Research Article |
Corresponding author: J. Hugo Cota-Sánchez ( hugo.cota@usask.ca ) Academic editor: Ilya Kirov
© 2018 Carina Gutiérrez-Flores, José L. León-de la Luz, Francisco J. García-De León, J. Hugo Cota-Sánchez.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Gutiérrez-Flores C, León-de la Luz JL, García-De León FJ, Cota-Sánchez JH (2018) Variation in chromosome number and breeding systems: implications for diversification in Pachycereus pringlei (Cactaceae). Comparative Cytogenetics 12(1): 61-82. https://doi.org/10.3897/compcytogen.v12i1.21554
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Polyploidy, the possession of more than two sets of chromosomes, is a major biological process affecting plant evolution and diversification. In the Cactaceae, genome doubling has also been associated with reproductive isolation, changes in breeding systems, colonization ability, and speciation. Pachycereus pringlei (S. Watson, 1885) Britton & Rose, 1909, is a columnar cactus that has long drawn the attention of ecologists, geneticists, and systematists due to its wide distribution range and remarkable assortment of breeding systems in the Mexican Sonoran Desert and the Baja California Peninsula (BCP). However, several important evolutionary questions, such as the distribution of chromosome numbers and whether the diploid condition is dominant over a potential polyploid condition driving the evolution and diversity in floral morphology and breeding systems in this cactus, are still unclear. In this study, we determined chromosome numbers in 11 localities encompassing virtually the entire geographic range of distribution of P. pringlei. Our data revealed the first diploid (2n = 22) count in this species restricted to the hermaphroditic populations of Catalana (ICA) and Cerralvo (ICE) Islands, whereas the tetraploid (2n = 44) condition is consistently distributed throughout the BCP and mainland Sonora populations distinguished by a non-hermaphroditic breeding system. These results validate a wider distribution of polyploid relative to diploid individuals and a shift in breeding systems coupled with polyploidisation. Considering that the diploid base number and hermaphroditism are the proposed ancestral conditions in Cactaceae, we suggest that ICE and ICA populations represent the relicts of a southern diploid ancestor from which both polyploidy and unisexuality evolved in mainland BCP, facilitating the northward expansion of this species. This cytogeographic distribution in conjunction with differences in floral attributes suggests the distinction of the diploid populations as a new taxonomic entity. We suggest that chromosome doubling in conjunction with allopatric distribution, differences in neutral genetic variation, floral traits, and breeding systems has driven the reproductive isolation, evolution, and diversification of this columnar cactus.
Diploid, Cactaceae , cryptic speciation, cytogeography, karyotype, Pachycereus pringlei , polyploidy
Polyploidy and hybridisation are major biological events in plant evolution and speciation (
The Cactaceae, a family with approximately 1,430 species (
Surveys of chromosome variation, both numerical and structural, have been successfully applied in systematic studies of the Cactaceae (
The columnar cactus Pachycereus pringlei (S. Watson, 1885) Britton & Rose, 1909, has been an excellent model plant for ecological and evolutionary studies because this species has an extensive distribution range in the Mexican portion of the Sonoran Desert (
The biogeographic distribution pattern of P. pringlei has been driven by longstanding climatic fluctuations associated with differential colonization abilities of genders, geographic variation of selfing and outcrossing rates, and the effect of biotic and abiotic factors (
Chromosome counts and allozyme data have revealed that Pachycereus pringlei is tetraploid (2n = 44), but these reports are supported by scanty evidence from northern BCP at El Rosario (
Pachycereus pringlei, a cactus commonly known as Cardón, is circumscribed within the subfamily Cactoideae. The species dominates rocky slopes and alluvial plains in the deserts of the BCP, most islands of the Gulf of California, and coastal areas of mainland Sonora, Mexico (
Typical vegetative and floral morphology of the emblematic cactus Pachycereus pringlei.A Mature individual with candelabra-like structure in the Cataviña region. B Archetypal funnel-form flower. C Main pollinator, the long-nosed bat Leptonycteris yerbabuenae. D Mature, fleshy fruit. Photo A by Jon Rebman; photo C by Merlin D. Tutle.
The chromosome numbers inspected in this study were obtained from individual plants from natural populations across the wide distributional range of P. pringlei encompassing variation in ecology, latitude, and longitude as well as floral morphology, breeding systems, and levels of genetic diversity. Fruits with mature seeds were collected in the field from three to 10 individuals in the following localities of the BCP: Bahía de Los Ángeles (BAN), Cabo San Lucas (CBS), Catalana Island (ICA), Cerralvo Island (ICE), El Comitán (COM), López Mateos (LMA), Loreto (LOR), San Felipe (SFE), Puente Querétaro (PQU), and Santa Rosalía (SRO). Also, one fruit collected in mainland Mexico from one locality, Álamos (ALA), Sonora, was scrutinised (Table
Sample sites of the columnar cactus Pachycereus pringlei for which chromosome numbers were investigated, including counts by
Locality | Code | Latitude | Longitude | Chromosome number | Ploidy level | Breeding system | Genetic diversity | Genetic population |
---|---|---|---|---|---|---|---|---|
Cabo San Lucas | CBS | 22.9438, -109.9905 | 2n = 44 | Tetraploid | Mainly dioecious | 0.38 | CBS | |
Catalana Island | ICA | 25.6768, -110.8087 | 2n = 22 | Diploid | Hermaphroditic | 0.40 | ICA | |
Cerralvo Island | ICE | 24.1868, -109.8775 | 2n = 22 | Diploid | Hermaphroditic | 0.26 | ICE | |
El Comitán | COM | 24.1332, -110.4317 | 2n = 44 | Tetraploid | Mainly trioecious | 0.45 | South | |
López Mateos | LMA | 25.2726, -111.8942 | 2n = 44 | Tetraploid | 0.45 | |||
Loreto | LOR | 25.8918, -111.4698 | 2n = 44 | Tetraploid | ND | |||
Puente Querétaro | PQU | 25.3508, -111.6094 | 2n = 44 | Tetraploid | 0.45 | |||
Santa Rosalía | SRO | 27.2408, -112.3615 | 2n = 44 | Tetraploid | Mainly gynodioecious | ND | North | |
Bahía de Los Ángeles | BAN | 28.9164, -113.5541 | 2n = 44 | Tetraploid | 0.35 | |||
El Rosario | ROS | 30.0861, -115.6795 | n = 22 | Tetraploid† | ND | |||
San Felipe | SFE | 30.3716, -114.8537 | 2n = 44 | Tetraploid | 0.35 | |||
Álamos | ALA | 26.8978, -109.4578 | 2n = 44 | Tetraploid | 0.35 | |||
Bahía Kino | BKI | 28.5000, -111.8063 | ND | Tetraploid* | ND |
Mitotic chromosome numbers were determined using meristematic cells from fresh root tips following a modified protocol by
For meiotic figures and counts, anthers from floral buds at different developmental stages were fixed in Carnoy’s solution, then stained with aceto-orcein at room temperature for 1 h, squashed, mounted in Hoyer’s medium and microscopically examined as indicated above. A minimum of three cytological figures from different individuals were scrutinized in each population for confirmation of chromosome number. Finally, the geographic distribution of diploid and polyploid cytotypes from the localities examined was plotted on a base map obtained from the Comisión Nacional para el Conocimiento y Uso de la Biodiversidad (CONABIO, http://www.conabio.gob.mx) using the ArcGIS 10.4 software (ESRI).
Idiograms were reconstructed based on microscopic observation of mitotic figures. Chromosome homology for diploid and tetraploid cytotypes follows
To compare floral variability among diploid and polyploid populations of P. pringlei, 17 floral traits were selected and a one-way ANOVA of morphological attributes from flowers gathered for the five genetic populations fide
Measurements of phenotypic floral traits, including sample size (mean ± SE) and statistical comparisons of bisexual flowers between diploid (ICE, ICA) and polyploid (CBS, SOUTH, NORTH) populations of Pachycereus pringlei in the Baja California Peninsula, Mexico. Lower case superscript letters indicate floral characters having statistically significant differences.
Floral trait | Diploid | Polyploid | |||
---|---|---|---|---|---|
ICE | ICA | CBS | SOUTH | NORTH | |
n = 50 | n = 35 | n = 7 | n = 20 | n = 38 | |
Corolla width (mm) | 33.3 ± 0.7 | 36.2 ± 0.9 | 29.4 ± 1.2 | 33.1 ± 1.0 | 36.0 ± 0.7 |
Floral length (mm) | 90.7 ± 1.2 | 86.8 ± 0.4 | 76.4 ± 2.8 | 85.1 ± 2.1 | 96.2 ± 1.5 |
Nectary length (mm) | 12.9 ± 0.3 | 13.0 ± 0.3 | 9.2 ± 0.4 | 11.9 ± 0.4 | 13.0 ± 0.3 |
Nectary width (mm) | 11.1 ± 0.2b | 10.7 ± 0.3b | 8.7 ± 0.5a | 9.3 ± 0.2a | 9.9 ± 0.3a |
No. of pollen grains (x106) | 2.8 ± 0.3c | 2.3 ± 0.2cb | 1.0 ± 0.2a | 1.3 ± 0.2a | 1.6 ± 0.2ab |
No. of stamens (in 0.5 cm2) | 48.6 ± 1.0ab | 43.0 ± 1.8a | 49.0 ± 3.14ab | 53.4 ± 1.9b | 51.6 ± 2.0b |
No. of tepals | 49.9 ± 0.6 | 46.7 ± 1.2 | 44.3 ± 1.6 | 47.9 ± 1.4 | 49.9 ± 0.8 |
P:O ratio (x103) | 2337 ± 255b | 2638 ± 307b | 1178 ± 289a | 911 ± 117a | 2166 ± 203a |
Stamen length (mm) | 10.0 ± 0.2 | 10.2 ± 0.2 | 11.9 ± 0.7 | 10.0 ± 0.2 | 10.9 ± 0.2 |
Tepal length (mm) | 23.0 ± 0.4 | 23.5 ± 0.4 | 19.0 ± 0.6 | 20.4 ± 0.6 | 23.0 ± 0.5 |
Tepal width (mm) | 7.7 ± 0.3 | 8.1 ± 0.3 | 5.9 ± 0.3 | 8.0 ± 0.3 | 9.1 ± 0.2 |
Ovary length (mm) | 14.0 ± 0.4 | 9.4 ± 0.4 | 8.3 ± 0.7 | 13.4 ± 0.7 | 15.5 ± 0.5 |
Ovary width (mm) | 8.9 ± 0.3 | 7.9 ± 0.3 | 8.1 ± 0.5 | 8.7 ± 0.4 | 9.6 ± 0.3 |
Pistil length (mm) | 44.6 ± 0.7 | 47.1 ± 0.9 | 43.8 ± 1.2 | 47.9 ± 1.4 | 51.0 ± 0.9 |
Stigma length (mm) | 9.8 ± 0.3 | 8.8 ± 0.3 | 7.7 ± 0.5 | 8.9 ± 0.3 | 10.7 ± 0.4 |
No. of ovules (mm) | 1550 ± 66b | 907 ± 40a | 849 ± 115a | 1505 ± 118b | 1614 ± 112b |
Stamen-stigma distance (mm) | 0.5 ± 0.5a | 2.0 ± 0.6ab | 1.5 ± 1.1ab | 4.2 ± 1.0bc | 5.4 ± 0.6c |
Chromosome counts performed in the populations of P. pringlei investigated revealed variation in ploidy level. Foremost, our survey unveiled the first diploid (2n = 2x = 22) count for this species and expanded the current cytological knowledge with additional tetraploid (2n = 4x = 44) cytotypes in different localities of the BCP. The diploid counts were consistently determined in all mitotic cells in plants from Cerralvo (ICE) and Catalana (ICA) Islands (Figs
Representative metaphase chromosomes of Pachycereus pringlei. Cerralvo Island (ICE) and Catalana Island (ICA) are the sole localities for diploid cytotypes. Remaining pictures in plate are typical chromosomes in different tetraploid populations. Scale Bar: 2 µm.
Distribution of diploid and tetraploid individuals in populations (black dots) of Pachycereus pringlei. Dark gray areas in ICA and ICE indicate distribution of diploid (2n = 22) cytotypes. Diagonal shaded area indicates the predicted coverage of tetraploid (2n = 44) cytotypes. See Table
The overall morphology of mitotic chromosomes for diploid and tetraploid cytotypes of this columnar cactus is, in general, symmetric and uniform in shape, i.e., chromosomes are mostly metacentric (M) with a few submetacentric (SM) and relatively small in size (measuring in average 2µm in length) (Figs
Morphological comparisons of bisexual flowers among populations revealed a few significant statistical differences that can be associated with variation in ploidy level. For instance, diploid individuals from ICE and ICA have wider nectaries (11.1 ± 0.2 and 10.7 ± 0.3 mm, respectively) and larger P:O ratios (2,337 ± 255 and 2,638 ± 307, respectively). The diploid cytotypes also tend to have larger amount of pollen grains (2.8 x106 and 2.3 x106, respectively), fewer number of stamens (48.6 ± 1.0, and 43.0 ± 1.8, respectively), fewer number of ovules (1,550 ± 66 and 907 ± 40, respectively), and closer proximity between stamen and stigmas (0.5 ± 0.5 and 2.0 ± 0.6 mm, respectively). See also Table
Chromosome number variation, especially polyploidy, is one of the major biological processes that has affected angiosperm evolution (
This study has unveiled the first report of diploid (2n = 2x = 22) cytotypes in P. pringlei and expands the distributional range of the tetraploid (2n = 4x = 44) condition known for this species. Geographically, our survey also revealed that the base diploid number is maintained exclusively in the two deep-water islands (ICA and ICE) of the Gulf of California characterised by the prevalence of hermaphrodite individuals. In turn, double chromosome dosage (tetraploidy) is consistent throughout the three BCP populations (CBS, North and South) (Fig.
Among polyploids, tetraploidy is the most successful condition (
During past geological events, the southern BCP, including ICE and ICA Islands, exhibited suitable niche conditions that served as refugia for populations of P. pringlei during the Last Glacial Maximum (LGM), as evidenced by Ecological Niche Modelling (
In terms of morphology, the chromosomes of P. pringlei follow the overall homogeneous pattern reported for the sister taxon P. weberi (J.M. Coulter, 1896) Backeberg, 1960 (
Foremost, it is worth noting that the correspondence of the base chromosome number (x = 11) with a hermaphroditic mating system in most cacti, including members of the basal subfamily (Pereskioideae) and species closely related to P. pringlei, such as P. weberi (Gama-López, 1994) and P. pecten-aboriginum (Engelmann ex S. Watson, 1886) Britton & Rose, 1909 (
Diploid and tetraploid plant populations may or may not be ecologically differentiated (
Unveiling polyploid individuals from diploid ancestors leads to the discovery of new cytotypes and potential taxonomic issues because different populations are frequently associated with an assortment of floral and/or vegetative phenotypes (Brickford et al. 2007). In this regard, the phenotypic variation reported in vegetative (
Evidently, genomic changes potentially produce new gene complexes, facilitating rapid evolution of individuals and their new attributes (
Merging chromosome number information, genetic data, breeding systems, and floral morphology has provided new insights to better understand the evolutionary history and reproductive success of this iconic cactus in northwestern Mexico. P. pringlei has diploid and tetraploid populations with distinctive distribution. Although tetraploids have not been named as distinct species due to the tradition of including multiple cytotypes derived from diploid relatives as a single species and the practicality of adhering to the general morphological species concept (
The authors thank the anonymous reviewers for providing feedback to improve the manuscript. This research was funded by the National Geographic Society, Grant No. 7382-02, to JHCS, and the Consejo Nacional de Ciencia y Tecnología (CONACYT), Grant No. CB-2008-01-106925, to FJGDL. CONACYT (Grant No. 165852) and the Emerging Leaders of the Americas Program, Government of Canada, subsidized scholarships for CGF. We are thankful to R. Domínguez-Cadena, A. Medel-Narváez, and A. Lozano-Garza for field assistance, to D. Litwiller and the Cota-Sánchez Plant Systematics Lab personnel for critical discussions and editorial comments in earlier drafts of the manuscript, and to Dr. J. Rebman and Merlyn D. Tutle for providing photographs.