| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Genetics and Molecular Biology |
2Instituto de Botánica del Nordeste (UNNE–CONICET), Casilla de Correo 209, 3400 Corrientes, Corrientes Province, Argentina; 3Facultad de Ciencias Agrarias (UNNE) 4Miembro de la Carrera del Investigador Científico (CONICET), Facultad de Ciencias Exactas y Naturales y Agrimensura (UNNE), Corrientes, Corrientes Province, Argentina
Received for publication June 26, 2001. Accepted for publication October 2, 2001.
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
Key Words: autopolyploidy • karyotype evolution • Leiocarpae • taxonomy • Turnera • Turneraceae
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
) that are distributed largely in tropical and subtropical areas of the Americas.
At least 34% of all species have been investigated karyologically. Chromosome numbers have been reported for 35 species (Raman and Kesavan, 1964
; Hamel, 1965
; Barrett, 1978
; Barrett and Shore, 1987; Arbo and Fernández, 1983
; Fernández, 1987
; Solís Neffa and Fernández, 1993
) but the karyotypes of only 21 have been described (Solís Neffa and Fernández, 1993
; Solís Neffa, 1996
). The basic chromosome number x = 7 was found in the series Salicifoliae, Stenodictyae, Mycrophyllae, and Leiocarpae and is clearly the most common, while x = 5 was found only in the Turnera (= Canaligerae) series and x = 13 in the Papilliferae (Fernández, 1987
). Cytological investigation has shown the occurrence of diploid to decaploid populations and that both autopolyploids and allopolyploids may be present (Raman and Kesavan, 1964
; Hamel, 1965
; Barrett, 1978
; Arbo and Fernández, 1983
; Barrett and Shore, 1987
; Fernández, 1987
; Shore, 1991a
).
Continuing the karyological investigation of the genus, in this article we describe the karyotypes of T. sidoides L., which belongs to series Leiocarpae and has the basic number x = 7. Turnera sidoides is a complex of obligately outcrossing perennial herbs, and it is the species with the most southerly distribution in America (Arbo, 1986
). It extends from the southern regions of Bolivia and Brazil, through Paraguay to Uruguay and Argentina, reaching 39° S (Arechavaleta, 1905
; Arbo, 1985, 1987
; Solís Neffa, 2000
).
After the first description of T. sidoides by Urban (1883)
, Arbo (1985)
revised this species extensively and, on the basis of the geographical distribution and the great variability of some morphological features, recognized five subspecies.
Different ploidy levels have been reported for all of the subspecies (Fernández, 1987
; Solís Neffa and Fernandez, 2001
), ranging from diploid to octoploid. Diploid, tetraploid, and hexaploid cytotypes occur in T. sidoides subsp. carnea (Cambess.) Arbo and T. sidoides subsp. pinnatifida (Juss. et Poir.) Arbo, whereas tetraploids and hexaploids are known in T. sidoides subsp. holosericea (Urban) Arbo. Turnera sidoides subsp. integrifolia (Griseb.) Arbo shows a polyploid series with ploidy levels from 2x to 8x. In T. sidoides subsp. sidoides, tetraploid cytotypes occur, although Fernández (1987)
reported the numbers 2n = 32, 34, and 39, which are presumably segregants from a pentaploid (2n = 5x = 35). Meiotic studies carried out in some polyploid cytotypes suggest an autopolyploid origin of the complex (Fernández, 1987
; Solís Neffa, 2000
).
In order to understand the evolutionary significance of autopolyploidy in T. sidoides, a biosystematic investigation is in progress in this complex. As a part of this study, in this paper the karyotypes of the five subspecies are described for the first time to (1) investigate the nature of polyploidy in the T. sidoides complex, (2) determine whether karyotypic differentiation has occurred among subspecies, and (3) discuss the results in relation to the taxonomic position of T. sidoides.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
|
) or 3 : 1 absolute ethanol:lacial acetic acid overnight, stored in 70% ethanol and stained using the Feulgen technique, and squashed in 3% aceto-orcein. Slides were made permanent in Euparal by mean of Bowen's (1956)
method.
At least ten metaphases were examined per population, with the five best selected for making idiograms. The length of chromosome arms and satellites were measured on drawings made with a camera lucida. Idiograms were drawn based on mean centromeric index (CI = short arm x 100/total length of the chromosome) and arranged in order of decreasing size. Chromosome nomenclature followed Levan, Fredga, and Sandberg (1964)
, the symbols m, sm, and st designating metacentric, submetacentric, and subtelocentric chromosomes, respectively. Satellites were classified according to Battaglia (1955)
.
The following parameters were estimated in each metaphase plate to characterize the karyotypes numerically: (1) haploid chromosome length (HCL); (2) mean chromosome length (ML); (3) mean centromeric index (CI); (4) ratio of shortest to longest pair (S/L); (5) intrachromosomal asymmetry index (A1) = 1 – [
(b/B)/n]; and (6) interchromosomal asymmetry index (A2) = s/x, where b and B are the mean length of short and long arms of each pair of homologues, respectively, n is the number of homologues, s is the standard deviation, and x the mean chromosome length.
Karyotype asymmetry was estimated using the mean centromeric index, the ratio of the shortest/longest pair and according to the A1 and A2 indices (Romero Zarco, 1986
).
Parameter means were compared by one-way ANOVA after Bartlett's test of homogeneity. Also, Tukey's test was carried out to test differences between each pair of means.
A cluster analysis of the karyotype data was carried out in order to examine karyotype similarity among the subspecies. A data matrix of 8 OTUs (operational taxonomic units) x 23 variables was constructed. For each chromosome pair, length and centromeric index were calculated; and for each genome, mean chromosome length, genome length, mean centromeric index, A1 and A2 indices, and the presence and position of satellites were determined. The NTSYS-PC program developed by Rohlf (1994)
was used to standarize the data matrix, to calculate the average taxonomic distance, and to generate a phenogram. Clustering was performed using the unweighted pair-group method (UPGMA). The distortion of phenogram was measured by computing the cophenetic correlation coefficient (r).
In addition, we also compared karyotype features of T. sidoides with those of other species of the genus using a principal component analysis (PCA). A data matrix of 26 OTUs x 11 variables describe above plus the number of metacentric, submetacentric, and subtelocentric pairs, was constructed using the data obtained in this paper and those obtained from Solís Neffa and Fernández (1993)
and from Solís Neffa (1996)
, which are summarized in Table 4. Standarized data were used for all multivariate analyses.
|
|
RESULTS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
|
|
|
|
The mean chromosome length is the only parameter analyzed that differed significantly among the subspecies (Table 3). The chromosomes are small (Tables 2 and 3; Figs. 1–11); the mean chromosome length for all the taxa is 2.19 µm, varying from 1.96 µm in subsp. holosericea to 2.86 µm in the population S57 of subsp. pinnatifida.
As a whole, karyotypes are symmetrical and generally present gradual differences in size among the chromosomes of each subspecies. The mean centromeric index for the species (CI = 41.48) falls into the metacentric category. Mean values obtained for the ratio of shortest/longest pair (S/L) and interchromosomal asymmetry index (A2) indicate there is little variation among the sizes of the different chromosomes in each subspecies. Data for the mean intrachromosomal asymmetry index (A1) show no sharp differences between the chromosome arms in the different taxa. The A1 and A2 indices are plotted in a scatter diagram (Fig. 12), which shows that the karyotypes of subsp. pinnatifida and subsp. sidoides are the most symmetrical and have the greatest variation in length among their chromosomes. On the other hand, subsp. carnea presents the most asymmetrical karyotype and the smaller values of A2.
|
The UPGMA phenogram based on karyotype similarities is shown in Fig. 13. The cophenetic correlation for it is 0.82, indicating a good fit between the cophenetic value matrix and the mean taxonomic distance matrix. The phenogram obtained shows the following clusters: (1) Turnera sidoides subsp. carnea is separated from the other subspecies because it possesses only one pair of satellited chromosomes and has the most asymmetrical karyotype; (2) subsp. holosericea and subsp. integrifolia show a great similarity of their karyotype characteristics because they share the karyotype formula, chromosome size, degree of karyotype asymmetry, and the presence of two pairs with microsatellites; (3) subsp. pinnatifida and subsp. sidoides have very similar karyotypes; (4) polyploid cytotypes and diploids from northwestern Argentina are grouped together in the same cluster because the polyploids exhibit the same karyotype as diploids but at the tetraploid and hexaploid level; and (5) the diploid population S57 of subsp. pinnatifida is isolated because its mean chromosome length makes it significantly different from the other OTUs.
|
|
|
DISCUSSION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
; Lewis, 1980
). Polyploidy has also played a prominent role in speciation within the genus Turnera as a whole as well as within the T. sidoides species complex (Raman and Kesavan, 1964
; Hamel, 1965
; Barrett, 1978
; Arbo and Fernández, 1983
; Fernández, 1987
; Barrett and Shore, 1987; Shore, 1991a, b
; Solís Neffa, 2000
; Solís Neffa and Fernández, 2001
). Furthermore, available data suggest that T. sidoides represents an example of autopolyploidy (Fernández, 1987
; Solís Neffa, 2000
).
Autopolyploidy apparently occurs less frequently than allopolyploids, but some workers suggest that the occurrence of autopolyploidy may have been underestimated (Levin, 1983
; Soltis and Rieseberg, 1986
). Autopolyploidy was originally defined using cytogenetic criteria and, as such, demonstration of autopolyploids requires a direct analysis of meiotic configurations. In this sense, autopolyploids are usually considered to be recognizable by multivalent chromosome configurations during meiotic metaphases. Morphological and chemosystematic characters may also provide additional information on the origin of autopolyploidy. Data on meiotic behavior obtained from some populations of T. sidoides showed high levels of multivalent formation, with quadrivalents, hexavalents, and octovalents in tetraploid, hexaploid, and octoploid cytotypes, respectively (Fernández, 1987
; Solís Neffa, 2000
). Furthermore, preliminary allozymic and biochemical data are also in agreement with the contention that the polyploid cytotypes are of autopolyploid origin (Solís Neffa, 2000
).
The morphology of chromosomes at mitotic metaphase has often been used as a criterion for distinguishing between autopolyploids and allopolyploids (Stebbins, 1971
). Our karyotype analyses of T. sidoides are in accordance with expectations in autopolyploids. In subsp. carnea, integrifolia, and pinnatifida, diploid and polyploid cytotypes were reported (Fernández, 1987
; Solís Neffa, 2000
; Solís Neffa and Fernández, 2001
). López (1987)
, describing mitotic metaphases of the diploid cytotype of subsp. carnea and integrifolia, did not analyze karyotypes in detail, but observed the presence of macrosatellites and microsatellites in subsp. carnea and integrifolia, respectively. The tetraploid cytotype of both subspecies analyzed in this paper exhibits the same features as in diploids but at the tetraploid level. In subsp. pinnatifida, chromosomes of diploids are grouped into pairs, while in tetraploid and hexaploid cytotypes, they can be grouped into sets of four and six, respectively. On the other hand, in subsp. holosericea and subsp. sidoides, only polyploid cytotypes are known (Fernández, 1987
; Solís Neffa, 2000
; Solís Neffa and Fernández, 2001
). The karyotypes of the hexaploid cytotype of subsp. holosericea and the tetraploid of subsp. sidoides studied are composed of chromosomes grouped into sets of six and four, respectively. Karyotype data, coupled with cytological observation of chromosome configuration at meiosis as well as preliminary biosystematic data (Fernández, 1987
, Solís Neffa, 2000
), lend further support to the existence of autopolyploidy within this species complex. Thus, T. sidoides may be added to the relatively short list of natural autopolyploid plants (reviewed in Soltis and Rieseberg, 1986
).
In Turnera sidoides subsp. pinnatifida, the karyotypes of diploid, tetraploid, and hexaploid cytotypes have been studied. This subspecies is the most widespread of the complex and occurs in southern Bolivia, Paraguayan "Chaco," and southwestern Uruguay to Argentina, where it reaches 39° S. It has a very broad ecological range, both climatically and edaphically, and grows on a wide range of soils. Habitats extend from sea level to mountain regions, ascending to about 2700 m above sea level (asl) (Arbo, 1985, 1987
; Solís Neffa, 2000
). Cytogeographical analysis has shown that diploids and hexaploids of this subspecies occupy disjunct areas and sites with different climatic regimes, while tetraploids are scattered over the entire geographical and climatic range of this subspecies (Solís Neffa and Fernández, 2001
). Our results have shown that diploid populations from northwestern Argentina and from Uruguay have similar karyotype formulae but differ in mean chromosome length and karyotype asymmetry, suggesting that the addition of genetic material may also be involved in their karyotype evolution. The spatial separation of these populations prevents gene flow between them, allowing the fixation of chromosome changes and, therefore, favoring karyotype differentiation. Furthermore, the phenogram (Fig. 13) based on karyotype data indicates that polyploid cytotypes of subsp. pinnatifida are more similar to those of diploid populations from northwestern Argentina than to those of diploids from Uruguay. These results, in agreement with cytogeographic, morphological, and biochemical data (Solís Neffa, 2000
; Solís Neffa and Fernández, 2001
), suggest that these polyploid populations would have originated from an ancestral diploid population with a karyotype similar to those of populations from Argentina.
The results of karyotype analysis suggest that the accumulation of chromosome rearrangements may also be involved in the karyotypic evolution of T. sidoides. Changes in chromosome morphology may either lag behind or precede change in external phenotype (Jackson, 1971
). Further, close morphological similarity of taxa with great differences in the basic karyotype suggests that the cytological differences have been brought about by sudden chromosome rearrangements (Jackson, 1971
). On the other hand, partial correspondence of morphology and karyotypes would imply parallel changes, which probably happened during the diversification of the Turnera sidoides complex because the subspecies are both morphologically and karyotypically differentiated. Morphologically, the leaf shape, the degree of incision of the leaf blade, the color of the flowers, and the expression of heterostyly (Arbo, 1985
) may distinguish the subspecies. Detailed analysis of karyotypes shows a high degree of intraspecific uniformity for all variables measured in the different samples studied; however, the subspecies may be differentiated by the number, type, and position of satellites.
Karyotype and systematics
The results of the karyotype analysis of T. sidoides are in agreement with earlier reports in Turneraceae (Lavia and Fernández, 1993
; Solís Neffa and Fernández, 1993
; Solís Neffa, 1996
). The karyotype of T. sidoides consists almost exclusively of m and sm chromosomes, except for one st pair present in subsp. carnea, pinnatifida, and sidoides. The st chromosomes are very rare in Turneraceae; besides T. sidoides, the only species known with this type of chromosome are T. chamaedryfolia and Piriqueta duarteana (Lavia and Fernández, 1993
; Solís Neffa and Fernández, 1993
).
The subspecies of T. sidoides, like the other species of the genus, present one or two pairs of chromosomes with satellites. Subspecies carnea possesses a macrosatellite in the short arm of an m pair, while subsp. holosericea and subsp. integrifolia have two pairs with microsatellites. Subspecies pinnatifida and sidoides present both macro- and microsatellites.
Chromosomes are small in Turnera (Solís Neffa and Fernández, 1993
; Solís Neffa, 1996
), and this is also true for the subspecies of T. sidoides. The average chromosome length for the genus as a whole is 2.25 µm, and species with a basic number of x = 7 generally possess smaller chromosomes (ML = 1.38 µm) than those with x = 5 (ML = 2.21 µm) and x = 13 (ML= 1.45 µm) (Solís Neffa and Fernández, 1993
; Solís Neffa, 1996
). Although T. sidoides has a basic number of x = 7, its mean chromosome length is in accordance with the mean values observed among species with x = 5.
Turnera is characterized by a moderate degree of karyotype asymmetry and by gradual differences in chromosome size, and T. sidoides follows this rule (Lavia and Fernández, 1993
; Solís Neffa and Fernández, 1993
; Solís Neffa, 1996
). An association between karyotype asymmetry and basic numbers was previously detected in the genus (Solís Neffa and Fernández, 1993
). In this sense, the scatter diagram (Fig. 15) shows that the x = 13 plant (T. chamaedryfolia) has the highest asymmetric tendency and the species with x = 5 have the smallest degree of asymmetry, while species with x = 7 have a moderate degree of karyotype asymmetry. Although T. sidoides has a basic number of x = 7, it showed a level of asymmetry intermediate between those species with x = 5 and x = 7.
|
and Arbo (1985)
, but it has morphological and anatomical features that are extremely different from those of the remaining species (Arbo, 1985
; González, 2000
) and, therefore, justify its exclusion from the series. Unlike the other species of Leiocarpae, T. sidoides is one of the few species without foliar nectaries and is the only one with tuberculate fruits. Furthermore, its seed morphology is extremely different from the remaining species of Turnera because, instead of being reticulate like those of the other species of the series, or striate as the species of other series, it presents irregular crests arranged in lines (Arbo, 1985
). From the cytological point of view, the PCA ordination carried out in this paper shows that Turnera species have very similar karyotypes, but they can be characterized not only by their basic number, but also by mean chromosome length and the degree of karyotype asymmetry. In this sense, although T. sidoides shares the basic number, x = 7, with the other species of the series, its mean chromosome length is in accordance with the values observed among the species of series Turnera (x = 5). Furthermore, T. sidoides is the only species of the series that possesses an st pair in its karyotype and shows an intermediate degree of karyotype asymmetry between species of Leiocarpae and Turnera series. Therefore, karyotype features coupled with morphological and anatomical data suggest that the taxonomic position of T. sidoides in the genus should be revised.
|
|
FOOTNOTES |
|---|
5 Author for reprint requests (viviana{at}agr.unne.edu.ar
)
|
LITERATURE CITED |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
Arbo M. M. 1986 Paraguay, centro importante de especiación en las Turneráceas. Candollea 41: 211-218
Arbo M. M. 1987 Turneraceae. In R. Spichiger [ed.], Flora del Paraguay 6: 1–65
Arbo M. M. A. Fernández 1983 Posición taxonómica, citología y palinología de tres niveles de ploidía de Turnera subulata Smith. Bonplandia 5: 212-226
Arechavaleta J. 1905 Enumeración y descripción breve de las plantas conocidas hasta hoy y de algunas nuevas que nacen espontáneamente y viven en la República Oriental del Uruguay. Flora Uruguaya Tomo II: 119–121. In Anales del Museo de Historia Natural de Montevideo. V. Montevideo, Uruguay
Barrett S. C. H. 1978 Heterostyly in a tropical weed: the reproductive biology of the Turnera ulmifolia complex (Turneraceae). Canadian Journal of Botany 56: 1713-1725
Barrett S. C. H. J. Shore 1987 Variation in breeding systems in the Turnera ulmifolia L. complex (Turneraceae). Evolution 41: 340-354[CrossRef][ISI]
Battaglia E. 1955 Chromosome morphology and terminology. Caryologia VIII: 179-187
Bowen C. C. 1956 Freezing by liquid carbone dioxide in making slides permanent. Stain Technology 31: 87-90[ISI][Medline]
Fernández A. 1973 El ácido láctico como fijador cromosómico. Boletín de la Sociedad Argentina de Botánica 15: 287-290
Fernández A. 1987 Estudios cromosómicos en Turnera y Piriqueta (Turneraceae). Bonplandia 6: 1-21
González A. M. 2000 Estudios anatómicos en los géneros Turnera y Piriqueta. Ph.D. thesis, Universidad Nacional de Cordoba, Córdoba Province, Argentina
Hamel J. L. 1965 Le noyau et les chromosomes somatiques de Turnera ulmifolia L. Mémoires du Muséum National d'Histoire Naturelle Série B, 16: 3-8
Jackson R. C. 1971 The karyotype in systematics. Annual Review of Ecology and Systematics 2: 327-368[CrossRef]
Lavia G. I. A. Fernández 1993 Cariotipos y estudios meióticos en varias especies de Piriqueta (Turneraceae). Bonplandia 7: 129-141
Levan A. K. Fredga A. A. Sandberg 1964 Nomenclature for centromeric position on chromosomes. Hereditas 52: 201-220[ISI]
Levin D. A. 1983 Polyploidy and novelty in flowering plants. American Naturalist 122: 1-25[CrossRef][ISI]
Lewis W. H. 1980 Polyploidy in species population. In W. H. Lewis [ed.], Polyploidy 103–142. Biological Relevance, Plenum Press, New York, New York, USA
López M. G. 1987 Significación morfológica y citológica de la poliploidía en el complejo Turnera sidoides L. Trabajo final de graduación para optar por el título de Ingeniero Agrónomo, Facultad de Cencias Agrarias, Universidad Nacional del Nordeste, Corrientes, Corrientes Province, Argentina
Raman V. S. P. C. Kesavan 1964 Meiosis and the nature of polyploidy in Turnera ulmifolia. Journal of the Indian Botanical Society 43: 495-497
Rohlf F. J. 1994 NTSYS-pc. Numerical taxonomy and multivariate analysis system, version 1.8. Exeter Software, New York, New York, USA
Romero Zarco C. 1986 A new method for estimating karyotype asymmetry. Taxon 35: 526-530[CrossRef][ISI]
Shore J. S. 1991a Chromosomal evidence for autotetraploidy in the Turnera ulmifolia complex (Turneraceae). Canadian Journal of Botany 69: 1302-1308[CrossRef]
Shore J. S. 1991b Tetrasomic inheritance and isozyme variation in T. ulmifolia vars. elegans Urb. and intermedia Urb. (Turneraceae). Heredity 66: 305-312[ISI]
Sols Neffa V. G. 1996 Cariotipos de especies de Turnera (Turneraceae). Bonplandia 9: 121-127
Sols Neffa V. G. 2000 Estudios biosistemáticos en el complejo Turnera sidoides L. (Turneraceae, Leiocarpae). Ph.D. thesis, Universidad Nacional de Cordoba, Córdoba Province, Argentina
Sols Neffa V. G. A. Fernández 1993 Estudios cromosómicos en especies de Turnera (Turneraceae). Bonplandia 7: 101-118
Sols Neffa V. G. A. Fernández 2001 Cytogeography of the South American Turnera sidoides L. complex (Turneraceae, Leiocarpae). Botanical Journal of the Linnean Society 137: 189-196[CrossRef]
Soltis D. E. L. H. Rieseberg 1986 Autopolyploidy in Tolmiea menziesii (Saxifragaceae): genetics insights from enzyme electrophoresis. American Journal of Botany 73: 310-318[CrossRef][ISI]
Stebbins G. L. 1971 Chromosomal evolution in higher plants. Arnold, London, UK
Urban I. 1883 Monographie der familie der Turneraceen. Jahrbuch des Königlichen Botanischen Gartens Berlin 2: 1-152
This article has been cited by other articles:
|
|
 |
|
  S. Truyens, M. M. Arbo, and J. S. Shore Phylogenetic relationships, chromosome and breeding system evolution in Turnera (Turneraceae): inferences from its sequence data Am. J. Botany, October 1, 2005; 92(10): 1749 - 1758. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
: x = 5. 
: x = 7. 
: x = 13. 
: T. sidoides complex