distances below 10% in phylogenetic distances. In this segment are included the species C. mezcalaensis , C. nudus , C. tetetzo , C. euphorbioides , and C. scoparius , which exhibit widespread or disjunct distributions (see Table 1 ). Three more species, C. apicicephalium , C. nizandensis , and C. totolapensis ( Neodawsonia clade), show distances standing in the same range of the intraspecific measures. Some differentiated species and species groups are distinguishable, as C. scoparius and the clade formed by C. parvispinus , C. euphorbioides , and C. polylophus . On the counterpart, a group of close or less differentiated species includes C. mezcalaensis , C. multiareolatus , C. nudus , C. macrocephalus , C. tetetzo , C. columna-trajani , and C. senilis (Fig. 4 A). The geographic distances alone show less structuration than phylogenetic distances. Climatic barriers are responsible for the spatial separation between genera at a higher spatial scale, while the geographic barrier appears to have a moderate influence on geographic distances between Cephalocereus species. The species occurring northward of the Transmexican Volcanic Belt show geographic distances from 11.4% in C. columna-trajani / C. senilis , to 16.8% in C. parvispinus / C. euphorbioides , and 15% in C. parvispinus / C. polylophus , respectively, and one locality of C. nudus at 35.3% distance collected far in western Jalisco (Fig. 4 B), indicating a possible isolation from other species’ populations. The distancing index ( d g ) reveals a normalizing effect, tending to the phylogenetic data. The only major change compared to phylogenetic distances alone is an increment in the average distance from C. nudus (Jalisco) (Fig. 4 C). No other significative change from the phylogenetic distance matrix is evident. Differentiation patterns The estimated slope (y = 0.5084x − 0.0059) and the coefficient of determination from Spearman’s correlation analysis (R 2 = 0.2682) indicate a significant deviation between phylogenetic and geographic distances. The scatterplot from correlation analysis shows three clearly separated groups, representing the genera Bergerocactus , Cephalocereus , and Marshallocereus (Fig. 5 A). Genus Marshallocereus is separated in the extreme of the phylogenetic distance axis, while Bergerocactus is separated in the geographic distance axis (Fig. 5 B). The dots representing Cephalocereus distances appear almost randomly distributed in the midst and lower values for both axes, but some groups can be detected in between. Patterns of phylogenetic differentiation are evident in two clades; the first formed by C. scoparius , C. apicicephalium , C. nizandensis , and C. totolapensis , (Fig. 5 C), and the second formed by C. euphorbioides , C. polylophus , and C. parvispinus (Fig. 5 D). On the other hand, the clade containing C. mezcalaensis , C. multiareolatus , C. nudus , C. macrocephalus , C. tetetzo , C. columna-trajani , and C. senilis , show less phylogenetic differentiation and proportionally major geographic distances (Fig. 5 E, F). The two remaining species, C. fulviceps and C. sanchezmejoradae , have low affinity for any other clade or species, also exhibiting a pattern of phylogenetic differentiation (Fig. 5 G). Lastly, the intra-species distances are presented for species with two or more representatives, and for the Neodawsonia clade, given the short distances between these species in both dimensions, phylogenetic and geographic (Fig. 5 H). Discussion Phylogenetic distancing . The most distant species were C. polylophus and M. aragonii ssp. eichlamii , which was taken as the intergeneric distance (100%). This distance was 31 times greater than that between the sister species C. apicicephalium and C. nizandensis . The distance between M. aragonii ssp. eichlamii and B. emoryi is 56.9%, while the most distant species within Cephalocereus lies in a 58.5% distance ( C. polylophus / C. sanchezmejoradae ). Other relevant comparisons are the distances among Neodawsonia species which barely reach 3.2% in C. apicicephalium / C. nizandensis , 4% in C. apicicephalium / C. totolapensis , and 5.4% in C. nizandensis / C. totolapensis . The intraspecific distances range from 4.3% in the disjunct C. euphorbioides to 9.1% in the widespread C. mezcalaensis , while the species C. columna-trajani and C. tetetzo , which present natural hybrids (Vite et al., 1996 ), lie in a 13.4% distance range. These results indicate not only the recentness of the Neodawsonia splitting, but the potential for other phylogenetically and geographically close species to produce genetic transferences or interchanges. Other examples are C. scoparius and C. nizandensis , separated barely 16 km away with a moderate phylogenetic distance from 25.9 to 28.8%, C. tetetzo and C. nudus , separated 25 km in a phylogenetic distance range from 15.9 to 20.4%, and C. macrocephalus and C. tetetzo , that make contact in one site and have phylogenetic distances from 11.5 to 13.1%. In this population of C. macrocephalus and C. tetetzo some putative hybrid individuals have been observed (Arias & Tapia, pers. obs.). Up to date, several intergeneric hybrids have been reported in the tribe Echinocereeae, where Cephalocereus belongs, as × Pachebergia ( Backebergia × Pachycereus ), × Pacherocactus ( Bergerocactus × Pachycereus ), × Myrtgerocactus ( Bergerocactus × Myrtillocactus ), × Myrtillenocereus ( Myrtillocactus × Stenocereus ), and some interspecific hybrids are documented in the sister tribe Hylocereeae, between Hylocereus undatus , H. monacanthus , and H. megalanthus , where H. megalanthus probably speciate from same species parentals by autopolyploidy (Anderson, 2001 ; Arias & Terrazas, 2008 ; Plume et al., 2013 ; Granados-Aguilar et al., 2022 ). The placement of hybridizing species in the same genus or in separated genera based on their morphology have implications on what we suppose to occur at cellular and genetic levels in such hybridization event, whereas morphology alone can be an overestimated indicator of differentiation and reproductive isolation (Plume et al., 2013 ). The hybrid between C. columna-trajani and C. tetetzo was described initially as an intergeneric hybrid between Neobuxbaumia tetetzo and Cephalocereus columna-trajani , later recovered as sister species in a phylogenetic analysis, along with C. senilis , and reassigned altogether in the same genus ( Cephalocereus ) (Tapia et al., 2017 ). Other example is Backebergia militaris , which presented enough morphological characters to be assigned to a monotypic genus according to Bravo-Hollis ( 1953 ), but phylogenetic studies resolved its position close to Pachycereus pecten-aboriginum , with which hybridize to produce x Pachebergia apicostata (Arias & Terrazas, 2008 ). The Cactaceae family, like other plant families, appears as a group inclined to hybridize at varying levels, since the same species, to interspecies, to intergeneric parentals, resulting in chaotic complexes as in wild opuntias, or in flashy cultivated varieties of Astrophytum (Granados-Aguilar et al., 2022 ). Attending to a morphometric assessment, the study on Cephalocereus areoles and spines revealed simultaneous patterns of character variation and conservation (Tapia et al., 2016 ). In some species a unique spination pattern allowed the univocal distinction of individuals as part of one species, as in C. polylophus , C. euphorbioides , and C. parvispinus , which correspond to a phylogenetically differentiated group in the present study. Other individuals show similarities that allowed the distinction of species groups, particularly the groups C. mezcalaensis / C. nudus / C. scoparius and C. tetetzo / C. macrocephalus / C. fulviceps are of particular interest due to the closeness between the included species, both in geographic and phylogenetic distances, and the possibility of genetic transferences or interchanges to occur, which needs to be studied further in detail. It is not intended to point that hybridization is responsible for all the evolutionary changes, but reproductive processes, either continuous or occasional, need to be accounted and comprehensible analyzed to untangle the genetic or genomic contribution of parental organisms, either if they belong to the same or to different species. Geographic barriers and splitting events . The effects of the climatic and geographic barriers between and among Cephalocereus species are consistent with a hypothesis of reproductive isolation. The clade formed by C. scoparius , C. apicicephalium , C. nizandensis , and C. totolapensis , which inhabit a region between the Tehuantepec Isthmus and the adjacent part of the Sierra Madre del Sur, represents an early divergence from the Marshallocereus / Cephalocereus ancestor, just followed by the Bergerocactus ancestor splitting. The species C. scoparius has a disjunct distribution occupying also a portion in the center of the Veracruzan province, caused probably by a relatively recent long distance dispersal event, denoted by a phylogenetic distance of 8.8%. The clade formed by C. euphorbioides , C. polylophus , and C. parvispinus becomes the next splitting event coupled with the dispersal northward into the Sierra Madre del Sur and then followed by the crossing of the Transmexican Volcanic Belt barrier. C. parvispinus remained in the Sierra Madre del Sur, probably ecologically isolated, and C. polylophus and C. euphorbioides ancestor colonized the Sierra Madre Oriental and the Veracruzan provinces. The phylogenetic distance between C. polylophus and C. euphorbioides is 20.3%, indicating an early divergence event between these species after their dispersal from the southern regions. On the counterpart, species non-isolated by hard or absolute barriers, show shorter distances either phylogenetic, geographic or both. This is the case of the clade formed by C. mezcalaensis , C. multiareolatus , C. nudus , C. macrocephalus , C. tetetzo , C. columna-trajani , and C. senilis . The only species from this clade that dispersed across the Transmexican Volcanic Belt is C. senilis . The phylogenetic distances among species of this clade ranges from 7.7% in the sister species C. columna-trajani / C. senilis , to 21.9% in C. senilis / C. nudus , while the geographic distances range from zero up to ~ 1’000 km. The species C. fulviceps and C. sanchezmejoradae have phylogenetic distances higher than 28% from the species belonging to the previous clade, despite the absence of a hard barrier, indicating the possible existence of ecological ancient barriers instead. Absolute geographic distances . Considering the size of Cephalocereus individuals and the type of pollinators (moths and bats), the distance for effective pollination ranges from meters up to several kilometers. The seeds display patterns of local dispersion led by fruit dehiscence and gravity action, or long-distance dispersal by migrating bats and birds, also up to several kilometers, and longer than in pollination (Valiente-Banuet et al., 1997 ; González-Terrazas et al., 2016 ). Phylogenetically differentiated clades and/or species are separated from other species or clades by barriers at least 80 kilometers wide or in altitudinal ranges of several hundreds of meters. In the absence of barriers, some species can occupy land stripes up to 800 km in length, like C. nudus which inhabits along the Pacific coastline in southern Mexico, from Jalisco and Colima to Oaxaca. In this case, the occupation of a broad region might had occurred gradually, pollination may be restricted to each population and seed dispersal limited to close populations. Given this structuration, it is likely that the most distant populations located in Colima and Jalisco are currently undergoing a differentiation process from same species populations in Oaxaca (> 600 km apart). Other species with disjunct distributions ( C. euphorbioides and C. scoparius ), show moderately to low phylogenetic distancing indicating recent dispersal events around 300 or 400 km away from their original distributions, probably driven by migrating birds or bats, and have become genetically isolated relatively few generations ago. Other component not-explicitly analyzed here is the populational genome, which act as a repository of evolutionary variance, where the genetic variants are gained and lost. Populational genome is involved in a third evolutionary process, the anagenesis (Emerson & Patiño, 2018 ), which is related to a slow and steady evolutionary phase where the main changes correspond to gene recombination and allelic fluctuation (Schnable et al., 1998 ), due to both determined and stochastic processes, as gene drift and selective factors, and require biparental and more variable molecular markers to be analyzed. Conclusions And Remarks This study was focused on the phylogenetic and geographic variation patterns, which concurrently account for the spatial and temporal isolation components. These components are conditionals that may or may not produce a noticeable differentiation in the resulting lineages, in morphological or genetical attributes, but if phylogenetic differentiation occurs it is always an indicative of spatial and temporal isolation (time since). Differentiation is related to the absence of reproduction among groups of individuals, while diversification is likely related to the added genetic variance from recent reproduction events between increasingly broader groups, in terms of number of individuals or occupied space, either if reproduction occurs in a continuous or in an occasional fashion. It must be noted that spatial barriers to reproduction are specific to each taxonomic group and need to be evaluated in their own ecological context. Practical applications of this type of analysis are currently under development and could serve to validate new species, to define significant spatial barriers to reproduction in evolutionary time, to distinguish and define species complexes where phylogenetic methods could not resolve the sisterhood of species, to assess the evolutionary trends (syngenetic or cladogenetic) in a taxonomic group, among others. Declarations Acknowledgments The first author thanks the Dirección General de Asuntos del Personal Acádemico (DGAPA) for the postdoctoral fellowship granted to carry out this research in the Laboratorio de Recursos Naturales, UBIPRO, FES Iztacala, Universidad Nacional Autónoma de México (UNAM). All authors thank Oswaldo Téllez and Gabriel Arroyo-Cosultchi for sharing some Cephalocereus geolocation records. Competing interests The authors have no competing interests to declare that are relevant to the content of this article. References Anderson EF (2001) The Cactus Family , USA: Timber Press, 776 pp. Arakaki M, Christin PA, Nyffeler R, Lendel A, Eggli U, Ogburn RM, Spriggs E, Moore MJ, Edwards EJ (2011) Contemporaneous and recent radiations of the world’s major succulent plant lineages. Proc Natl Acad Sci U S A 108: 8379–8384. https://doi.org/10.1073/pnas.1100628108 Arias S, Tapia HJ, Guzmán U (2019) A new species of Cephalocereus (Cactaceae) from southern Mexico. Phytotaxa 392(2): 147–156. https://doi.org/10.11646/phytotaxa.392.2.4 Arias S, Terrazas T (2008) x Pachebergia (Cactaceae), a nothogenus from western Mexico. Rev Mex Biodivers 79(1): 23–28. Bárcenas-Argüello ML, Gutiérrez-Castorena MC, Terrazas T, López-Mata L (2010) Rock–soil preferences of three Cephalocereus (Cactaceae) species of tropical dry forests. Soil Sci Soc Am J 74: 1374–1382. https://doi.org/10.2136/sssaj2009.0310 Barthlott W, Burstedde K, Geffert JL, Ibisch PL, Korotkova N, Miebach A, Rafiqpoor MD, Stein A, Mutke J (2015) Biogeography and biodiversity of Cacti. Schumannia 7 . 205pp Bravo-Hollis H (1953) Un nuevo género de la Familia de las Cactáceas Backebergia . An Inst Bio UNAM 24: 215–232. Bravo-Hollis H (1978). Las Cactáceas de México, vol. I . 2da Ed. México, DF: UNAM, 755pp. Darwin C (1859) On the origin of species by means of natural selection , London: J. Murray , 502pp. De Queiroz K (2007) Species concepts and species delimitation. Syst Biol 56(6): 879–886. https://doi.org/10.1080/10635150701701083 Emerson BC, Patiño J (2018) The evolution of Cladogenetic and Anagenetic concepts in Island Biogeography. Trends Ecol Evol 33(7): 488–491. Fitzpatrick BM, Fordyce JA, Gavrilets S (2009) Pattern, process and geographic modes of speciation. J Evol Biol 22: 2342–2347. Franck AR, Cochrane BJ, Garey JR (2012) Low-copy nuclear primers and ycf1 primers in Cactaceae. Am J Bot 99(10): e405-7. https://doi.org/10.3732/ajb.1200128 Gibson AC, Nobel P (1986) The cactus primer , USA: Harvard University Press, 286pp. González-Terrazas TP, Martel C, Milet-Pinheiro P, Ayasse M, Kalko EKV, Tschapka M (2016) Finding flowers in the dark: Nectar-feeding bats integrate olfaction and echolocation while foraging for nectar. R Soc Open Sci 3 (8): 160199. https://doi.org/10.1098/rsos.160199 Granados-Aguilar X, Rosas U, González-Rodríguez A, Arias S (2022) The prickly problem of interwoven lineages: hybridization process in Cactaceae. Bot Sci, 100(4): 797-813. Hernández-Hernández T, Hernández HM, De-Nova JA, Puente R, Eguiarte LE, Magallón S (2011) Phylogenetic relationships and evolution of growth form in Cactaceae (Caryophyllales, Eudicotyledoneae). Am J Bot 98(1): 44–61. https://doi.org/10.3732/ajb.1000129 Huelsenbeck JP, Ronquist F (2001) MrBayes: Bayesian inference of phylogenetic trees. Bioinformatics 17 (8): 754–755. Retrieved from http://www.ncbi.nlm.nih.gov/pubmed/11524383 Hunt DR (2006) The New Cactus Lexicon , dh Books, 527pp. Karlsson-Green K, Svensson EI, Bergsten J, Härdling R, Hansson B (2014) The interplay between local ecology, divergent selection, and genetic drift in population divergence of a sexually antagonistic female trait. Evolution 7: 1934–1946. Korotkova N, Aquino D, Arias S, Eggli U, Franck AR, Gómez-Hinostrosa C, Guerrero PC, Hernández HM, Kohlbecker A, Köhler M, Luther K, Majure LC, Müller A, Metzing D, Nyffeler R, Sánchez D, Schlumpberger B, Berendsohn W (2021) Cactaceae at Caryophyllales.org – a dynamic online species-level taxonomic backbone for the family. Willdenowia 51(2): 251–270. Lande R (1980) Genetic variation and phenotypic evolution during allopatric speciation. Am Nat 116: 463–479. Morrone JJ (2019) Regionalización biogeográfica y evolución biótica de México: encrucijada de la biodiversidad del Nuevo Mundo. Rev Mex Biodivers, 90: e902980. Morrone JJ, Escalante T, Rodríguez-Tapia G, Carmona A, Arana M, Mercado-Gómez JD (2022) Biogeographic regionalization of the Neotropical region: new map and shapefile. An Acad Bra Cienc 94: e20211167. Murillo J, Valencia J, Orozco CI, Parra C, Neubig KM (2022) Incomplete lineage sorting and reticulate evolution mask species relationships in Brunelliaceae, an Andean family with rapid, recent diversification. Am J Bot 109: 1139–1156. Nei M (1972) Genetic Distance between Populations. Am Nat 106(949): 283–292. Ocampo G, Columbus JT (2010) Molecular phylogenetics of suborder Cactineae (Caryophyllales), including insights into photosynthetic diversification and historical biogeography. Am J Bot 97(11): 1827–1847. https://doi.org/10.3732/ajb.1000227 Peakall R, Smouse PE (2012) GenAlEx 6.5: genetic analysis in Excel. Population genetic software for teaching and research-an update. Bioinformatics 28: 2537–2539. Peters EM, Martorell C, Ezcurra E (2008) Nurse rocks are more important than nurse plants in determining the distribution and establishment of globose cacti ( Mammillaria ) in the Tehuacán Valley, Mexico. J Arid Environ 70: 593–601. https://doi.org/10.1016/j.jaridenv.2007.10.004 Phillips SJ, Anderson RP, Schapire RE (2006) Maximum entropy modeling of species geographic distributions. Ecol Modell 190: 231–259. Plume O, Straub SCK, Tel-Zur N, Cisneros A, Schneider B, Doyle JJ (2013) Testing a Hypothesis of Intergeneric Allopolyploidy in Vine Cacti (Cactaceae: Hylocereeae). Syst Bot 38(3): 737–751. https://doi.org/10.1600/036364413X670421 QGIS Development Team (2016) QGIS Geographic Information System RStudio Team (2020) RStudio: Integrated Development for R. RStudio, PBC, Boston, MA http://www.rstudio.com/. Sang T, Crawford DJ, Stuessy TF (1997) Chloroplast DNA phylogeny, reticulate evolution, and biogeography of Paeonia (Paeoniaceae). Am J Bot 84(9): 1120–1136. http://www.amjbot.org/content/84/8/1120.abstract Schnable PS, Hsia AP, Nikolau BJ (1998) Genetic recombination in plants. Curr Opin Plant Biol 1(2): 123–129. https://doi.org/10.1016/S1369-5266(98)80013-7 Shaw J, Lickey EB, Schilling EE, Small RL (2007) Comparison of whole chloroplast genome sequences to choose noncoding regions for phylogenetic studies in angiosperms: the tortoise and the hare III. Am J Bot 94(3): 275–288. Slatkin M (1987) Gene Flow and the Geographic Structure of Natural Populations. Science 236(4803): 787–792. Slatkin M (1993) Isolation by Distance in Equilibrium and Non-Equilibrium Populations. Evolution 47(1): 264–279. Soltis PS, Soltis DE (2009) The role of hybridization in plant speciation. Ann Rev Plant Biol 60: 561–588. https://doi.org/10.1146/annurev.arplant.043008.092039 Taberlet P, Gielly L, Pautou G, Bouvet J (1991) Universal primers for amplification of three non-coding regions of chloroplast DNA. Plant Mol Biol 17(5): 1105–1109. http://www.ncbi.nlm.nih.gov/pubmed/1932684 Tapia HJ, Arias S, Yáñez-Espinosa L, Terrazas T (2016) El uso de espinas del tallo en la identificación de las especies de Neobuxbaumia (Cactaceae). Rev Mex Biodivers 87(2): 288–300. https://doi.org/10.1016/j.rmb.2016.04.006 Tapia HJ, Bárcenas-Argüello ML, Terrazas T, Arias S (2017) Phylogeny and Circumscription of Cephalocereus (Cactaceae) Based on Molecular and Morphological Evidence. Syst Bot 42(4): 1–15. https://doi.org/10.1600/036364417X696546 Tate JA, Simpson BB (2003) Paraphyly of Tarasa (Malvaceae) and Diverse Origins of the Polyploid Species. Syst Bot 28(4): 723–737. https://doi.org/10.1043/02-64.1 Valiente-Banuet A, Rojas-Martínez A, Arizmendi MC, Dávila P (1997) Pollination biology of two columnar cacti ( Neobuxbaumia mezcalaensis and Neobuxbaumia macrocephala ) in the Tehuacan Valley, central Mexico. Am J Bot 84(4): 452–455. Vite F, Portilla E, Zavala-hurtado JA, Valverde PL, Díaz-Solís A (1996) A natural hybrid population between Neobuxbaumia tetetzo and Cephalocereus columna-trajani (Cactaceae). Journal of Arid Environments , 32: 395–405. Additional Declarations No competing interests reported. Supplementary Files 01CONCsequence.nex DISTGENET.csv DISTGEO.csv DISTINDEX.csv Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-2481800","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":168156175,"identity":"2ab91baf-c869-4158-9ce2-0dd15e8b9d02","order_by":0,"name":"Héctor J. 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Morrone","email":"","orcid":"","institution":"Universidad Nacional Autónoma de México (UNAM)","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Juan","middleName":"J.","lastName":"Morrone","suffix":""},{"id":168156178,"identity":"d7b6dcb5-7bd4-43c7-85e4-b8484d7b4340","order_by":3,"name":"Patricia Dávila","email":"","orcid":"","institution":"UBIPRO, FES Iztacala, Universidad Nacional Autónoma de México (UNAM), Estado de México","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Patricia","middleName":"","lastName":"Dávila","suffix":""}],"badges":[],"createdAt":"2023-01-16 00:14:11","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-2481800/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-2481800/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":31774614,"identity":"c958234e-2fea-4222-92e8-197d88c97989","added_by":"auto","created_at":"2023-01-18 23:35:18","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":773595,"visible":true,"origin":"","legend":"\u003cp\u003eApproximate location of collecting sites.\u003c/p\u003e","description":"","filename":"Figure1final.png","url":"https://assets-eu.researchsquare.com/files/rs-2481800/v1/36c2cf26af1d1835ec23e671.png"},{"id":31773463,"identity":"7895423c-d8bc-43ae-8728-c0cf45646f40","added_by":"auto","created_at":"2023-01-18 23:27:18","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":183782,"visible":true,"origin":"","legend":"\u003cp\u003eObserved distributions for the genera \u003cem\u003eBergerocactus\u003c/em\u003e (★), \u003cem\u003eCephalocereus\u003c/em\u003e (+), and \u003cem\u003eMarshallocereus\u003c/em\u003e(*), and potential distributions for \u003cem\u003eBergerocactus\u003c/em\u003e and \u003cem\u003eCephalocereus\u003c/em\u003e. Three inferred barriers are indicated: climatic barrier in south Veracruz (A), geographic barrier on Transmexican Volcanic Belt (B), and climatic barrier in Baja California (C). Probabilities under 50% are discarded in the potential distribution map. Internal lines represent biogeographic provinces (Morrone et al., 2022).\u003c/p\u003e","description":"","filename":"Figure2final.png","url":"https://assets-eu.researchsquare.com/files/rs-2481800/v1/192a2deff2c942bddf509f2c.png"},{"id":31774613,"identity":"0ef8f177-a672-429b-ae0a-997cf2b9aaf0","added_by":"auto","created_at":"2023-01-18 23:35:18","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":678863,"visible":true,"origin":"","legend":"\u003cp\u003eBayesian tree (phylogram) showing main clades and branch lengths. Posterior probabilities under 1 are annotated close to the branch. Bold lines represent posterior probabilities = 1.\u003c/p\u003e","description":"","filename":"Figure3final.png","url":"https://assets-eu.researchsquare.com/files/rs-2481800/v1/e9903a94316d5d30c9571647.png"},{"id":31773471,"identity":"2cc4757b-ec2c-4c03-9778-f2a799c3f347","added_by":"auto","created_at":"2023-01-18 23:27:18","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":2090553,"visible":true,"origin":"","legend":"\u003cp\u003eHeatmap representation of pairwise distances: A) phylogenetic distances (DGEN), B) geographic distances (DGEO), and C) differentiation index (\u003cem\u003ed\u003c/em\u003e\u003csub\u003e\u003cem\u003eg\u003c/em\u003e\u003c/sub\u003e).\u003c/p\u003e","description":"","filename":"Figure4final.png","url":"https://assets-eu.researchsquare.com/files/rs-2481800/v1/7c3f04c8cd988315a328108c.png"},{"id":31773470,"identity":"0ea85005-c3d4-4c78-8343-9b0070a7d270","added_by":"auto","created_at":"2023-01-18 23:27:18","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":1949077,"visible":true,"origin":"","legend":"\u003cp\u003eCorrelation scatterplots showing distribution of pairwise distances: A) uncategorized data; B) outgroup, \u003cem\u003eMarshallocereus\u003c/em\u003e, and \u003cem\u003eBergerocactus\u003c/em\u003e; C) first differentiated clade: \u003cem\u003eC. scoparius\u003c/em\u003e, \u003cem\u003eC. apicicephalium\u003c/em\u003e, \u003cem\u003eC. nizandensis\u003c/em\u003e, and \u003cem\u003eC. totolapensis\u003c/em\u003e, D) second differentiated clade: \u003cem\u003eC. euphorbioides\u003c/em\u003e, \u003cem\u003eC. polylophus\u003c/em\u003e, and \u003cem\u003eC. parvispinus\u003c/em\u003e; E) diversified clade [partial] (1): \u003cem\u003eC. macrocephalus\u003c/em\u003e, \u003cem\u003eC. mezcalaensis\u003c/em\u003e, \u003cem\u003eC. multiareolatus\u003c/em\u003e, \u003cem\u003eC. nudus\u003c/em\u003e; F) diversified clade [partial] (2): \u003cem\u003eC. tetetzo\u003c/em\u003e, \u003cem\u003eC. columna-trajani\u003c/em\u003e, and \u003cem\u003eC. senilis\u003c/em\u003e; G) two differentiated species not strongly grouped in any clade: \u003cem\u003eC. fulviceps\u003c/em\u003e, and \u003cem\u003eC. sanchezmejoradae\u003c/em\u003e; H) intra-species distances: \u003cem\u003eC. euphorbioides\u003c/em\u003e, \u003cem\u003eC. mezcalaensis\u003c/em\u003e, \u003cem\u003eC. scoparius\u003c/em\u003e, \u003cem\u003eC. nudus\u003c/em\u003e, \u003cem\u003eC. tetetzo\u003c/em\u003e, and \u003cem\u003eNeodawsonia\u003c/em\u003e (\u003cem\u003eC. apicicephalium\u003c/em\u003e, \u003cem\u003eC. nizandensis\u003c/em\u003e, and \u003cem\u003eC. totolapensis\u003c/em\u003e).\u003c/p\u003e","description":"","filename":"Figure5final.png","url":"https://assets-eu.researchsquare.com/files/rs-2481800/v1/6fc9438212ad44c52c68a744.png"},{"id":52738316,"identity":"6d21c7ec-5090-4c47-944f-cdf3dd7e8160","added_by":"auto","created_at":"2024-03-15 07:07:55","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1425388,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-2481800/v1/32dd0c86-dbbd-4bb0-9aae-bd7a72ddbfd9.pdf"},{"id":31773464,"identity":"5b3285f6-47d7-440c-b4a1-d6d681dcc0c6","added_by":"auto","created_at":"2023-01-18 23:27:18","extension":"nex","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":172421,"visible":true,"origin":"","legend":"","description":"","filename":"01CONCsequence.nex","url":"https://assets-eu.researchsquare.com/files/rs-2481800/v1/d40277dc721d2f7352823cf0.nex"},{"id":31774615,"identity":"65ea5801-9c0b-4f1d-9b15-113ca65f518a","added_by":"auto","created_at":"2023-01-18 23:35:18","extension":"csv","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":18784,"visible":true,"origin":"","legend":"","description":"","filename":"DISTGENET.csv","url":"https://assets-eu.researchsquare.com/files/rs-2481800/v1/87e305badd78e1a09f985596.csv"},{"id":31773466,"identity":"6545231a-26e9-403c-9ca3-5ef866d9a971","added_by":"auto","created_at":"2023-01-18 23:27:18","extension":"csv","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":17496,"visible":true,"origin":"","legend":"","description":"","filename":"DISTGEO.csv","url":"https://assets-eu.researchsquare.com/files/rs-2481800/v1/193bf79edb74ce827e2c62f4.csv"},{"id":31773468,"identity":"83694169-4a8e-406d-b5e8-051e81441ad4","added_by":"auto","created_at":"2023-01-18 23:27:18","extension":"csv","order_by":4,"title":"","display":"","copyAsset":false,"role":"supplement","size":16874,"visible":true,"origin":"","legend":"","description":"","filename":"DISTINDEX.csv","url":"https://assets-eu.researchsquare.com/files/rs-2481800/v1/2e8e0fad402e99e87ef60a83.csv"}],"financialInterests":"No competing interests reported.","formattedTitle":"Phylogenetic and geographic diversification/differentiation as an evolutionary avenue in the genus Cephalocereus (Cactaceae) Evolutionary Avenue in Cephalocereus","fulltext":[{"header":"Introduction","content":"\u003cp\u003eAfter Darwin (\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e1859\u003c/span\u003e), geography has been acknowledged as a factor that affects the composition of biotas and drives to morphological change of living beings after uncountable generations in isolation have passed, as some of the most evident aspects of biological evolution. Currently, the emergence of new kinds of organisms are strongly related to spatial and reproductive isolation (allopatric speciation), which is considered an ordinary way for biological diversity to be produced and to be maintained functionally separated in distinguishable classes, in contrast to other alternative processes as sympatric speciation (Lande, \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e1980\u003c/span\u003e, De Queiroz, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2007\u003c/span\u003e; Fitzpatrick et al., \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). Must isolation be understood as an evident and sustained discontinuity in spatial or temporal distributional traits; isolation is equivalent to separation, regardless of the appearance of genetic or morphological differentiation. Then, isolation can account for genetic and morphological differentiation in a conditional manner, when genetic flow ceases and mutation and genetic drift produce noticeable among-populations changes (Slatkin, \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e1987\u003c/span\u003e). Given the independence between isolation and differentiation it is necessary to measure the actual distances in structural (morphologic or genetic) or functional (ecologic or physiologic) attributes, solely or in correlation with the spatial or temporal (time since) separation.\u003c/p\u003e \u003cp\u003eTo date, most of the evolutionary methods and related approaches have emphasized on an intrinsic inclination of living beings to drift apart from their relatives, as in population genetics, systematics, biogeography and ecology, among others (e.g. Karlsson-Green et al., \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). The cladogenetic events depicted in a branching phylogenetic diagram or a phylogenetic tree are cases of this logic, even when other evolutionary processes would also be accounted for (Lande, \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e1980\u003c/span\u003e; Schnable et al., \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e1998\u003c/span\u003e; Soltis \u0026amp; Soltis, \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). The ecological isolation and the isolation by distance are also examples of divergentist models, where spatial or ecological isolation and differentiation are expected to occur simultaneously. On the other hand, processes as hybridization and incomplete lineage sorting (ILS) reveal discrepancies in observed characters, either morphological, genetical or both (Soltis \u0026amp; Soltis, \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2009\u003c/span\u003e; Murillo et al., \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Hybridization events in plants are responsible for the appearance of a noticeable number of new species, but in the overall results insufficient to be considered a main evolutionary process. The present study advocate for an alternative point of view, where convergence may indeed be as important as divergence is to the evolutionary processes, none of them being exclusive over the other. Convergence is referred here as a syngenetic or reproductive process and is opposed to the cladogenetic or reproductive isolation process, either if it is intrinsically or extrinsically driven. This alternative approach requires the development of new methodologies or new interpretations of current methods. For example, common distance methods estimate rates of evolutionary change or change accumulation through time, between and among populations and species, including geographic and genetic distances, and populational parameters as the flow of individuals and their genes (Nei, \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e1972\u003c/span\u003e; Slatkin, \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e1993\u003c/span\u003e). The present study follows an exploratory viewpoint and is intended to quantify the amount of evolutionary change accumulated between and among species and species groups, as well as to contrast the geographic distances in a structured and relativized framework, such that the different groupings (from populations to genera) can be located and compared in a simplified graphical and statistical manner.\u003c/p\u003e \u003cp\u003eThe family Cactaceae is an outstanding model for the study of geographic and climatic isolation because it includes species highly adapted to harsh environments in arid and semiarid regions of the Americas (Bravo-Hollis, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e1978\u003c/span\u003e, Gibson \u0026amp; Nobel, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e1986\u003c/span\u003e, Anderson, \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2001\u003c/span\u003e). Several authors have related the ability to survive in these environments to structural and physiological features, as the presence of spines, reduced or absent leaves, succulent tissues, growth forms, and CAM metabolism, among others (Bravo-Hollis, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e1978\u003c/span\u003e; Anderson, \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2001\u003c/span\u003e; Hern\u0026aacute;ndez-Hern\u0026aacute;ndez et al, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). Extrinsic microhabitat features also play a key role in the establishment and survival of plantlets, as occur with rocky outcrops, cliffs, nurse plants and nurse rocks (Peters et al., \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2008\u003c/span\u003e, B\u0026aacute;rcenas-Arg\u0026uuml;ello et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2010\u003c/span\u003e, Ocampo \u0026amp; Columbus, \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2010\u003c/span\u003e, Arakaki et al., \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). Geographically, it has been recognized the influence of topographic and climatic factors limiting distributions and affecting the diversity of cactus species, as high elevations, or high precipitations, added to the seasonal and historical variation in climatic conditions (see Barthlott et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). \u003cem\u003eCephalocereus\u003c/em\u003e is endemic to Mexico and a common representative of the columnar cacti in the central and southern regions (Bravo-Hollis, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e1978\u003c/span\u003e). Much of its species\u0026rsquo; diversity occurs among the states of Jalisco, Colima, Michoac\u0026aacute;n, Guerrero, Oaxaca, Puebla, and Chiapas, within the Southeast Lowlands, Balsas Basin, and Sierra Madre del Sur biogeographic provinces. Some species are distributed northward, crossing the Transmexican Volcanic Belt, into the Sierra Madre Oriental and Veracruzan provinces, in the states of Hidalgo, Quer\u0026eacute;taro, San Luis Potos\u0026iacute;, Guanajuato, Veracruz, and Tamaulipas. Provinces are referred following Morrone (\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Morrone et al., \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Particularly, the Tehuacan-Cuicatlan valley region (Oaxaca and Puebla) has the higher species\u0026rsquo; diversity with five out of sixteen species occurring there (Bravo-Hollis, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e1978\u003c/span\u003e, Gibson \u0026amp; Nobel, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e1986\u003c/span\u003e, Anderson, \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2001\u003c/span\u003e, Hunt et al., 2006).\u003c/p\u003e \u003cp\u003eOur hypothesis is that geographic and climatic barriers have produced the isolation or separation of the \u003cem\u003eCephalocereus\u003c/em\u003e genus into phylogenetically differentiated species and species groups, while the absence of barriers in part of its distributional range has led to processes of phylogenetic diversification while developing less differentiated instead.\u003c/p\u003e"},{"header":"Material And Methods","content":"\u003cp\u003e\u003cem\u003eTaxonomic and genetic sampling.\u003c/em\u003e Taxon sampling includes 26 terminals corresponding to the 16 recognized species in the genus \u003cem\u003eCephalocereus\u003c/em\u003e, and two closely related species as outgroup: \u003cem\u003eBergerocactus emoryi\u003c/em\u003e and \u003cem\u003eMarshallocereus aragonii\u003c/em\u003e ssp. \u003cem\u003eeichlamii\u003c/em\u003e (Tapia et al., \u003cspan class=\"CitationRef\"\u003e2017\u003c/span\u003e; Arias et al., \u003cspan class=\"CitationRef\"\u003e2019\u003c/span\u003e; Arias in Korotkova et al., \u003cspan class=\"CitationRef\"\u003e2021\u003c/span\u003e); \u003cem\u003eBergerocactus\u003c/em\u003e and \u003cem\u003eMarshallocereus\u003c/em\u003e hereafter. Disjunct or widespread species’ distributions are represented with two and three terminals respectively. Collected species and distributions are detailed in Table \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e, and approximate collecting sites are shown in Fig. \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e.\u003c/p\u003e\u0026nbsp;\u003ctable border=\"1\" id=\"Tab1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eSpecies, locations, and distribution types.\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\u0026nbsp;\u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eSpecies\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eKey\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eCollect number\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eDistribution type\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\" rowspan=\"24\"\u003e\n \u003cp\u003e\u003cem\u003eCephalocereus\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. apicicephalium\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSA1257\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eRestricted\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. columna-trajani\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSA1377\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eRestricted\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. euphorbioides\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(VER)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS001\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eDisjunct\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(TAMPS)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHamman (cult.)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. fulviceps\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS029\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMedium\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. macrocephalus\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS028\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eRestricted\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. mezcalaensis\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(GRO)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS036\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eWidespread\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(MOR)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS042\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(PUE)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS044\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. multiareolatus\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS034\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eRestricted\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. nizandensis\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eTT633\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eRestricted\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. nudus\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(JAL)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS030\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eWidespread\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(OAXa)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS011\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(OAXb)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS024\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. parvispinus\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS038\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSingle site\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. polylophus\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS007\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMedium\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. sanchezmejoradae\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS039\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSingle site\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. scoparius\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(OAX)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS010\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eDisjunct\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(VER)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS002\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. senilis\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eTT529\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eRestricted\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. tetetzo\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(OAXa)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS020\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eWidespread\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(OAXb)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHJTS041\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e(PUE)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSA1376\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. totolapensis\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eTT628\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eRestricted\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eBergerocactus emoryi\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSA1307\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMedium\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eMarshallocereus eichlamii\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSA1363\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMedium\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003c/p\u003e\n\u003cp\u003e\u003cbr\u003e\u003c/p\u003e\n\u003cp\u003e\u003cbr\u003e\u003c/p\u003e\n\u003cp\u003eGenetic sampling includes 7 chloroplast loci: \u003cem\u003erpl16\u003c/em\u003e, \u003cem\u003etrnL-F\u003c/em\u003e, \u003cem\u003epsbA-trnH\u003c/em\u003e, \u003cem\u003erpl32-trnL\u003c/em\u003e, \u003cem\u003etrnQ-rps16\u003c/em\u003e, \u003cem\u003epetL-psbE\u003c/em\u003e, and \u003cem\u003eycf1\u003c/em\u003e. All the sequences have been generated and deposited previously in the GenBank (see Table \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e). PCR conditions are detailed in Tapia et al. (\u003cspan class=\"CitationRef\"\u003e2017\u003c/span\u003e). DNA sequences were aligned and trimmed, producing a slightly different matrix length in base pairs from the previous analysis, due to the absence of some indels in the actual taxa sample. Primers, sizes, and basic statistics for each region are shown in Table \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e.\u003c/p\u003e\u0026nbsp;\u003ctable border=\"1\" id=\"Tab2\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eGenBank accessions.\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eTerminal\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eAccession numbers (\u003cem\u003erpl16\u003c/em\u003e, \u003cem\u003etrnL-F\u003c/em\u003e, \u003cem\u003epsbA-trnH\u003c/em\u003e, \u003cem\u003erpl32-trnL\u003c/em\u003e, \u003cem\u003etrnQ-rps16\u003c/em\u003e, \u003cem\u003epetL-psbE\u003c/em\u003e, \u003cem\u003eycf1\u003c/em\u003e)\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. apicicephalium\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eDQ099996, DQ099927, KY624684, KY624717, KY624756, KY624787, KY624825\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. columna-trajani\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAY181599, AY181648, KY624685, KY624718, KY624757, KY624788, ND\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. euphorbioides\u003c/em\u003e (VER)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eKY624637, KY624651, KY624666, KY624699, KY624738, KY624769, KY624808\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. euphorbioides\u003c/em\u003e (TAMPS)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAY181595, AY181635, KY624667, KY624700, KY624739, KY624770, KY624809\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. fulviceps\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAY181602, AY181621, KY624683, KY624716, KY624755, KY624786, KY624824\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. mezcalaensis\u003c/em\u003e (PUE)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eKY624638, KY624652, KY624669, KY624702, KY624741, KY624772, KY624811\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. mezcalaensis\u003c/em\u003e (MOR)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eKY624639, KY624653, KY624670, KY624703, KY624742, KY624773, KY624812\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. mezcalaensis\u003c/em\u003e (GRO)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAY181600, AY181645, KY624671, KY624704, KY624743, KY624774, KY624813\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. macrocephalus\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eDQ100013, DQ099944, KY624668, KY624701, KY624740, KY624771, KY624810\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. multiareolatus\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAY181594, AY181636, KY624672, KY624705, KY624744, KY624775, KY624814\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. nizandensis\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eDQ099997, DQ099928, KY624686, KY624719, ND, KY624789, ND\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. nudus\u003c/em\u003e (JAL)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eKY624644, KY624658, KY624679, KY624712, KY624751, KY624782, KY624821\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. nudus\u003c/em\u003e (OAXa)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eKY624643, KY624657, KY624678, KY624711, KY624750, KY624781, KY624820\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. nudus\u003c/em\u003e (OAXb)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eKY624642, KY624656, KY624677, KY624710, KY624749, KY624780, KY624819\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. parvispinus\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMK165433, MK165434, MK165435, MK165436, MK165437, MK165438, MK165439\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. polylophus\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAY181597, AY181644, KY624673, KY624706, KY624745, KY624776, KY624815\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. sanchezmejoradae\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eKY624640, KY624654, KY624674, KY624707, KY624746, KY624777, KY624816\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. scoparius\u003c/em\u003e (OAX)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAY181596, AY181625, KY624675, KY624708, KY624747, KY624778, KY624817\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. scoparius\u003c/em\u003e (VER)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eKY624641, KY624655, KY624676, KY624709, KY624748, KY624779, KY624818\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. senilis\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAY181616, AY181638, KY624687, KY624720, KY624758, KY624790, KY624826\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. tetetzo\u003c/em\u003e (PUE)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAY181592, AY181632, KY624680, KY624713, KY624752, KY624783, ND\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. tetetzo\u003c/em\u003e (OAXa)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eKY624645, KY624659, KY624681, KY624714, KY624753, KY624784, KY624822\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. tetetzo\u003c/em\u003e (OAXb)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eKY624646, KY624660, KY624682, KY624715, KY624754, KY624785, KY624823\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eC. totolapensis\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eKY624647, KY624661, KY624688, KY624721, KY624759, KY624791, KY624827\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eB. emoryi\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eDQ099994, DQ099925, KF783478, KY624730, KF783697, KY624800, KY624834\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eM. eichlamii\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAY181610, AY181629, KY624690, KY624723, KY624760, KY624793, KY624828\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003c/p\u003e\n\u003cp\u003e\u003c/p\u003e\u0026nbsp;\u003ctable border=\"1\" id=\"Tab3\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003ePlastid regions employed, and primer sequences.\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eRegion\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\u0026nbsp;\u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eSequence\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eExpected length\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eReference\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003erpl16\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eF(1F)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eGCT ATG CTT AGT GTG TGA CTC GTT\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e900–1,200\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eHernández-Hernández et al. \u003cspan class=\"CitationRef\"\u003e2011\u003c/span\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eR(3R)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCTT CTA TTT GTC TAG GCG TGA TCC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003etrnL-F\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eF€\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCGA AAT CGG TAG ACG CTA CG\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e700–1,100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eTaberlet et al. \u003cspan class=\"CitationRef\"\u003e1991\u003c/span\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eR(f)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eATT TGA ACT GGT GAC ACG AG\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eF-int€\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eGGT TCA AGT CCC TCT ATC CC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eR-int(d)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eGGG GAT AGA GGG ACT TGA AC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003epsbA-trnH\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eF(psbA)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eGTT ATG CAT GAA CGT AAT C\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e300–700\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSang et al. \u003cspan class=\"CitationRef\"\u003e1997\u003c/span\u003e\u003c/p\u003e\n \u003cp\u003eTate and Simpson \u003cspan class=\"CitationRef\"\u003e2003\u003c/span\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eR(trnH2)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCGC GCA TGG TGG ATT CAC A\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003erpl32-trnL\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eF(rpl32)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCAG TTC CAA AAA AAC GTA CTT C\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e500–1,400\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eShaw et al. \u003cspan class=\"CitationRef\"\u003e2007\u003c/span\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eR(trnL\u003csup\u003eUAG\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCTG CTT CCT AAG AGC AGC GT\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eF-int\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eGTA ACT CTT GAA ATC ATT ATT TC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ePlume et al. \u003cspan class=\"CitationRef\"\u003e2013\u003c/span\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eR-int\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eGTT ATC TTA GGT TTC AAC AAA CC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003etrnQ-rps16\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eF(trnQ\u003csup\u003eUUG\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eGCG TGG CCA AGY GGT AAG GC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e600–1,500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eShaw et al. \u003cspan class=\"CitationRef\"\u003e2007\u003c/span\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eR(rps16\u003csup\u003eXI\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eGTT GCT TTY TAC CAC ATC GTT T\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003epetL-psbE\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eF(petL)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAGT AGA AAA CCG AAA TAA CTA GTT A\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e400–700\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eShaw et al. \u003cspan class=\"CitationRef\"\u003e2007\u003c/span\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eR(psbE)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eTAT CGA ATA CTG GTA ATA ATA TCA GC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eycf1\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eF(4182)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAAA TAY RRA TAG AAA ATA TTT KGA TT\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e900–1,100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eFranck et al. \u003cspan class=\"CitationRef\"\u003e2012\u003c/span\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eR(5248)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eGAA TTC TYA ATT CTC TAC GAC G\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003ePhylogenetic analysis.\u003c/em\u003e A Bayesian Inference analysis was conducted in MrBayes 3.2.6 (Huelsenbeck \u0026amp; Ronquist, \u003cspan class=\"CitationRef\"\u003e2001\u003c/span\u003e), using the GTR + I + G model, during 10’000’000 generations, without matrix partitions due to their same-organelle origin, it was applied a burnin fraction = 0.5. Statistics for trees and probabilities were summarized to visualize topologies, convergence of chains, branch lengths, and posterior probabilities. The consensus Bayesian tree with branch lengths and posterior probabilities appended was rooted using the outgroup, exported in newick format, and retained for further analyses.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eDistance analyses.\u003c/em\u003e A phylogenetic pairwise distance matrix (DGEN) was computed from the Bayesian consensus tree, imported, and analyzed in R environment (RStudio 3.6.3, 2020), using the ‘ape’ library. The resulting table (26x26) contains two symmetrical triangles and a diagonal with only-zero values that can be reduced to 325 single comparison values. This table was then transformed into relative values (positives from ~ 0 to 1), dividing each cell by the maximum value in the table. Geographic pairwise distance matrix (DGEO) was computed in Qgis software (2016; ver. 3.22), using geographic data (latitude and longitude) for the same 26 terminals included in the phylogenetic analysis. Linear Geographic distance was computed resulting in an identical data structure. These values were standardized and reduced in the same way as for phylogenetic distance matrix. Absolute and relative values for both datasets are provided as supplemental data.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eSpearman’s correlation and Mantel test.\u003c/em\u003e Phylogenetic distances were tested for spatial correlation against geographic distances, to evaluate the existence of deviations from null hypothesis. These analyses were implemented in Excel (tm) using the complement GenAlEx6.5 (Peakall \u0026amp; Smouse, \u003cspan class=\"CitationRef\"\u003e2012\u003c/span\u003e). Non-parametric correlation analysis was performed using Spearman’s coefficient and a Mantel test with the maximum allowed permutations (9’999).\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eClimatic barriers.\u003c/em\u003e Geographic data from samples were integrated into a larger geolocation dataset. This matrix contains 423 records for species from the three genera, 407 of them corresponding to \u003cem\u003eCephalocereus\u003c/em\u003e. At the generic level, all the records were considered, but at species’ level some records were discarded from species with restricted distributions or species known from a single locality. The species not considered were \u003cem\u003eC. apicicephalium\u003c/em\u003e, \u003cem\u003eC. nizandensis\u003c/em\u003e, \u003cem\u003eC. parvispinus\u003c/em\u003e, \u003cem\u003eC. sanchezmejoradae\u003c/em\u003e, \u003cem\u003eC. senilis\u003c/em\u003e, and \u003cem\u003eMarshallocereus\u003c/em\u003e. Climatic models were performed using MaxEnt 3.4.4 (Phillips et al., \u003cspan class=\"CitationRef\"\u003e2006\u003c/span\u003e), generating potential distribution maps to identify climatic barriers and environmental factors limiting species distributions.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eDistancing index (d).\u003c/em\u003e Both datasets, phylogenetic and geographic, were verified as Euclidean values, then relativized, and identically pairwise organized. A combined distance measure was implemented using standardized phylogenetic and geographic distance matrices between pairs of terminals, as follows:\u003c/p\u003e\n\u003cdiv class=\"Equation\" id=\"Equa\"\u003e\n \u003cdiv class=\"mathdisplay\" id=\"FileID_Equa\" name=\"EquationSource\"\u003e$$d=\\sqrt{{p}^{2}+{g}^{2}}$$\u003c/div\u003e\n\u003c/div\u003e\n\u003cp\u003e\u003c/p\u003e\n\u003cp\u003ewhere \u003cem\u003ep\u003c/em\u003e and \u003cem\u003eg\u003c/em\u003e represent respectively the phylogenetic and geographic distances between the same pair of terminals. The limit of this index is \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\sqrt{2}\\)\u003c/span\u003e\u003c/span\u003e (≈ 1.4142).\u003c/p\u003e\n\u003cp\u003e\u003c/p\u003e\n\u003cp\u003eThe following assumptions were taken into account to interpret the \u003cem\u003edistancing index\u003c/em\u003e: a) phylogenetic distance is indicative of the time since two lineages split up, b) distancing index can be translated as a probability of reproductive isolation between two lineages, c) differentiated lineages reach the higher distance values in both dimensions, d) non-differentiated lineages will have lower distance values in one dimension, phylogenetic or geographic, and could be interpreted as diversified groups, and e) existence of a geographic or climatic barrier is crucial to decide whether or not exists a differentiation or a diversification process. Given that assumptions, a decision matrix was constructed to evaluate the results as follows:\u003c/p\u003e\n\u003cp\u003e\u003cimg src=\"data:image/png;base64,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\"\u003e\u003cbr\u003e\u003c/p\u003e\n\u003cp\u003eWhere higher distance values (+ +) indicate a complete differentiation process, combination of low and high distance values (- +, + -) indicate diversification processes, and both lower values (- -) indicate an identity process, or a continuous reproductive system. Distance and diversification/differentiation matrices were represented as heatmaps as a visual guide to identify multiple simultaneous relationships.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e \u003cem\u003eGeographic and climatic barriers.\u003c/em\u003e Three main barriers were inferred from species\u0026rsquo; distributions and climatic models, not only as unoccupied areas but also as areas with a factor accountable for impeding the establishment of \u003cem\u003eCephalocereus\u003c/em\u003e, \u003cem\u003eBergerocactus\u003c/em\u003e or \u003cem\u003eMarshallocereus\u003c/em\u003e species. The first climatic barrier is in southeastern Mexico, as a stripe broader than 80 km running from southern Veracruz towards the Caribbean Sea, along the Gulf of M\u0026eacute;xico Lowlands province, and separating the Yucat\u0026aacute;n peninsula from the continent. The model for all the \u003cem\u003eCephalocereus\u003c/em\u003e species exhibits a pattern of avoidance related to temperature and moisture factors: temperature annual range (bio7), precipitation of coldest quarter (bio19), and mean temperature of warmest quarter (bio10). The second climatic barrier is in northwestern Mexico, between Baja California peninsula and the continent, separating \u003cem\u003eBergerocactus\u003c/em\u003e from all other species. The climatic variables are also related to temperature and moisture factors: mean temperature of warmest quarter (bio10), precipitation of coldest quarter (bio19) and precipitation of warmest quarter (bio18). Statistics for these potential distribution models are presented in Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e4\u003c/span\u003e, following the same abbreviations as in text.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab4\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 4\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eClimatic and geographic barriers, main variables, and contributions.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eBarrier\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eType\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eVariable\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePercent contribution\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003ePermutation importance\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSouth Veracruzan\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eClimatic\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ebio10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e29.4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.4\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ebio19\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e27.1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e10.2\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ebio18\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e21.1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1.8\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ebio17\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e11.8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e76.7\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTransmexican Volcanic Belt\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGreographic\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eBaja-californian\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eClimatic\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ebio7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e56.4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e19.8\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ebio19\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e12.8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e11.8\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ebio15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e11.7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e18\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eThe third barrier is located along the Transmexican Volcanic Belt and is explained geographically, possibly related to altitudinal or topographical constrains, given that modelled and observed distributions did not match in this region. Observed and modelled distributions for the genera \u003cem\u003eCephalocereus\u003c/em\u003e and \u003cem\u003eBergerocactus\u003c/em\u003e, as well as climatic and geographic barriers are shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e. The species\u0026rsquo; distribution models are mostly compatible with the generic models (data not shown).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cem\u003eTree topology.\u003c/em\u003e The Bayesian tree shows high support values in most of the internal nodes. This tree is represented as a phylogram to show the correspondence with the phylogenetic distance matrix (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). Two clades derived basally: the first including the species \u003cem\u003eC. scoparius\u003c/em\u003e, \u003cem\u003eC. apicicephalium\u003c/em\u003e, \u003cem\u003eC. nizandensis\u003c/em\u003e, and \u003cem\u003eC. totolapensis\u003c/em\u003e, and the second including \u003cem\u003eC. parvispinus\u003c/em\u003e, \u003cem\u003eC. euphorbioides\u003c/em\u003e, and \u003cem\u003eC. polylophus\u003c/em\u003e. Posterior probabilities for the first clade are low, although it is spatially congruent. Two species, \u003cem\u003eC. fulviceps\u003c/em\u003e, and \u003cem\u003eC. sanchezmejoradae\u003c/em\u003e, appear as sister taxa of a major clade, which includes the most variable and widespread species \u003cem\u003eC. mezcalaensis\u003c/em\u003e, \u003cem\u003eC. nudus\u003c/em\u003e, and \u003cem\u003eC. tetetzo\u003c/em\u003e, along with the restricted species \u003cem\u003eC. macrocephalus\u003c/em\u003e, \u003cem\u003eC. multiareolatus\u003c/em\u003e, \u003cem\u003eC. columna-trajani\u003c/em\u003e, and \u003cem\u003eC. senilis\u003c/em\u003e. Species within this clade occupy an area extended from the Sierra Norte de Oaxaca to the Sierra Madre del Sur, the Balsas Basin, and the Pacific Lowlands, except for \u003cem\u003eC. senilis\u003c/em\u003e, which inhabits the Sierra Madre Oriental, at the north of the Transmexican Volcanic Belt.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cem\u003eDistancing Index and Differentiation.\u003c/em\u003e Spatial and temporal dimensions are presented in a relative scale (0\u0026ndash;1, or 0\u0026ndash;100%) and provide a framework to explain relationships from both phylogenetic and geographic variation. Samples at species or under species ranks exhibit relative distances below 10% in phylogenetic distances. In this segment are included the species \u003cem\u003eC. mezcalaensis\u003c/em\u003e, \u003cem\u003eC. nudus\u003c/em\u003e, \u003cem\u003eC. tetetzo\u003c/em\u003e, \u003cem\u003eC. euphorbioides\u003c/em\u003e, and \u003cem\u003eC. scoparius\u003c/em\u003e, which exhibit widespread or disjunct distributions (see Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Three more species, \u003cem\u003eC. apicicephalium\u003c/em\u003e, \u003cem\u003eC. nizandensis\u003c/em\u003e, and \u003cem\u003eC. totolapensis\u003c/em\u003e (\u003cem\u003eNeodawsonia\u003c/em\u003e clade), show distances standing in the same range of the intraspecific measures. Some differentiated species and species groups are distinguishable, as \u003cem\u003eC. scoparius\u003c/em\u003e and the clade formed by \u003cem\u003eC. parvispinus\u003c/em\u003e, \u003cem\u003eC. euphorbioides\u003c/em\u003e, and \u003cem\u003eC. polylophus\u003c/em\u003e. On the counterpart, a group of close or less differentiated species includes \u003cem\u003eC. mezcalaensis\u003c/em\u003e, \u003cem\u003eC. multiareolatus\u003c/em\u003e, \u003cem\u003eC. nudus\u003c/em\u003e, \u003cem\u003eC. macrocephalus\u003c/em\u003e, \u003cem\u003eC. tetetzo\u003c/em\u003e, \u003cem\u003eC. columna-trajani\u003c/em\u003e, and \u003cem\u003eC. senilis\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eA).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe geographic distances alone show less structuration than phylogenetic distances. Climatic barriers are responsible for the spatial separation between genera at a higher spatial scale, while the geographic barrier appears to have a moderate influence on geographic distances between \u003cem\u003eCephalocereus\u003c/em\u003e species. The species occurring northward of the Transmexican Volcanic Belt show geographic distances from 11.4% in \u003cem\u003eC. columna-trajani\u003c/em\u003e/\u003cem\u003eC. senilis\u003c/em\u003e, to 16.8% in \u003cem\u003eC. parvispinus\u003c/em\u003e/\u003cem\u003eC. euphorbioides\u003c/em\u003e, and 15% in \u003cem\u003eC. parvispinus\u003c/em\u003e/\u003cem\u003eC. polylophus\u003c/em\u003e, respectively, and one locality of \u003cem\u003eC. nudus\u003c/em\u003e at 35.3% distance collected far in western Jalisco (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eB), indicating a possible isolation from other species\u0026rsquo; populations.\u003c/p\u003e \u003cp\u003eThe distancing index (\u003cem\u003ed\u003c/em\u003e\u003csub\u003e\u003cem\u003eg\u003c/em\u003e\u003c/sub\u003e) reveals a normalizing effect, tending to the phylogenetic data. The only major change compared to phylogenetic distances alone is an increment in the average distance from \u003cem\u003eC. nudus\u003c/em\u003e (Jalisco) (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eC). No other significative change from the phylogenetic distance matrix is evident.\u003c/p\u003e \u003cp\u003e \u003cstrong\u003eDifferentiation patterns\u003c/strong\u003e \u003cp\u003eThe estimated slope (y\u0026thinsp;=\u0026thinsp;0.5084x \u0026minus;\u0026thinsp;0.0059) and the coefficient of determination from Spearman\u0026rsquo;s correlation analysis (R\u003csup\u003e2\u003c/sup\u003e\u0026thinsp;=\u0026thinsp;0.2682) indicate a significant deviation between phylogenetic and geographic distances. The scatterplot from correlation analysis shows three clearly separated groups, representing the genera \u003cem\u003eBergerocactus\u003c/em\u003e, \u003cem\u003eCephalocereus\u003c/em\u003e, and \u003cem\u003eMarshallocereus\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eA). Genus \u003cem\u003eMarshallocereus\u003c/em\u003e is separated in the extreme of the phylogenetic distance axis, while \u003cem\u003eBergerocactus\u003c/em\u003e is separated in the geographic distance axis (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eB). The dots representing \u003cem\u003eCephalocereus\u003c/em\u003e distances appear almost randomly distributed in the midst and lower values for both axes, but some groups can be detected in between. Patterns of phylogenetic differentiation are evident in two clades; the first formed by \u003cem\u003eC. scoparius\u003c/em\u003e, \u003cem\u003eC. apicicephalium\u003c/em\u003e, \u003cem\u003eC. nizandensis\u003c/em\u003e, and \u003cem\u003eC. totolapensis\u003c/em\u003e, (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eC), and the second formed by \u003cem\u003eC. euphorbioides\u003c/em\u003e, \u003cem\u003eC. polylophus\u003c/em\u003e, and \u003cem\u003eC. parvispinus\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eD). On the other hand, the clade containing \u003cem\u003eC. mezcalaensis\u003c/em\u003e, \u003cem\u003eC. multiareolatus\u003c/em\u003e, \u003cem\u003eC. nudus\u003c/em\u003e, \u003cem\u003eC. macrocephalus\u003c/em\u003e, \u003cem\u003eC. tetetzo\u003c/em\u003e, \u003cem\u003eC. columna-trajani\u003c/em\u003e, and \u003cem\u003eC. senilis\u003c/em\u003e, show less phylogenetic differentiation and proportionally major geographic distances (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eE, F). The two remaining species, \u003cem\u003eC. fulviceps\u003c/em\u003e and \u003cem\u003eC. sanchezmejoradae\u003c/em\u003e, have low affinity for any other clade or species, also exhibiting a pattern of phylogenetic differentiation (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eG). Lastly, the intra-species distances are presented for species with two or more representatives, and for the \u003cem\u003eNeodawsonia\u003c/em\u003e clade, given the short distances between these species in both dimensions, phylogenetic and geographic (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eH).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003e \u003cem\u003ePhylogenetic distancing\u003c/em\u003e. The most distant species were \u003cem\u003eC. polylophus\u003c/em\u003e and \u003cem\u003eM. aragonii\u003c/em\u003e ssp. \u003cem\u003eeichlamii\u003c/em\u003e, which was taken as the intergeneric distance (100%). This distance was 31 times greater than that between the sister species \u003cem\u003eC. apicicephalium\u003c/em\u003e and \u003cem\u003eC. nizandensis\u003c/em\u003e. The distance between \u003cem\u003eM. aragonii\u003c/em\u003e ssp. \u003cem\u003eeichlamii\u003c/em\u003e and \u003cem\u003eB. emoryi\u003c/em\u003e is 56.9%, while the most distant species within \u003cem\u003eCephalocereus\u003c/em\u003e lies in a 58.5% distance (\u003cem\u003eC. polylophus\u003c/em\u003e/\u003cem\u003eC. sanchezmejoradae\u003c/em\u003e). Other relevant comparisons are the distances among \u003cem\u003eNeodawsonia\u003c/em\u003e species which barely reach 3.2% in \u003cem\u003eC. apicicephalium\u003c/em\u003e/\u003cem\u003eC. nizandensis\u003c/em\u003e, 4% in \u003cem\u003eC. apicicephalium\u003c/em\u003e/\u003cem\u003eC. totolapensis\u003c/em\u003e, and 5.4% in \u003cem\u003eC. nizandensis\u003c/em\u003e/\u003cem\u003eC. totolapensis\u003c/em\u003e. The intraspecific distances range from 4.3% in the disjunct \u003cem\u003eC. euphorbioides\u003c/em\u003e to 9.1% in the widespread \u003cem\u003eC. mezcalaensis\u003c/em\u003e, while the species \u003cem\u003eC. columna-trajani\u003c/em\u003e and \u003cem\u003eC. tetetzo\u003c/em\u003e, which present natural hybrids (Vite et al., \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e1996\u003c/span\u003e), lie in a 13.4% distance range. These results indicate not only the recentness of the \u003cem\u003eNeodawsonia\u003c/em\u003e splitting, but the potential for other phylogenetically and geographically close species to produce genetic transferences or interchanges. Other examples are \u003cem\u003eC. scoparius\u003c/em\u003e and \u003cem\u003eC. nizandensis\u003c/em\u003e, separated barely 16 km away with a moderate phylogenetic distance from 25.9 to 28.8%, \u003cem\u003eC. tetetzo\u003c/em\u003e and \u003cem\u003eC. nudus\u003c/em\u003e, separated 25 km in a phylogenetic distance range from 15.9 to 20.4%, and \u003cem\u003eC. macrocephalus\u003c/em\u003e and \u003cem\u003eC. tetetzo\u003c/em\u003e, that make contact in one site and have phylogenetic distances from 11.5 to 13.1%. In this population of \u003cem\u003eC. macrocephalus\u003c/em\u003e and \u003cem\u003eC. tetetzo\u003c/em\u003e some putative hybrid individuals have been observed (Arias \u0026amp; Tapia, pers. obs.).\u003c/p\u003e \u003cp\u003eUp to date, several intergeneric hybrids have been reported in the tribe Echinocereeae, where \u003cem\u003eCephalocereus\u003c/em\u003e belongs, as \u0026times;\u003cem\u003ePachebergia\u003c/em\u003e (\u003cem\u003eBackebergia\u003c/em\u003e \u0026times; \u003cem\u003ePachycereus\u003c/em\u003e), \u0026times;\u003cem\u003ePacherocactus\u003c/em\u003e (\u003cem\u003eBergerocactus\u003c/em\u003e \u0026times; \u003cem\u003ePachycereus\u003c/em\u003e), \u0026times;\u003cem\u003eMyrtgerocactus\u003c/em\u003e (\u003cem\u003eBergerocactus\u003c/em\u003e \u0026times; \u003cem\u003eMyrtillocactus\u003c/em\u003e), \u0026times;\u003cem\u003eMyrtillenocereus\u003c/em\u003e (\u003cem\u003eMyrtillocactus\u003c/em\u003e \u0026times; \u003cem\u003eStenocereus\u003c/em\u003e), and some interspecific hybrids are documented in the sister tribe Hylocereeae, between \u003cem\u003eHylocereus undatus\u003c/em\u003e, \u003cem\u003eH. monacanthus\u003c/em\u003e, and \u003cem\u003eH. megalanthus\u003c/em\u003e, where \u003cem\u003eH. megalanthus\u003c/em\u003e probably speciate from same species parentals by autopolyploidy (Anderson, \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2001\u003c/span\u003e; Arias \u0026amp; Terrazas, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2008\u003c/span\u003e; Plume et al., \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2013\u003c/span\u003e; Granados-Aguilar et al., \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). The placement of hybridizing species in the same genus or in separated genera based on their morphology have implications on what we suppose to occur at cellular and genetic levels in such hybridization event, whereas morphology alone can be an overestimated indicator of differentiation and reproductive isolation (Plume et al., \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). The hybrid between \u003cem\u003eC. columna-trajani\u003c/em\u003e and \u003cem\u003eC. tetetzo\u003c/em\u003e was described initially as an intergeneric hybrid between \u003cem\u003eNeobuxbaumia tetetzo\u003c/em\u003e and \u003cem\u003eCephalocereus columna-trajani\u003c/em\u003e, later recovered as sister species in a phylogenetic analysis, along with \u003cem\u003eC. senilis\u003c/em\u003e, and reassigned altogether in the same genus (\u003cem\u003eCephalocereus\u003c/em\u003e) (Tapia et al., \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Other example is \u003cem\u003eBackebergia militaris\u003c/em\u003e, which presented enough morphological characters to be assigned to a monotypic genus according to Bravo-Hollis (\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e1953\u003c/span\u003e), but phylogenetic studies resolved its position close to \u003cem\u003ePachycereus pecten-aboriginum\u003c/em\u003e, with which hybridize to produce x\u003cem\u003ePachebergia apicostata\u003c/em\u003e (Arias \u0026amp; Terrazas, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2008\u003c/span\u003e). The Cactaceae family, like other plant families, appears as a group inclined to hybridize at varying levels, since the same species, to interspecies, to intergeneric parentals, resulting in chaotic complexes as in wild opuntias, or in flashy cultivated varieties of \u003cem\u003eAstrophytum\u003c/em\u003e (Granados-Aguilar et al., \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2022\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eAttending to a morphometric assessment, the study on \u003cem\u003eCephalocereus\u003c/em\u003e areoles and spines revealed simultaneous patterns of character variation and conservation (Tapia et al., \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). In some species a unique spination pattern allowed the univocal distinction of individuals as part of one species, as in \u003cem\u003eC. polylophus\u003c/em\u003e, C. \u003cem\u003eeuphorbioides\u003c/em\u003e, and \u003cem\u003eC. parvispinus\u003c/em\u003e, which correspond to a phylogenetically differentiated group in the present study. Other individuals show similarities that allowed the distinction of species groups, particularly the groups \u003cem\u003eC. mezcalaensis\u003c/em\u003e/\u003cem\u003eC. nudus\u003c/em\u003e/\u003cem\u003eC. scoparius\u003c/em\u003e and \u003cem\u003eC. tetetzo\u003c/em\u003e/\u003cem\u003eC. macrocephalus\u003c/em\u003e/\u003cem\u003eC. fulviceps\u003c/em\u003e are of particular interest due to the closeness between the included species, both in geographic and phylogenetic distances, and the possibility of genetic transferences or interchanges to occur, which needs to be studied further in detail. It is not intended to point that hybridization is responsible for all the evolutionary changes, but reproductive processes, either continuous or occasional, need to be accounted and comprehensible analyzed to untangle the genetic or genomic contribution of parental organisms, either if they belong to the same or to different species.\u003c/p\u003e \u003cp\u003e \u003cem\u003eGeographic barriers and splitting events\u003c/em\u003e. The effects of the climatic and geographic barriers between and among \u003cem\u003eCephalocereus\u003c/em\u003e species are consistent with a hypothesis of reproductive isolation. The clade formed by \u003cem\u003eC. scoparius\u003c/em\u003e, \u003cem\u003eC. apicicephalium\u003c/em\u003e, \u003cem\u003eC. nizandensis\u003c/em\u003e, and \u003cem\u003eC. totolapensis\u003c/em\u003e, which inhabit a region between the Tehuantepec Isthmus and the adjacent part of the Sierra Madre del Sur, represents an early divergence from the \u003cem\u003eMarshallocereus\u003c/em\u003e/\u003cem\u003eCephalocereus\u003c/em\u003e ancestor, just followed by the \u003cem\u003eBergerocactus\u003c/em\u003e ancestor splitting. The species \u003cem\u003eC. scoparius\u003c/em\u003e has a disjunct distribution occupying also a portion in the center of the Veracruzan province, caused probably by a relatively recent long distance dispersal event, denoted by a phylogenetic distance of 8.8%. The clade formed by \u003cem\u003eC. euphorbioides\u003c/em\u003e, \u003cem\u003eC. polylophus\u003c/em\u003e, and \u003cem\u003eC. parvispinus\u003c/em\u003e becomes the next splitting event coupled with the dispersal northward into the Sierra Madre del Sur and then followed by the crossing of the Transmexican Volcanic Belt barrier. \u003cem\u003eC. parvispinus\u003c/em\u003e remained in the Sierra Madre del Sur, probably ecologically isolated, and \u003cem\u003eC. polylophus\u003c/em\u003e and \u003cem\u003eC. euphorbioides\u003c/em\u003e ancestor colonized the Sierra Madre Oriental and the Veracruzan provinces. The phylogenetic distance between \u003cem\u003eC. polylophus\u003c/em\u003e and \u003cem\u003eC. euphorbioides\u003c/em\u003e is 20.3%, indicating an early divergence event between these species after their dispersal from the southern regions.\u003c/p\u003e \u003cp\u003eOn the counterpart, species non-isolated by hard or absolute barriers, show shorter distances either phylogenetic, geographic or both. This is the case of the clade formed by \u003cem\u003eC. mezcalaensis\u003c/em\u003e, \u003cem\u003eC. multiareolatus\u003c/em\u003e, \u003cem\u003eC. nudus\u003c/em\u003e, \u003cem\u003eC. macrocephalus\u003c/em\u003e, \u003cem\u003eC. tetetzo\u003c/em\u003e, \u003cem\u003eC. columna-trajani\u003c/em\u003e, and \u003cem\u003eC. senilis\u003c/em\u003e. The only species from this clade that dispersed across the Transmexican Volcanic Belt is \u003cem\u003eC. senilis\u003c/em\u003e. The phylogenetic distances among species of this clade ranges from 7.7% in the sister species \u003cem\u003eC. columna-trajani\u003c/em\u003e/\u003cem\u003eC. senilis\u003c/em\u003e, to 21.9% in \u003cem\u003eC. senilis\u003c/em\u003e/\u003cem\u003eC. nudus\u003c/em\u003e, while the geographic distances range from zero up to ~\u0026thinsp;1\u0026rsquo;000 km. The species \u003cem\u003eC. fulviceps\u003c/em\u003e and \u003cem\u003eC. sanchezmejoradae\u003c/em\u003e have phylogenetic distances higher than 28% from the species belonging to the previous clade, despite the absence of a hard barrier, indicating the possible existence of ecological ancient barriers instead.\u003c/p\u003e \u003cp\u003e \u003cem\u003eAbsolute geographic distances\u003c/em\u003e. Considering the size of \u003cem\u003eCephalocereus\u003c/em\u003e individuals and the type of pollinators (moths and bats), the distance for effective pollination ranges from meters up to several kilometers. The seeds display patterns of local dispersion led by fruit dehiscence and gravity action, or long-distance dispersal by migrating bats and birds, also up to several kilometers, and longer than in pollination (Valiente-Banuet et al., \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e1997\u003c/span\u003e; Gonz\u0026aacute;lez-Terrazas et al., \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). Phylogenetically differentiated clades and/or species are separated from other species or clades by barriers at least 80 kilometers wide or in altitudinal ranges of several hundreds of meters. In the absence of barriers, some species can occupy land stripes up to 800 km in length, like \u003cem\u003eC. nudus\u003c/em\u003e which inhabits along the Pacific coastline in southern Mexico, from Jalisco and Colima to Oaxaca. In this case, the occupation of a broad region might had occurred gradually, pollination may be restricted to each population and seed dispersal limited to close populations. Given this structuration, it is likely that the most distant populations located in Colima and Jalisco are currently undergoing a differentiation process from same species populations in Oaxaca (\u0026gt;\u0026thinsp;600 km apart). Other species with disjunct distributions (\u003cem\u003eC. euphorbioides\u003c/em\u003e and \u003cem\u003eC. scoparius\u003c/em\u003e), show moderately to low phylogenetic distancing indicating recent dispersal events around 300 or 400 km away from their original distributions, probably driven by migrating birds or bats, and have become genetically isolated relatively few generations ago.\u003c/p\u003e \u003cp\u003eOther component not-explicitly analyzed here is the populational genome, which act as a repository of evolutionary variance, where the genetic variants are gained and lost. Populational genome is involved in a third evolutionary process, the anagenesis (Emerson \u0026amp; Pati\u0026ntilde;o, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2018\u003c/span\u003e), which is related to a slow and steady evolutionary phase where the main changes correspond to gene recombination and allelic fluctuation (Schnable et al., \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e1998\u003c/span\u003e), due to both determined and stochastic processes, as gene drift and selective factors, and require biparental and more variable molecular markers to be analyzed.\u003c/p\u003e"},{"header":"Conclusions And Remarks","content":"\u003cp\u003eThis study was focused on the phylogenetic and geographic variation patterns, which concurrently account for the spatial and temporal isolation components. These components are conditionals that may or may not produce a noticeable differentiation in the resulting lineages, in morphological or genetical attributes, but if phylogenetic differentiation occurs it is always an indicative of spatial and temporal isolation (time since). Differentiation is related to the absence of reproduction among groups of individuals, while diversification is likely related to the added genetic variance from recent reproduction events between increasingly broader groups, in terms of number of individuals or occupied space, either if reproduction occurs in a continuous or in an occasional fashion. It must be noted that spatial barriers to reproduction are specific to each taxonomic group and need to be evaluated in their own ecological context.\u003c/p\u003e \u003cp\u003ePractical applications of this type of analysis are currently under development and could serve to validate new species, to define significant spatial barriers to reproduction in evolutionary time, to distinguish and define species complexes where phylogenetic methods could not resolve the sisterhood of species, to assess the evolutionary trends (syngenetic or cladogenetic) in a taxonomic group, among others.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe first author thanks the Direcci\u0026oacute;n General de Asuntos del Personal Ac\u0026aacute;demico (DGAPA) for the postdoctoral fellowship granted to carry out this research in the Laboratorio de Recursos Naturales, UBIPRO, FES Iztacala, Universidad Nacional Aut\u0026oacute;noma de M\u0026eacute;xico (UNAM). All authors thank Oswaldo T\u0026eacute;llez and Gabriel Arroyo-Cosultchi for sharing some \u003cem\u003eCephalocereus\u003c/em\u003e geolocation records.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors have no competing interests to declare that are relevant to the content of this article.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eAnderson EF (2001) \u003cem\u003eThe Cactus Family\u003c/em\u003e, USA: Timber Press, 776 pp.\u003c/li\u003e\n\u003cli\u003eArakaki M, Christin PA, Nyffeler R, Lendel A, Eggli U, Ogburn RM, Spriggs E, Moore MJ, Edwards EJ (2011) Contemporaneous and recent radiations of the world\u0026rsquo;s major succulent plant lineages. \u003cem\u003eProc Natl Acad Sci U S A\u003c/em\u003e 108: 8379\u0026ndash;8384. https://doi.org/10.1073/pnas.1100628108\u003c/li\u003e\n\u003cli\u003eArias S, Tapia HJ, Guzm\u0026aacute;n U (2019) A new species of \u003cem\u003eCephalocereus\u003c/em\u003e (Cactaceae) from southern Mexico. \u003cem\u003ePhytotaxa\u003c/em\u003e 392(2): 147\u0026ndash;156. https://doi.org/10.11646/phytotaxa.392.2.4\u003c/li\u003e\n\u003cli\u003eArias S, Terrazas T (2008) x\u003cem\u003ePachebergia\u003c/em\u003e (Cactaceae), a nothogenus from western Mexico. \u003cem\u003eRev Mex Biodivers\u003c/em\u003e 79(1): 23\u0026ndash;28.\u003c/li\u003e\n\u003cli\u003eB\u0026aacute;rcenas-Arg\u0026uuml;ello ML, Guti\u0026eacute;rrez-Castorena MC, Terrazas T, L\u0026oacute;pez-Mata L (2010) Rock\u0026ndash;soil preferences of three \u003cem\u003eCephalocereus\u003c/em\u003e (Cactaceae) species of tropical dry forests. \u003cem\u003eSoil Sci Soc Am J\u003c/em\u003e 74: 1374\u0026ndash;1382. https://doi.org/10.2136/sssaj2009.0310\u003c/li\u003e\n\u003cli\u003eBarthlott W, Burstedde K, Geffert JL, Ibisch PL, Korotkova N, Miebach A, Rafiqpoor MD, Stein A, Mutke J (2015) Biogeography and biodiversity of Cacti. \u003cem\u003eSchumannia\u003c/em\u003e \u003cem\u003e7\u003c/em\u003e. 205pp\u003c/li\u003e\n\u003cli\u003eBravo-Hollis H (1953) Un nuevo g\u0026eacute;nero de la Familia de las Cact\u0026aacute;ceas \u003cem\u003eBackebergia\u003c/em\u003e. \u003cem\u003eAn Inst Bio\u003c/em\u003e \u003cem\u003eUNAM\u003c/em\u003e 24: 215\u0026ndash;232.\u003c/li\u003e\n\u003cli\u003eBravo-Hollis H (1978). \u003cem\u003eLas Cact\u0026aacute;ceas de M\u0026eacute;xico, vol. I\u003c/em\u003e. 2da Ed. M\u0026eacute;xico, DF: UNAM, 755pp.\u003c/li\u003e\n\u003cli\u003eDarwin C (1859) \u003cem\u003eOn the origin of species by means of natural selection\u003c/em\u003e, London: \u003cem\u003eJ. Murray\u003c/em\u003e, 502pp.\u003c/li\u003e\n\u003cli\u003eDe Queiroz K (2007) Species concepts and species delimitation. \u003cem\u003eSyst Biol\u003c/em\u003e 56(6): 879\u0026ndash;886. https://doi.org/10.1080/10635150701701083\u003c/li\u003e\n\u003cli\u003eEmerson BC, Pati\u0026ntilde;o J (2018) The evolution of Cladogenetic and Anagenetic concepts in Island Biogeography. \u003cem\u003eTrends Ecol Evol\u003c/em\u003e 33(7): 488\u0026ndash;491.\u003c/li\u003e\n\u003cli\u003eFitzpatrick BM, Fordyce JA, Gavrilets S (2009) Pattern, process and geographic modes of speciation. \u003cem\u003eJ Evol Biol\u003c/em\u003e 22: 2342\u0026ndash;2347. \u003c/li\u003e\n\u003cli\u003eFranck AR, Cochrane BJ, Garey JR (2012) Low-copy nuclear primers and \u003cem\u003eycf1\u003c/em\u003e primers in Cactaceae. \u003cem\u003eAm J Bot\u003c/em\u003e 99(10): e405-7. https://doi.org/10.3732/ajb.1200128\u003c/li\u003e\n\u003cli\u003eGibson AC, Nobel P (1986) \u003cem\u003eThe cactus primer\u003c/em\u003e, USA: Harvard University Press, 286pp.\u003c/li\u003e\n\u003cli\u003eGonz\u0026aacute;lez-Terrazas TP, Martel C, Milet-Pinheiro P, Ayasse M, Kalko EKV, Tschapka M (2016) Finding flowers in the dark: Nectar-feeding bats integrate olfaction and echolocation while foraging for nectar. \u003cem\u003eR Soc Open Sci\u003c/em\u003e \u003cem\u003e3\u003c/em\u003e(8): 160199. https://doi.org/10.1098/rsos.160199\u003c/li\u003e\n\u003cli\u003eGranados-Aguilar X, Rosas U, Gonz\u0026aacute;lez-Rodr\u0026iacute;guez A, Arias S (2022) The prickly problem of interwoven lineages: hybridization process in Cactaceae. Bot Sci, 100(4): 797-813.\u003c/li\u003e\n\u003cli\u003eHern\u0026aacute;ndez-Hern\u0026aacute;ndez T, Hern\u0026aacute;ndez HM, De-Nova JA, Puente R, Eguiarte LE, Magall\u0026oacute;n S (2011) Phylogenetic relationships and evolution of growth form in Cactaceae (Caryophyllales, Eudicotyledoneae). \u003cem\u003eAm J Bot\u003c/em\u003e 98(1): 44\u0026ndash;61. https://doi.org/10.3732/ajb.1000129\u003c/li\u003e\n\u003cli\u003eHuelsenbeck JP, Ronquist F (2001) MrBayes: Bayesian inference of phylogenetic trees. \u003cem\u003eBioinformatics\u003c/em\u003e \u003cem\u003e17\u003c/em\u003e(8): 754\u0026ndash;755. Retrieved from http://www.ncbi.nlm.nih.gov/pubmed/11524383\u003c/li\u003e\n\u003cli\u003eHunt DR (2006) \u003cem\u003eThe New Cactus Lexicon\u003c/em\u003e, dh Books, 527pp.\u003c/li\u003e\n\u003cli\u003eKarlsson-Green K, Svensson EI, Bergsten J, H\u0026auml;rdling R, Hansson B (2014) The interplay between local ecology, divergent selection, and genetic drift in population divergence of a sexually antagonistic female trait. \u003cem\u003eEvolution\u003c/em\u003e 7: 1934\u0026ndash;1946.\u003c/li\u003e\n\u003cli\u003eKorotkova N, Aquino D, Arias S, Eggli U, Franck AR, G\u0026oacute;mez-Hinostrosa C, Guerrero PC, Hern\u0026aacute;ndez HM, Kohlbecker A, K\u0026ouml;hler M, Luther K, Majure LC, M\u0026uuml;ller A, Metzing D, Nyffeler R, S\u0026aacute;nchez D, Schlumpberger B, Berendsohn W (2021) Cactaceae at Caryophyllales.org \u0026ndash; a dynamic online species-level taxonomic backbone for the family. \u003cem\u003eWilldenowia\u003c/em\u003e 51(2): 251\u0026ndash;270.\u003c/li\u003e\n\u003cli\u003eLande R (1980) Genetic variation and phenotypic evolution during allopatric speciation. \u003cem\u003eAm Nat\u003c/em\u003e 116: 463\u0026ndash;479. \u003c/li\u003e\n\u003cli\u003eMorrone JJ (2019) Regionalizaci\u0026oacute;n biogeogr\u0026aacute;fica y evoluci\u0026oacute;n bi\u0026oacute;tica de M\u0026eacute;xico: encrucijada de la biodiversidad del Nuevo Mundo. \u003cem\u003eRev Mex Biodivers,\u003c/em\u003e 90: e902980.\u003c/li\u003e\n\u003cli\u003eMorrone JJ, Escalante T, Rodr\u0026iacute;guez-Tapia G, Carmona A, Arana\u003csup\u003e \u003c/sup\u003eM, Mercado-G\u0026oacute;mez JD (2022) Biogeographic regionalization of the Neotropical region: new map and shapefile. \u003cem\u003eAn Acad Bra Cienc\u003c/em\u003e 94: e20211167.\u003c/li\u003e\n\u003cli\u003eMurillo J, Valencia J, Orozco CI, Parra C, Neubig KM (2022) Incomplete lineage sorting and reticulate evolution mask species relationships in Brunelliaceae, an Andean family with rapid, recent diversification. \u003cem\u003eAm J Bot\u003c/em\u003e 109: 1139\u0026ndash;1156.\u003c/li\u003e\n\u003cli\u003eNei M (1972) Genetic Distance between Populations. \u003cem\u003eAm Nat\u003c/em\u003e 106(949): 283\u0026ndash;292.\u003c/li\u003e\n\u003cli\u003eOcampo G, Columbus JT (2010) Molecular phylogenetics of suborder Cactineae (Caryophyllales), including insights into photosynthetic diversification and historical biogeography. \u003cem\u003eAm J Bot\u003c/em\u003e 97(11): 1827\u0026ndash;1847. https://doi.org/10.3732/ajb.1000227\u003c/li\u003e\n\u003cli\u003ePeakall R, Smouse PE (2012) GenAlEx 6.5: genetic analysis in Excel. Population genetic software for teaching and research-an update. \u003cem\u003eBioinformatics\u003c/em\u003e 28: 2537\u0026ndash;2539.\u003c/li\u003e\n\u003cli\u003ePeters EM, Martorell C, Ezcurra E (2008) Nurse rocks are more important than nurse plants in determining the distribution and establishment of globose cacti (\u003cem\u003eMammillaria\u003c/em\u003e) in the Tehuac\u0026aacute;n Valley, Mexico. \u003cem\u003eJ Arid Environ\u003c/em\u003e 70: 593\u0026ndash;601. https://doi.org/10.1016/j.jaridenv.2007.10.004\u003c/li\u003e\n\u003cli\u003ePhillips SJ, Anderson RP, Schapire RE (2006) Maximum entropy modeling of species geographic distributions. \u003cem\u003eEcol Modell\u003c/em\u003e 190: 231\u0026ndash;259.\u003c/li\u003e\n\u003cli\u003ePlume O, Straub SCK, Tel-Zur N, Cisneros A, Schneider B, Doyle JJ (2013) Testing a Hypothesis of Intergeneric Allopolyploidy in Vine Cacti (Cactaceae: Hylocereeae). \u003cem\u003eSyst Bot\u003c/em\u003e 38(3): 737\u0026ndash;751. https://doi.org/10.1600/036364413X670421\u003c/li\u003e\n\u003cli\u003eQGIS Development Team (2016) QGIS Geographic Information System\u003c/li\u003e\n\u003cli\u003eRStudio Team (2020) RStudio: Integrated Development for R. RStudio, PBC, Boston, MA http://www.rstudio.com/.\u003c/li\u003e\n\u003cli\u003eSang T, Crawford DJ, Stuessy TF (1997) Chloroplast DNA phylogeny, reticulate evolution, and biogeography of \u003cem\u003ePaeonia\u003c/em\u003e (Paeoniaceae). \u003cem\u003eAm J Bot\u003c/em\u003e 84(9): 1120\u0026ndash;1136. http://www.amjbot.org/content/84/8/1120.abstract\u003c/li\u003e\n\u003cli\u003eSchnable PS, Hsia AP, Nikolau BJ (1998) Genetic recombination in plants. \u003cem\u003eCurr Opin Plant Biol\u003c/em\u003e 1(2): 123\u0026ndash;129. https://doi.org/10.1016/S1369-5266(98)80013-7\u003c/li\u003e\n\u003cli\u003eShaw J, Lickey EB, Schilling EE, Small RL (2007) Comparison of whole chloroplast genome sequences to choose noncoding regions for phylogenetic studies in angiosperms: the tortoise and the hare III. \u003cem\u003eAm J Bot\u003c/em\u003e 94(3): 275\u0026ndash;288.\u003c/li\u003e\n\u003cli\u003eSlatkin M (1987) Gene Flow and the Geographic Structure of Natural Populations. \u003cem\u003eScience\u003c/em\u003e 236(4803): 787\u0026ndash;792.\u003c/li\u003e\n\u003cli\u003eSlatkin M (1993) Isolation by Distance in Equilibrium and Non-Equilibrium Populations. \u003cem\u003eEvolution\u003c/em\u003e 47(1): 264\u0026ndash;279.\u003c/li\u003e\n\u003cli\u003eSoltis PS, Soltis DE (2009) The role of hybridization in plant speciation. \u003cem\u003eAnn Rev Plant Biol\u003c/em\u003e 60: 561\u0026ndash;588. https://doi.org/10.1146/annurev.arplant.043008.092039\u003c/li\u003e\n\u003cli\u003eTaberlet P, Gielly L, Pautou G, Bouvet J (1991) Universal primers for amplification of three non-coding regions of chloroplast DNA. \u003cem\u003ePlant Mol Biol\u003c/em\u003e 17(5): 1105\u0026ndash;1109. http://www.ncbi.nlm.nih.gov/pubmed/1932684\u003c/li\u003e\n\u003cli\u003eTapia HJ, Arias S, Y\u0026aacute;\u0026ntilde;ez-Espinosa L, Terrazas T (2016) El uso de espinas del tallo en la identificaci\u0026oacute;n de las especies de \u003cem\u003eNeobuxbaumia\u003c/em\u003e (Cactaceae). \u003cem\u003eRev Mex Biodivers\u003c/em\u003e 87(2): 288\u0026ndash;300. https://doi.org/10.1016/j.rmb.2016.04.006\u003c/li\u003e\n\u003cli\u003eTapia HJ, B\u0026aacute;rcenas-Arg\u0026uuml;ello ML, Terrazas T, Arias S (2017) Phylogeny and Circumscription of \u003cem\u003eCephalocereus\u003c/em\u003e (Cactaceae) Based on Molecular and Morphological Evidence. \u003cem\u003eSyst Bot\u003c/em\u003e 42(4): 1\u0026ndash;15. https://doi.org/10.1600/036364417X696546\u003c/li\u003e\n\u003cli\u003eTate JA, Simpson BB (2003) Paraphyly of \u003cem\u003eTarasa\u003c/em\u003e (Malvaceae) and Diverse Origins of the Polyploid Species. \u003cem\u003eSyst Bot\u003c/em\u003e 28(4): 723\u0026ndash;737. https://doi.org/10.1043/02-64.1\u003c/li\u003e\n\u003cli\u003eValiente-Banuet A, Rojas-Mart\u0026iacute;nez A, Arizmendi MC, D\u0026aacute;vila P (1997) Pollination biology of two columnar cacti (\u003cem\u003eNeobuxbaumia mezcalaensis\u003c/em\u003e and \u003cem\u003eNeobuxbaumia macrocephala\u003c/em\u003e) in the Tehuacan Valley, central Mexico. \u003cem\u003eAm J Bot\u003c/em\u003e 84(4): 452\u0026ndash;455.\u003c/li\u003e\n\u003cli\u003eVite F, Portilla E, Zavala-hurtado JA, Valverde PL, D\u0026iacute;az-Sol\u0026iacute;s A (1996) A natural hybrid population between \u003cem\u003eNeobuxbaumia tetetzo\u003c/em\u003e and \u003cem\u003eCephalocereus columna-trajani\u003c/em\u003e (Cactaceae). \u003cem\u003eJournal of Arid Environments\u003c/em\u003e, 32: 395\u0026ndash;405.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"divergence, cladogenesis, reproductive isolation, syngenesis","lastPublishedDoi":"10.21203/rs.3.rs-2481800/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-2481800/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003ePhylogenetic and geographic distances were estimated to produce a combined Distancing Index as a measure of historical reproductive isolation in the genus \u003cem\u003eCephalocereus\u003c/em\u003e. Geographic and climatic barriers were inferred from potential and observed distributions. Distances were extracted from a Bayesian tree for seven chloroplast regions and 26 terminals, and from the geolocation dataset for the exact same sampling. Distance matrices were relativized, and a Mantel test was applied to identify deviations between phylogenetic and geographic distances. Heatmaps and scatterplots were implemented to visualize combined trends. Two basal clades show patterns of differentiation or complete isolation; the first includes \u003cem\u003eC. scoparius\u003c/em\u003e, \u003cem\u003eC. apicicephalium\u003c/em\u003e, \u003cem\u003eC. nizandensis\u003c/em\u003e, and \u003cem\u003eC. totolapensis\u003c/em\u003e, and the second includes \u003cem\u003eC. parvispinus\u003c/em\u003e, \u003cem\u003eC. polylophus\u003c/em\u003e, and \u003cem\u003eC. euphorbioides\u003c/em\u003e. The species \u003cem\u003eC. fulviceps\u003c/em\u003e, and \u003cem\u003eC. sanchezmejoradae\u003c/em\u003e appear in a differentiated grade as sisters of a well-defined clade that includes \u003cem\u003eC. mezcalaensis\u003c/em\u003e, \u003cem\u003eC. macrocephalus\u003c/em\u003e, \u003cem\u003eC. tetetzo\u003c/em\u003e, \u003cem\u003eC. senilis\u003c/em\u003e, \u003cem\u003eC. columna-trajani\u003c/em\u003e, \u003cem\u003eC. multiareolatus\u003c/em\u003e, and \u003cem\u003eC. nudus\u003c/em\u003e, where geographic or phylogenetic distances lie below the mean, indicating a diversification process in absence of hard barriers. At the generic level, separation is related to climatic factors as temperature and moisture, while factors as the altitude could be determinants of separation at the species level. The steady accumulation of variants may lead to opposed evolutionary outcomes: differentiation or diversification, in isolated and non-isolated lineages, respectively. More studies are needed on how genetic variation is transferred or interchanged between and among lineages, and how morphological differentiation of diverging lineages account for reproductive isolation.\u003c/p\u003e","manuscriptTitle":"Phylogenetic and geographic diversification/differentiation as an evolutionary avenue in the genus Cephalocereus (Cactaceae) Evolutionary Avenue in Cephalocereus","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2023-01-18 23:27:13","doi":"10.21203/rs.3.rs-2481800/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"4de35af4-1568-4844-a979-84cc6fe8a08d","owner":[],"postedDate":"January 18th, 2023","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2024-03-15T06:59:46+00:00","versionOfRecord":[],"versionCreatedAt":"2023-01-18 23:27:13","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-2481800","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-2481800","identity":"rs-2481800","version":["v1"]},"buildId":"cBFmMYwuxLRRLfASyISRj","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}