382 s. Mr. 1. Bot. 1997,63(6) 382- 399 The systematic value of the surface mitromorphology and anatomy of cypselae of some members of the Senecioneae, Liabeae and Vernonieae (Asteraceae) P. Leszek, D. Vincent' and Sharon L. Wilson Oepartment of Botan), Un iversity of the Witwatersrand , Private Bag 3, Wits, 2050 Repub li c of South Africa email: [email protected] Recl!i\:ed 23 April /997; revised 26 Augusl /997 Twenty one characters of the surface micromorphology and 12 characters of the anatomy of cypseJae of species in the tribes Liabeae (Uabum disc%r, L. ovalum), Senecioneae (subtribe Senecioninae: Senecio inaequidens, S. madagascariensis, S. vulgaris, Bra~1yglottis repanda, subtribe Blennospermatinae: Abrotanella emarginata, A seapigera , Blennosperma nanum) and Vernoneae (Vern onia oligoeephala, V. paueiflora , V. poskeana) were investigated using light microscopy (LM) and scanning electron microscopy (SEM). The data we re analysed phenetically with the assistance of cluster (SAHN) and principal component analyses. Both sources of data were biologically very informative. Taxonomically, the SEM data was most useful in recognising discontinuities between the taxa followed by the combined SEM and LM data sets. The placement of the Blennospermatinae in the Senecioneae is confirmed as being systematically discordant. Keywords: Cypselae, micromorphology, anatomy, Asteraceae. 'To whom correspondence should be addressed. Introduction muci laginous cells (Roth 1977). The Compo sitae or Asteraceae is one of the largest and most Nevertheless, characters dealing with surface micromorphol­ familiar fa milies of fl owering plants and has been researched for ogy and anatomy of cypseJae of the As teraceae have not been centuries (Heywood el al. 1977). i nv ~t i gated very much. The pu rpose of thi s study of Asteraceae A diverse range of characters has been used to delimit the fam­ carpology was primarily to evaluate a wide range of micromor­ ily. Micromorphol0gical characters of the florets have played an phological and anatomical characters of the cypselae of certain important role in the systematics of the Asteraceae since the members of the Senecioneae. Liabeae and Vernonicae (A ppendix investigations of Cassini (Wagenitz 1976). The basic plan of the I). Furthermore, the data was used to elucidate the systematic wood anatomy of the family has also provided useful characters, affinities of the taxa investigated as the systematic affinity of although li ttle intertribal variation has been found (Carlquist these three tribes has been the subject of much argum ent (Bremer 1966). Embryology (e.g. Pandoy & Singh 1980), phytochemistry 1994). (e.g. Robins 1977), cytology, palynology (Vincent & Getliffe Norris t989), mi cromorphol ogy (e.g. Nordenstam 1978; Vincent Materials and Methods & Getliffe 1988, 1992; Wagenitz 1976; Wetter 1983) and 111 01ec­ The choice of the thrce tribes included in this study is based prima­ ular studies (e.g. Baldwin 1992, 1993) have all provided consid­ rily on recent argument concerning the systematic affinitie s of these erable data for systematic studies of various taxa in the three tribes (B reill er 1994). The tax a investigated are n:pL.;clltatiws Asteraceae. of the tribes Senecion!'!a!'! (subtribcs Se necioninae & Bknnospc.:nna­ However, the anatomy of the fruits (cypseJae), sometimes tinae), Liabeae and Vernoneae (Appcndix I). erroneously referred to as achenes (Pope 1983), has not been As little or no information is avail abh:: as to \vhich species is mor· investi gated widely in the fam il y. According to Wagenitz (1976) phological\ y the most re presentat ive of each of the tribes and suh­ the study of the fruit anatomy (termed carpology) is a promising tribes investigated (Appendix I). it was decided to obtain eypselae fie ld for elucidative studies on the systematics of the Asteraceae. from the type species of the type genu s wherever po ss ible. nolwith· The systematic importance of cypsela anatomy of the Asteraceae standing the fact that the type species is no! necessarily the mosl rep­ has been demonstrated in the Anthemideae by Bruhl and Quinn resentative taxon of the genus. Cypselae or putati vel y closely all ied (1990) and Kallersj6 ( 1985). Roth (1977) has provided consider­ taxa to thc various type species were also sampkd. The he rbariulll able evidence on the use of cypsela anatomy data in systematics. material investigated was obtained from PRE and K (Append ix 1). Bremer (1987) stated that cypselae provide a wealth of character Intraspecific variation was noted by samrling. cypselac from two or information at the lower taxonomic levels and that variation in three herbarium specimens of each of the speci es studied. morphology is so great that it is not very useful at the tri bal level. Note that the use of the term cypseJa here is as defined by Scanning Electron Microscopy (SEM) Jackson (1949). Dehydrated cypsdae from herbarium speci mens were secured The cypsela is a special form of dry indehiscent fruit in which directl y on to aluminium viewing stubs using doubk-sided tape. The the seed coat (testa) and fruit wall (pericarp) are ti ghtl y attached stubs were coated with gold/palladium lIsi ng a Polaron sputter coaleI' to one another and is characteri stic of the family Asteraceae and viewed with a .k ol JSM 840 scann ing ek ctro n mic roscope. Data (Roth 1977). were recorded from the photographs that were taken. Differences in The cypselae of the Asteraceae are usually elongate in shape, magnification \verc taken into accollnt when recording the dat a. and are often compressed and show a characteristic scu lpturing on their surface. Appendages are usually present and may be in Light Microscopy (L M) th e form of enl arged ribs, wi ngs or hooks, specialized epidermal Several cypselac frolll each specimen were rehydrated in warlll outgrowths sll ch as protuberances, special trichomes, or water (40°C) on a hot tray lor three days and then Cllt in hal r. S. All'. J. Bot. 11)97, (13(6) .183 Figures ,-6 Scanning electron micrographs of cypsclae: L S. madagascariensis; 2. S. inaeqllidells: 3 . S vII/Karis: 4. A. scapi~(!ra: 5. A.ell1argl1lala: 6. B. repanda. Scale: bars: 0. 1 mm. trallsvt!rsd~. 10 fm:ilitale rl!l1etratioll or the fixatives. Fixation was most inslances nominal-level (non-rarametric) coding \vas lls(:d for dnnc in formalin i1ceto-alcohol (fAA) overnight. The samples wen: tl1(: characters. Standard ization ellnhJcd each <.:haractcr to contribute then suojectcd 10 a graded alcohol (ethanol) dehydration series toward the ovt:rall re~c111blance illver~dy in proportion to it s varia­ (70~o. ~W% . 90% & 95%) for 30 m in each and then twice in absolute bi li ty amongst the enti n: set or OTU·s. Dis tance and corrdati on alcohol !(lr J() min ellcll time. Initial embedding was in Spurr's resin. matrices were computed for each matrix llsing the SIMINT suhrou­ but it was found that thl! resin failed to infiltrate the ti ssues suffi­ tine, to dt:tcrmine the similarity ofth~ taxa in each matri x in terms of cientl) r(lr sl!ctiolling and viewing. Subsequent cmbcdding was in distance and correlation. This gives an indication of the inter-taxon LR White resin. Dehydrated cypselac: were removed rrom Ihe abso­ phenetic relationship or atlinit)' ( Dunn & Ewrilt 1982). The SAHN lu te alcohol anti infiltrated in LR White resin:absolule alcohol s ubroutine was used to r crform various clusteri ng patterns (e.g. (50:50) fix 20 m in and then in resin:absolute alcohol (67:33) for a UPGMA, WPGMA). The CONSENSUS subroutine was used to further 20 min. Finally the cypselae were embedded in pure resin produce a consensus trce when st!veral. equally parsimonius trees overnight after being placed in a vacuum oven at 100 mbar for 4-5 were generated. The COPH subroutine was llsed to test the goodneSS hours of lit or lhe clustering mntrices to the original distance or wrrdation Tnll1::\verse ::\ecliolls ( 1.0-2.5 pm thick) \H!re cut using II Reichert matrices. The cophen(:tic correlation coefficient (r) was computed 2030 microtome. In most cases good quality sections wen: obtained for each data set. Principle Componellts Analysis (PCA) (3 c()m rO~ (see Appendi;.; I for list of spec imens that yielded good quality sec­ nents) was performed to produce a visual n:presentation of Ihe puta­ lions for U\-'O. Se<..' tillIlS were stained in Safran in and Fast Green tive phenetic relationships amongst the tnxa (OTU's) as well as (Johan::\cn 1<J40). All observations were made using a Zeiss MC63 determining which characters are most imrortClnt in defining these photo microscore. phenetic relationships hetwcen tht: taxa in low-dimensional space. Choice of characters investigated Results and Observations ('wenty one characters (Appendix 2 and 4) were recorded for t!ach Characters and Character States (Appendix 2) of the specimen (Appendix I) using the SEM photographs. TwelVt! charac­ cypsela micromorphology recorded via the SEM study ters (Appendix 3 and 5) \ven: recorded using the LM photographs. (character state abbreviated to 'c/state'). Analysis of data Shape of cypselae (SEM Char. I): The variation in the shape of the cypselae of the taxa investigated was represented by 5 char­ rhree t x n (taxon )( character) data matrices were developed (SEM acter states. The specimens of .S. l1Iadagascariensis (Figure I) data. LM data ami combined SEM & LM data - the combined data and (Figure 3) had narrowly elliptic cypselae (c/state matri;.; is not provided here). Each data matrix was analysed lIsing S vlIlgaris the nlllnt!rical phenetics programme NTSYS-pc (Rohlf 1988). The I). Those of S. inoeqllidens (Figure 2) were ell iptic (c/state 2) - data \vere standardized (Sneath & Sakal 1973) lIsing the STAND having a greater width than those described by c/state I .