Amer. J. Bot. 76(5): 657-665. 1989. CHROMOSOMENUMBERS IN BROMELIACEAE' GREGORY K. BROWN AND AMY JEAN GILMARTIN2,3 Departmentof Botany and Rocky Mountain Herbarium,University of Wyoming, Laramie,Wyoming 82071; and 2Departmentof Botany and Marion Ownbey Herbarium,Washington State University, Pullman, Washington99164 ABSTRACT Eighty-threechromosome counts are reportedfor 72 taxa of the Bromeliaceae.Fifty-eight of these counts are the firstknown chromosomenumber reports for their respectivetaxa. A model of chromosomalevolution in the Bromeliaceae(n = 25) is presented.The model is parsimonious and consistent with existing data on meiotic chromosome numbers within the family and in the closely related Velloziaceae (n = 9). Two hypothesizedpaleodiploids (n = 8 and n = 9) hybridizedto form a tetraploidthat in turn hybridizedwith the n = 8 lineage. The resultantn = 25 is the extant base number for the family. Two alternativehypotheses could explain the unique extant base number(n = 17) for Cryptanthus:1) Cryptanthusrepresents the paleotetra- ploid level, i.e., prior to the second round of hybridization,or 2) the lower numberrepresents the result of a more recent series of aneuploidreductions from n = 25. Given the existence of intergenerichybrids involving Cryptanthus,aneuploid reduction is the morelikely interpretation. RECENT RESEARCH concerning Bromeliaceae ploidy, chromosome size bimodality, and the systematics and evolution (e.g., Brown and correlationof nonconcordancein meiotic and Gilmartin, 1984, 1986; Gilmartin and Brown, mitotic chromosome numbers with the epi- 1985, 1986b) has sparkedrenewed interest in phytic mode of growth. the study of Bromeliaceae chromosomes and The purpose of this paper is to describe re- chromosome evolution. Past chromosome sults of an ongoing meiotic chromosome num- number surveys in the family (i.e., Lindschau, ber survey within the Bromeliaceae, and es- 1933; Gauthe, 1965; Weiss, 1965; Sharmaand pecially subfamily Tillandsioideae. We also Ghosh, 1971; Till, 1984) have relied mostly presenta model for chromosome base number on mitotic material.The only major exception evolution for the family that is consistent and to this was Marchant (1967) who utilized parsimonious with existing data. meiotically active microsporocytes. There is great variability in reported mitotic chromo- METHODS AND MATERIALS- Floralbuds were some numbers (Brown and Gilmartin, 1986), collected in the field, or obtained from culti- and lack of concordance between mitotic and vated material at Marie Selby Botanical Gar- meiotic numbersfor some taxa within the fam- dens, Sarasota,Florida (SEL).Buds were fixed ily. This variability in mitotic number is re- in field-mixedFarmer's solution (100%EtOH: flected in the variable interpretationsof chro- glacial acetic acid; 3: l/v:v) to which a drop of mosome base numbers for the family. Brown saturatedaqueous ferric chloride (FeCl3-6H20) and Gilmartin (1986) summarized the pre- had been added. The latter enhances chro- vious controversy over base number deter- mosome stainability. After a minimum of 24 mination for Bromeliaceae,and discussed the hr, fixed buds are transferredto 70% EtOH. current level of knowledge concerning poly- See Gilmartin and Brown (1986a) for a com- plete description of the field collaboratornet- ' Received for publication 13 October 1987; revision work and its operation. accepted 28 October 1988. For chromosome squash preparations, in- We thank a dedicated group of field collaborators, with- dividual anthers were removed from the bud out whom this project would not have been possible: James Ackerman, Puerto Rico; Stephan Beck, Bolivia; Olga Be- in 70% EtOH and transferredto a pool of 1% navides, Colombia; Elizabeth Bravo, Ecuador; David acetic carmine on a microscope slide. While in Brunner, Paraguay; I. Chacon, Costa Rica; Hermes Cua- the stain, the anther is cut transverselyin half. dros, Colombia; Linda Escobar, Colombia; Gert Hatsch- Using ultrafine-tippedneedle and forceps, the bach, Brazil; Stephen Koch, Mexico; Gustavo Martinelli, Brazil; Fernando Ortiz, Ecuador; Isidoro Sanchez Vega, sporogenous masses are squeezed from each Peru; and Rosa Subils, Argentina. Expert technical help microsporangiumthrough the median trans- was supplied by Carol Annable. We thank Ron Hartman, verse cut. The sporogenous masses are posi- Don Hauber, and two anonymous reviewers for their com- tioned toward the center of the stain pool and ments. This work was supported by collaborative research grants BSR-8607 187 (GKB) and BSR-8407573 (AJG) from a coverslip and gentle finger pressure are ap- the National Science Foundation. plied. The preparationis further flattened by 3 Deceased 10 February 1989. passingthe slide throughan alcohol flame sev- 657 658 AMERICAN JOURNAL OF BOTANY [Vol. 76 eral times. Heating the slide helps to rupture far homogeneous for the base number of x = the callose that encapsulates the microsporo- 25 (also see Brown and Gilmartin, 1986). All cyte. Squasheswere examined with phase con- repeatchromosome number reports made here trast microscopy, and documented using Ko- for Pitcairnioideae taxa corroborate one or dak Technical Pan 2415 and drawings. As more previous counts (i.e., Lindschau, 1933; standard practice, a minimum of five micro- Di Fulvio, 1967; Marchant, 1967; Brown et sporocytes with unambiguous meiotic figures al., 1984). With the addition of new genera (usually in diplotene, diakinesis, metaphase I, Brewcariaand Steyerbromeliapublished since or metaphase II) serve as the basis for chro- Smith and Downs (1974), Pitcairnioideaecon- mosome numberdetermination. In caseswhere tains 15 genera.Chromosome data are lacking a new chromosome number (e.g., Tillandsia for the followinggenera: Abromeitiella (2 spp.), leiboldiana) or abnormality (e.g., B-chromo- Ayensua (1 sp.), Brewcaria(1 sp.), Brocchinia somes or fragments)was encountered,as many (18 spp.), Connellia (4 spp.), Cottendorfia(24 as 18 unambiguous meiotic figureswere doc- spp.), Encholirium (12 spp.), Navia (74 spp.), umented. Voucher herbarium specimens are and Steyerbromelia(1 sp.). at WS unless otherwiseindicated (see Table 1). All graphicdocumentation of chromosomes is Tillandsioideae-With over 800 species in at RM. The nomenclaturefollowed here is that six genera(Catopsis, 19 spp.; Glomeropitcairn- of Smith and Downs (1974, 1977, 1979). ia, 2 spp.; Guzmania, 126 spp.;Mezobromelia, 3 spp.; Tillandsia,410 spp.; Vriesea,250 spp.), RESULTS-Eighty-threechromosome counts this is the largestof the subfamilies. Published are reportedfor 72 taxa (Table 1). For the most chromosome number information now is part, reports are either the first known pub- available for all but Mezobromelia. The pri- lished chromosome number, or represent a mary focus of this research is Tillandsia, the previously unreported number for a taxon. largestgenus in the family. Representativesquash preparations are shown A comparison of published chromosome in Fig. 1-4. number data for Tillandsia (Brown and Gil- martin, 1986) has revealed a strikingdiscrep- Bromelioideae-Approximately 15%of the ancy between mitotic (root tip) and meiotic ca. 570 specieswithin Bromelioideaeare known chromosome numbers. Prior attempts to dis- by at least one chromosome number report. cover trendsof chromosomalevolution within Unfortunately, these reports are not evenly Tillandsioideae(e.g., Lindschau,1933; Gauthe, spreadacross the 27 generarecognized by Smith 1965) had been hinderedby this variability in and Downs (1979). Most come from five gen- mitotic chromosome number reports (see era (Aechmea, Billbergia, Cryptanthus,Neo- Brownand Gilmartin, 1986, for additionaldis- regelia, Nidularium), while 14 (51%) of the cussion). An explicit goal of our researchhas genera are unknown chromosomally (Andrea, been to determine the level of meiotic chro- 1 sp.; Androlepis, 1 sp.; Araeococcus, 5 spp.; mosome numbervariability within Tillandsia. Disteganthus, 2 spp.; Fascicularia, 5 spp.; The count of n = 22 for T. complanata(subg. Fernseea, 1 sp.; Greigia,26 spp.;Hohenbergia, Allardtia)differs from an earlierreport (Brown 40 spp.; Hohenbergiopsis,1 sp.; Neoglaziovia, et al., 1984; n = 20). Tillandsia complanata is 2 spp.; Ochagavia, 3 spp.; Orthophytum, 17 a wide ranging forest epiphyte (Greater An- spp.; Ronnbergia, 8 spp.; Wittrockia,7 spp.). tilles, CostaRica to Bolivia andN. Brazil;Smith Except for Cryptanthusand Aechmea til- and Downs, 1977) and may comprise cyto- landsioides (Martius ex Schultes f.) Baker, all geographicraces. These two counts correspond Bromelioideae genera thus far studied appear closely to the known northern and southern to have a meiotically established extant base limits for this species. number of x = 25 (see Brown and Gilmartin, The report for T. leiboldiana (Fig. 2) is the 1986). Cryptanthusis anomalous in having a first record of n = 19 within the family. This, base of x = 17 (Marchant, 1967) and the pos- and the T. complanata chromosome numbers sible significance of this is discussed later. presumably arose via dysploidy from an an- Aechmea tillandsioides (n = 21; Marchant, cestral n = 25. We view such chromosome 1967) would appear to be an aneuploid deriv- numbersas being derived. The two reportsfor ative. T. complanata and the remaining 13 reports for species of the subg.Allardtia presented here Pitcairnioideae-Chromosome counts are are the only known chromosome number re- available for six pitcairnioid genera (Deuter- ports for this subgenus. ocohnia, Dyckia, Fosterella, Hechtia, Pitcair- The chromosome number