170 Cytologia 28 A Cytotaxonomic Study of the Genus Cyrtanthus R. Wilsenach Department of Botany, University of the Witwatersrand, Johannesburg, Republic of South Africa Received November 19, 1962 The members of the family Amaryllidaceae have relatively large chromo somes and this facilitates the computing of accurate idiograms which are a help in the understanding of the phylogeny of plants. Many workers, e.g. Taylor (1925), Heitz (1926), Sato (1938, 1942), Gouws (1949), Mookerjea (1955), Sharma and Bhattacharya (1956), and Sharma and Bal (1956), have studied the karyology of amaryllids in order to obtain a clearer picture of their relationships. The phylogenetic relationships of the amaryllid groups as proposed by Gouws (1949), for example, must be considered as a great im provement on the earlier systems which were based only on morphological characters. Unfortunately the karyological data are still far from complete, and serious discrepancies still exist as regards the interpretation of results. This is especially true of the genera Cyrtanthus, Vallota and Anoiganthus. Taylor (1925), Sato (1938, 1942), Gouws (1949), Mookerjea (1955) and Ising (1962), have studied several species belonging to these genera. These authors differ concerning the relationships of these plants and also as to their basic chromo some numbers, the diploid numbers of 16, 18, 20 and 22 having been reported. The present investigation attempts to throw further light on this problem by an extended study of the genus Cyrtanthus, which has South Africa as its main centre of distribution. Intention was, firstly, to find out more about the variation in chromosome number reported by other workers, and secondly, to correlate the karyological observations with morphological data to obtain a better understanding of the phylogeny of the genus. Material and methods Almost all the plants were collected in their natural habitat and replanted in a sandy loam in 4 "and 6" pots. The meiotic divisions of the pollen mother cells were found to take place before the inflorescences appear, with the result that preparations of meiotic stages would result in the destruction of numerous bulbs, and this could not be afforded. For this reason only mitotic stages in the root tips were recorded. The root tips were fixed in Craf (Randolph 1935) for 24 hours , except in the case of C. carneus in which case the root tips were fixed for about 1 hour in 3 alcohol: 1 propionic acid. The root tips of C. herrei were fixed in both fixing fluids. After fixation in Craf the root tips were transferred 1963 A Cytotaxonomic Study of the Genus Cyrtanthus 171 to 70% ethyl alcohol and dehydrated in the ethyl alcohol-normal butyl alcohol series according to Randolph (1935), embedded in paraffin wax, and cut 16 18 microns thick. Sections were stained in a 1% aqueous solution of crystal violet, following the method described by Smith (1934), and mounted in canada balsam. Six year old preparations show very little sign of bleaching. After fixation in 3 alcohol: 1 propionic acid the root tips were squashed and stained according to Warmke's (1935) prescription, but propionic acid was substituted for acetic acid. In order to make these slides permanent the methods of McClintock (1929), Conger and Fairchild (1953) and Cave and Pocock (1951) were tried. None of these proved to be without drawbacks, with the result that the slides were made semipermanent by sealing off the sides of the coverslip with "nail varnish". Table 1. Species investigated Description of somatic chromosomes In computing the idiograms of each species several metaphase plates were compared. The chromosomes of the species investigated are relatively long, with the result that the chromosome arms in polar view twist both towards and away from the observer, and this had to be taken into account. This was done by determining how far the chromosome arm twists towards the pole, and this distance is often as much as 6 microns. In the idiograms the chromosomes are generally arranged from the longest chromosome to the shortest. This is, however, not true of all the idiograms 172 R. Wilsenach Cytologia 28 e.g. C. parviflorus chromosome G is longer than chromosome F (Idiog. 9). These exceptions occur only to facilitate comparison of the different idiograms, and it infers a homology between e.g. chromosome G of C. par viflorus and the G chromosomes of the other species. The chromosomes were arbitrarily divided into long, medium and short, Table 2. Genome formulae and chromosome numbers of species investigated 1963 A Cytotaxonomic Study of the Genus Cyrtanthus 173 Figs. 1-18. Somatic chromosomes (2n) of Cyrtanthus species. 1, C. erubescens, 16. 2, C. thorncroftii, 16. 3, C. huttonii, 16. 4, C. species (from Northern Transvaal), 16. 5, C. mackenii, 16. 6, C. mackenii var. cooperi, 16. 7, C. brachyscyphus, 16. 8, C. parviflorus, 16. 9. C. ochroleucus, 16. 10, C. smithiae, 16. 11, C. helictus, 16. 12, C. obliquus, 16. 13, C. carneus (fixed in 3 alcohol: 1 propionic acid), 16. 14a, C. herrei (Craf fixation), 16. 14b, C. herrei (fixed in 3 alcohol: 1 propionic acid), 16. 15, C. contractus, 16. 16, C. sanguineus, 16. 17, C. galpinii, 16. 18, C. balenii, 16. 174 R. Wilsenach Cytologia 28 for which the letters 1 (longum), m (medium) and b (brevis) are used. In addition the letters V, L and f are used to indicate the position of the primary (kinetic) constriction: V for a median or near median constriction; L for sub median; and f for subterminal, i.e. when the proximal arm is 1/3 or less of the length of the distal arm. Discussion The karyology of the genus Cyrtanthus. Sato (1938) recorded 2n=22 as the somatic chromosome number of Cyrtanthus obliquus. According to Taylor (1925) the diploid number of C. parviflorus is 16, and Gouws (1949) reported the same number for C. tuckii var. transvaalensis. Flory (1955) investigated six species, viz. C. angustifolius, C. falcatus, C. ochroleucus (=C. lutescens), C. mackenii, C. O'Brieni and C. sanguineus, and quoted the diploid number 16 for all. Mookerjea (1955) studied C. sanguineus and a horticultural variety called Cyrtanthus "ifafa lily", and reported the diploid chromosome numbers as 18 and 20 respectively, and stated that it was not possible to make any sug gestions regarding the basic number for the genus. In the present work the three species, namely C. obliquus, C. parvi florus, and C. sanguineus were re-investigated and 16 chromosomes were regularly present. This is also true of all the other species investigated here, with result that the number 16 can be considered as the normal diploid number of this genus. Ising (private communication) made the following remark: "The number 2n=22 reported for C . obliquus... I think is wrong, as I found 2n=16 in my plants". The counts of 22, 20 and 18 reported by Sato (1938) and Mookerjea (1955) are difficult to interpret, unless they resulted from incorrect technique. It is apparently the first time that C. helictus, C. huttonii, C. mackenii var. cooperi, C. contractus, C. balenii, C. galpinii, C. brachyscyphus, C. smithiae, C. herrei, C. carneus, C. thorncroftii and C. erubescens have been investi gated cytologically. The idiograms of the different species show a fair degree of resemblance but vary from very asymmetrical ones to more symmetrical ones (e.g. C. brachyscyphus and C. obliquus respectively). Chromosome A is V-shaped or almost so in most species and varies in length between 15-19 microns. Chromo some B and C are often similar, are usually fairly long (10-18 microns) and usually have sub-median kinetic constrictions. Chromosome D is the most variable one in the complex and varies in length between 8 and 13 microns. Chromosomes E and F are often very similar and usually have sub-terminal kinetic constrictions, their length varies between 6-12 microns. Chromosome G is usually of mV type, the most striking exception being C. brachyscy phus, where this chromosome has a sub-terminal kinetic constriction. Its length varies between 5 and 10 microns. Chromosome H is always of the bf 1963 A Cytotaxonomic Study of the Genus Cyrtanthus 175 Idiograms 1-22. Idiograms of Cyrtanthus, Anoiganthus and Vallota species. 1, Anoigan thus breviforus (according to Gouws, 1949). 2, Cyrtanthus erubescens. 3, C. thorncroftii. 4, C. huttonii. 5, C. species (from Northern Transvaal). 5, C. mackenii. 7, C. mackenii var. cooperi. 8, C. brachyscyphus. 9, C. parviflorus. 10, C. ochroleucus. 11, C. smithiae. 12, C. helictus. 13, C. obliquus. 14, C. carneus (fixed in 3 alcohol: 1 propionic acid). 15, C. herrei (Craf fixation). 16, C. herrei (fixed in 3 alcohol: 1 propionic acid). 17, C. con tractus. 18, C. sanguineus. 19, C. galpinii. 20, C. balenii. 21, Vallota purpurea (accord ing to Gouws, 1949). 22, Cryptostephanus vansonii (according to Gouws, 1949). 176 R. Wilsenach Cytologia 28 type or nearly so, it varies in length between 5 and 10 microns. (The lengths quoted refer only to the determinations based on Craf-fixed material). Phylogeny. The genus Cyrtanthus has a rather interesting history. The first two species recorded were described under the generic name Crinum by Linnaeus in 1781. Eight years later the genus Cyrtanthus was created by Aiton (1789) to accommodate these species, which had curved flowers. Herbert originally separated the species which are now considered as belonging to Cyrtanthus into three genera viz. Monella, Cyrtanthus and Gastronema. When C. carneus became known to him he realised that it represented a link between Cyrtanthus and Monella, with the result that he combined these two genera. Baker in 1888 combined Gastronema with Cyrtanthus and the genus was composed of three sub-genera, Cyrtanthus proper, Monella and Gastr onema.