Mexico harbors ~45% of world's cacti species richness. Their biogeography and phylogenomics were integrated to elucidate the evolutionary history of the genera Coryphantha, Escobaria, Mammillaria, Mammilloydia, Neolloydia, Ortegocactus, and Pelecyphora (Mammilloid Clade). We analyzed 52 orthologous loci from 142 complete genomes of chloroplast (103 taxa) to generate a cladogram and a chronogram; in the latter, the ancestral distribution was reconstructed with the Dispersal-Extinction-Cladogenesis model. The ancestor of these genera arose ~7 Mya on the Mexican Plateau, from which nine evolutionary lineages evolved. This region was the site of 52% of all the biogeographical processes. The lineages 2, 3 and 6 were responsible for the colonization of the arid southern territories. In the last 4 Mya, the Baja California Peninsula has been a region of prolific evolution, particularly for lineages 8 and 9. Dispersal was the most frequent process and vicariance had relevance in the isolation of cacti distributed in the south of Mexico. The 70 taxa sampled as Mammillaria were distributed in six distinct lineages; one of these presumably corresponded to this genus, which likely had its center of origin in the southern part of the Mexican Plateau. We recommend detailed studies to further determine the taxonomic circumscription of the seven genera.

Background Structural descriptions of complete genomes have elucidated evolutionary processes in angiosperms. In Cactaceae (Caryophyllales), a high structural diversity of the chloroplast genome has been identified within and among genera. In this study, we assembled the first mitochondrial genome (mtDNA) for the short-globose cactus Mammillaria huitzilopochtli . For comparative purposes, we used the published genomes of 19 different angiosperms and the gymnosperm Cycas taitungensis as an external group for phylogenetic issues. Results The mtDNA of M. huitzilopochtli was assembled into one linear chromosome of 2,052,004 bp, in which 65 genes were annotated. These genes account for 57,606 bp including 34 protein-coding genes (PCGs), 27 tRNAs, and three rRNAs. In the non-coding sequences, repeats were abundant, with a total of 4,550 (179,215 bp). In addition, five complete genes ( psaC and four tRNAs) of chloroplast origin were documented. Negative selection was estimated for most (23) of the PCGs. The phylogenetic tree showed a topology consistent with previous analyses based on the chloroplast genome. Conclusions The number and type of genes contained in the mtDNA of M. huitzilopochtli were similar to those reported in 19 other angiosperm species, regardless of their phylogenetic relationships. Although other Caryophyllids exhibit strong differences in structural arrangement and total size of mtDNA, these differences do not result in an increase in the typical number and types of genes found in M. huitzilopochtli . We concluded that the total size of mtDNA in angiosperms increases by the lengthening of the non-coding sequences rather than a significant gain of coding genes.

In angiosperms, huge advances in massive DNA sequencing technologies have impacted phylogenetic studies. Probe sets have been developed with the purpose of recovering hundreds of orthologous loci of targeted DNA sequences (TDS) across different plant lineages. We tested in silico the effectiveness of two universal probe sets in the whole available genomes of Caryophyllids, emphasizing phylogenetic issues in cacti species. A total of 870 TDS (517 TDS from Angiosperm v.1 and 353 from Angiosperms353) were individually tested in nine cacti species and Amaranthus hypochondriacus (external group) with ≥17 Gbp of available DNA data. The effectiveness was measured by the total number of orthologous loci recovered and their length, the percentage of loci discarded by paralogy, and the proportion of informative sites (PIS) in the alignments. The results showed that, on average, Angiosperms353 was better than Angiosperm v.1 for cacti species, since the former obtained an average of 275.6 loci that represent 123,687 bp, 2.48% of paralogous loci, and 4.32% of PIS in alignments, whereas the latter recovered 148.4 loci (37,683 bp), 10.38% of paralogous loci, and 3.49% of PIS. We recommend the use of predesigned universal probe sets for Caryophyllids, since these recover a high number of orthologous loci that resolve phylogenetic relationships.

The Military Macaw (Ara militaris) faces a number of serious conservation threats. The use of genetic markers and assignment tests may help to identify the geographic origin of captive individuals and improve conservation and management programs. The purpose of this study was to identify the possible geographic origin of a captive individual using genetic markers. We used a reference database of genotypes of 86 individuals previously shown to belong to two different genetic groups to determine the genetic assignment of the captive individual of unknown origin (captive specimen) and five individuals of known geographic origin (as positive controls). We evaluated the accuracy of three assignment/exclusion criteria to determine the success of correct assignment of the individual of unknown origin and the five positive control individuals. WICHLOCI estimated that eight loci were required to achieve an assignment success of 83%. The correct geographic origin of positive controls was identified with 83% confidence. All of the analyses assigned the captive individual to the genetic group from the Sierra Madre Oriental. Bayesian assignment tests, tests for genetic distance and allele frequency tests assigned the unknown individual to the locations from the Sierra Madre Oriental with a probability of 71.2–82.4%. We show that the use of genetic markers provides a promising tool for determining the origin of pets and individuals seized from the illegal animal trade to better inform decisions on reintroduction and improve conservation programs.

In plants, partial DNA sequences of chloroplasts have been widely used in evolutionary studies. However, the Cactaceae family (1500–1800 species) lacks molecular markers that allow a phylogenetic resolution between species and genera. In order to identify sequences with high variation levels, we compared previously reported complete chloroplast genomes of seven species of Mammillaria. We identified repeated sequences (RSs) and two types of DNA variation: short sequence repeats (SSRs) and divergent homologous loci. The species with the highest number of RSs was M. solisioides (256), whereas M. pectinifera contained the highest amount of SSRs (84). In contrast, M. zephyranthoides contained the lowest number (35) of both RSs and SSRs. In addition, five of the SSRs were found in the seven species, but only three of them showed variation. A total of 180 homologous loci were identified among the seven species. Out of these, 20 loci showed a molecular variation of 5% to 31%, and 12 had a length within the range of 150 to 1000 bp. We conclude that the high levels of variation at the reported loci represent valuable knowledge that may help to resolve phylogenetic relationships and that may potentially be convenient as molecular markers for population genetics and phylogeographic studies.

The complete sequence of chloroplast genome (cpDNA) has been documented for single large columnar species of Cactaceae, lacking inverted repeats (IRs). We sequenced cpDNA for seven species of the short-globose cacti of Mammillaria and de novo assembly revealed three novel structures in land plants. These structures have a large single copy (LSC) that is 2.5 to 10 times larger than the small single copy (SSC), and two IRs that contain strong differences in length and gene composition. Structure 1 is distinguished by short IRs of <1 kb composed by rpl23-trnI-CAU-ycf2; with a total length of 110,189 bp and 113 genes. In structure 2, each IR is approximately 7.2 kb and is composed of 11 genes and one Intergenic Spacer-(psbK-trnQ)-trnQ-UUG-rps16-trnK-UUU-matK-trnK-UUU-psbA-trnH-GUG-rpl2-rpl23-trnI-CAU-ycf2; with a total size of 116,175 bp and 120 genes. Structure 3 has divergent IRs of approximately 14.1 kb, where IRA is composed of 20 genes: psbA-trnH-GUG-rpl23-trnI-CAU-ycf2-ndhB-rps7-rps12-trnV-GAC-rrn16-ycf68-trnI-GAU-trnA-AGC-rrn23-rrn4.5-rrn5-trnR-ACG-trnN-GUU-ndhF-rpl32; and IRB is identical to the IRA, but lacks rpl23. This structure has 131 genes and, by pseudogenization, it is shown to have the shortest cpDNA, of just 107,343 bp. Our findings show that Mammillaria bears an unusual structural diversity of cpDNA, which supports the elucidation of the evolutionary processes involved in cacti lineages.

The Military Macaw (Ara militaris) is a globally threatened species with a fragmented distribution, and assessing the genetics of populations could help identify conservation units. Nine microsatellites were used to analyze 86 samples in seven localities along the Sierra Madre Occidental, the Sierra Madre del Sur, and the Sierra Madre Oriental in Mexico. Results showed that the Military Macaw has moderate levels of genetic diversity, similar to that found in other macaw species in Latin America. This species shows a high genetic structure; we find a genetic break between localities separated by the Central Plateau and the Trans-Mexican Volcanic Belt, which serve as geographic barriers. However, the locations within each genetic group are not genetically differentiated. It was observed that three locations of the Military Macaw have excess homozygotes, which could indicate a small effective size of the population and in combination with genetic isolation could increase the risk of extinction of the species. We propose two genetic groups for the species, the first comprising localities in the Sierra Madre Occidental and the Sierra Madre del Sur, and the second comprising localities of the Sierra Madre Oriental. According to the genetic differentiation, which was significant between the physiographic regions, and the unique allelic richness shown in this study, these two groups should be considered as independent conservation units. We strongly recommend the conservation and restoration of the natural habitats of the Military Macaw to maintain and increase the size of its population and recover and expand its original geographical distribution.

Mammillaria groups 200 species. Mexico harbors 164 species, 85 % are endemic and 113 are listed in the Official Mexican Standard NOM-059 SEMARNAT-2010. In spite of their importance in biodiversity and serious conservation crisis, the scarce information documented for these species does not allow proposals for protection strategies. Our objective was to estimate the population genetic diversity levels of M. hernandezii, M. kraehenbuehlii and M. napina to identify genetic conservation units. Twenty-four individuals from two M. hernandezii populations and 120 individuals from five populations for the other two species were genotyped with ten microsatellite loci. In the three species the observed heterozygosity levels were lower than the expected ones (M. hernandezii 0.58, 0.65; M. kraehenbuehlii 0.61, 0.73; and M. napina 0.56, 0.74). The allelic diversity ranged from five (M. hernandezii) to eight alleles in the other two species. The three species showed heterozygotic deficiency possibly due to genetic drift since their populations are small, but selfing may also be involved. Gene flow levels were high for the three especies indicating that genetic drift, pollen and seed dispersal systems determine the genetic structure. Three genetic groups are proposed for M. kraehenbuehlii and M. napina to be considered as reference for conservation programs of the species and their habitats. The main threats for the three species are severe landscape transformation, which isolates the populations, and looting. For M. hernandezii, the number of studied populations should increase to guarantee conclusive results of its population genetic diversity. Currently, the geographic distribution pattern in small patches indicates severe habitat fragmentation that urges to take actions for its protection and management.

Mammillaria agrupa a 200 especies, 164 registradas en México, 85 % son endémicas y 113 están en la Norma Oficial Mexicana NOM-059 SEMARNAT-2010. A pesar de su importancia en la biodiversidad y de su grave crisis de conservación, la escasa información documentada en estas especies no permite proponer estrategias para protegerlas. Nuestro objetivo fue estimar los niveles de diversidad genética poblacional en M. hernandezii, M. kraehenbuehlii y M. napina para identificar unidades genéticas de conservación. Se genotiparon con 10 loci de microsatélites 24 individuos de dos poblaciones de M. hernandezii y 120 individuos de cinco poblaciones para cada una de las otras dos especies. En las tres especies se estimaron niveles de heterocigosidad observada menores que los esperados (M. hernandezii 0.58, 0.65; M. kraehenbuehlii 0.61, 0.73; y M. napina 0.56, 0.74), la diversidad alélica varió de cinco (M. hernandezii) a ocho alelos en las otras dos especies. Las tres especies mostraron una deficiencia de heterocigotos que puede deberse a deriva genética porque sus poblaciones son pequeñas, aunque la autofecundación también podría participar. Entre las poblaciones de las tres especies los niveles de flujo génico fueron altos, lo que indica que podría ser la deriva genética y el sistema de dispersión de polen y semillas lo que determina la estructura genética. Para M. kraehenbuehlii y M. napina se proponen tres grupos genéticos para que sean considerados como referencias para programas de conservación de estas especies y de sus hábitats. Las principales amenazas para las tres especies son la severa transformación del paisaje que aisla a sus poblaciones y el saqueo. En M. hernandezii se debe incrementar el número de poblaciones estudiadas para tener resultados concluyentes de su diversidad genética poblacional, por el momento el patrón de distribución geográfica en parches pequeños indica una severa fragmentación que insta a tomar medidas urgentes para su protección y manejo.

The endemic plant species with extremely narrow geographical range (<100 km2) often have few populations of small size and tend to be more vulnerable to extinction by genetic drift and inbreeding effects. For these species, we tested if intraspecific genetic diversity can be applied to identify conservation priorities. The biological model was Mammillaria albiflora—a Mexican cactus that numbers ~1000 individuals distributed in four nearby patches covering 4.3 km2. A total of 96 individuals were genotyped with 10 microsatellite loci to describe the genetic substructure and diversity. There is significant population substructure: the genetic diversity is distributed in three genetic neighbors and varies among the patches, the genotypes are not randomly distributed and three genetic barriers restrict the gene flow. The current population size is 15 times smaller than in the past. The restricted gene flow and genetic drift are the processes that have shaped population substructure. To conserve the genetic diversity of this cactus we recommend that two patches, which are not private property, be legally protected; to include M. albiflora in the Red List Species of Mexico in the category of extinction risk; and a legal propagation program may help to diminish the illegal harvesting.

Cerca de 168 especies of Mammillaria están amenazadas a nivel global, la mayoría de estas especiesmuestran una distribución restringida y/o poblaciones pequeñas. Recientemente, los primeros estudios de genéticapoblacional mostraron niveles de heterocigosidad bajos a moderados en este género. Este trabajo estudia a Mammillariahuitzilopochtli y M. supertexta, dos cactus amenazados que son endémicos a la región semiárida central de Méxicopara proponer acciones de conservación así como para identificar los procesos subyacentes que determinan susniveles de diversidad genética poblacional. Un total de 106 y de 148 individuos de 5 poblaciones se muestrearonpara M. huitzilopochtli y M. supertexta, respectivamente. Para cada individuo se determinó su genotipo con 8 loci demicrosatélites. Los niveles de heterocigosidad fueron altos en las dos especies, pero fueron detectadas diferencias ensu riqueza alélica. La diferenciación genética entre poblaciones fue significativa en las dos especies, con evidenciade aislamiento por distancia en M. supertexta pero no en M. huitzilopochtli. Proponemos que la deriva génica, elaislamiento geográfico y la endogamia son los procesos más importantes que moldean la variabilidad genética de laspoblaciones y la diferenciación en estas especies. Identificamos 3 y 4 grupos genéticos para M. huitzilopochtli y M.supertexta, respectivamente. En el valle de Tehuacán-Cuicatlán estos grupos podrían ser usados como una referenciapara guiar los esfuerzos de conservación de las plantas bajo una perspectiva de conservación del ecosistema.

Florestina is shown to consist of six annual species occurring mostly in arid and semiarid regions of Mexico. Florestina species are morphologically similar and consequently phylogenetic relationships within the genus are poorly understood. We present a phylogenetic study based on morphological characters, DNA sequences of nuclear non-coding spacers (ETS and ITS) and chloroplast non-coding spacers ( rpl32 - trn L and trn C- pet N). The ETS and ITS spacer-based phylogenies allowed several well-supported conclusions: (1) the genus Florestina is monophyletic and Palafoxia is its closest relative; (2) Florestina latifolia and F. platyphylla form a strongly supported clade; (3) four taxa that are morphologically very similar, F. liebmannii , F. pedata , F. simplicifolia , and F. tripteris , are phylogenetically closely related and based on the sequence data we suggest that these should be recognized as only two species, one comprising F. pedata and F. simplicifolia , which shows wide morphological variation throughout its distributional range; and the other comprising F. liebmannii and F. tripteris ; (4) F. lobata and F. purpurea are species very distinct from the remainder of the species in Florestina . Our phylogenetic analyses suggest that hybridization and introgression may be involved in the evolutionary history of Florestina.

Background: Rare cactus in the Americas and other species worldwide are threatened species because of their high level of habitat specialisation, narrow distribution range and continuing population decline.Aims: To identify management units (MUs) based on genetic variability and demographic structure in order to propose assertive conservation actions for Mammillariacrucigera and to provide a model case study for other species that are under similar threats.Methods: We genotyped through eight microsatellite loci in 171 individuals and described demographic structures in six populations of this cactus based on plots of 1m(2).Results: Across populations with a mean density of 2.6m(-2) and a total of similar to 500 individuals counted, 30% of the individuals were reproductive (diameter>2cm). The total heterozygosity was low (H-O=0.54), but the inbreeding coefficient (F-IS=0.29) and the allele diversity (N-A=20) were high. Four genetic groups were distinguished, although considering the demographic structure, we propose three MUs.Conclusions: It is critical to maintain the genetic connectivity within and among MUs, which can only be achieved through cooperation between government authorities and local habitants to halt the degradation and further destruction of the remnant populations. Searching MUs allows the identification of critical areas for conservation issues for all species whose extant populations are in a fragmented landscape.

Forest structure and composition have been used to assess the habitat characteristics that determine bird distributions. The patterns of distribution have been shaped by historical and ecological factors that play different roles at both temporal and spatial scales. The objectives of this research were to characterize the habitat of the endangered Military Macaw (Ara militaris) and evaluate the potential distribution of this species based on trends of land use changes in Mexico. We characterized the community structure and floristic composition of 8 forests that are currently used by the Military Macaw for breeding and feeding and compared the results with 6 similar forests characterized in other studies but without historical records of the presence of the Military Macaw. The Military Macaw preferred sites with high diversity of plant species dominated by trees from 4 to 15 m in height and from 5 to 90 cm in diameter at breast height. We identified 236 plant species in the 8 forests with 20 species (8.4%) used for nesting and feeding by the Military Macaw. The floristic composition is important for the presence of the Military Macaw because there were significant differences between forests with and without its presence. The potential area of distribution of the Military Macaw had decreased by 32% and the remnant areas are included in only 8 National Protected Areas. The protected areas of natural forests should be increased to preserve the sites of potential distribution and consequently the habitat of the Military Macaw in Mexico.

In this study, we analyzed the genetic variation of quetzals ( Pharomachrus mocinno ) throughout their geographic distribution to determine conservation targets. This species is found in patchy isolated cloud forests from Mexico to Panama. A multidimensional scaling and UPGMA analysis of a 286 RAPD fragment set resolved 3 genetic groups: cluster 1 (Mexican localities), cluster 2 (Guatemala, Nicaragua and El Salvador) and cluster 3 (Panama). The mean genetic diversity estimated by the Shannon index was 0.38, 0.22 and 0.32, for clusters 1, 2, and 3, respectively. The genetic differentiation among clusters was statistically significant. The highest percentage of genetic variation (70.86%) was found within populations using an AMOVA analysis. Our results suggest that within the quetzal species, there are 3 genetic groups that should be considered as independent conservation targets and included in a global Mesoamerican conservation program.

The resplendent Quetzal (Pharomachrus mocinno) is an endemic Mesoamerican bird species of conservation concern. Within this species, the subspecies P. m. costaricensis and P. m. mocinno, have been recognized by apparent morphometric differences; however, presently there is no sufficient data for confirmation. We analyzed eight morphometric attributes of the body from 41 quetzals: body length, tarsus and cord wing, as well as the length, wide and depth of the bill, body weight; and in the case of the males, the length of the long upper-tail cover feathers. We used multivariate analyses to discriminate morphometric differences between subspecies and contrasted each morphometric attribute between and within subspecies with paired non-parametric Wilcoxon test. In order to review the intraspecific taxonomic status of this bird, we added phylogenetic analysis, and genetic divergence and differentiation based on nucleotide variations in four sequences of mtDNA. The nucleotide variation was estimated in control region, subunit NDH6, and tRNAGlu and tRNAPhe in 26 quetzals from eight localities distributed in five countries. We estimated the genetic divergence and differentiation between subspecies according to a mutation-drift equilibrium model. We obtained the best mutation nucleotide model following the procedure implemented in model test program. We constructed the phylogenetic relationships between subspecies by maximum parsimony and maximum likelihood using PAUP, as well as with Bayesian statistics. The multivariate analyses showed two different morphometric groups, and individuals clustered according to the subspecies that they belong. The paired comparisons between subspecies showed strong differences in most of the attributes analyzed. Along the four mtDNA sequences, we identified 32 nucleotide positions that have a particular nucleotide according to the quetzals subspecies. The genetic divergence and the differentiation was strong and markedly showed two groups within P. mocinno that corresponded to the quetzals subspecies. The model selected for our data was TVM+G. The three phylogenetic methods here used recovered two clear monophyletic clades corresponding to each subspecies, and evidenced a significant and true partition of P. mocinno species into two different genetic, morphometric and ecologic groups. Additionally, according to our calculations, the gene flow between subspecies is interrupted at least from three million years ago. Thus we propose that P. mocinno be divided in two independent species: P. mocinno (Northern species, from Mexico to Nicaragua) and in P. costaricensis (Southern species, Costa Rica and Panama). This new taxonomic classification of the quetzal subspecies allows us to get well conservation achievements because the evaluation about the kind and magnitude of the threats could be more precise.

The cactus Mammillaria crucigera is a threatened species endemic to central Mexico. As a means of assessing population genetic status of these species, eight microsatellite markers were developed. These primers were tested in 40 individual from two wild populations. The results showed that these primers will be useful to describe population structure and aid to the conservation of species. The eight primers were tested in other Mammillaria species and most of them showed successful amplification.