A Hexaploid Wild Form of Ipomoea Was Collected by the Author in Mexico in 1955

Bot. Maj. Tokyo 84: 377--387, 1971

Evolution and Domestication of the Sweet Potato

ICHIZO NISHIYAMA

Hazamacho 18, Shugakuin, Sakyoku, Kyoto 606

The discovery of a direct ancestor, Ipomoea trifida (6x), of the sweet potato (6x) in 1961-1962 has been followed by a further finding of its diploid (I, leucantha) and tetraploid (I. littoralis) predecessors. On the basis of cytogenetical study it was reasonably assumed that I. littoralis (4x), I. trifida-(3x)-6x, I. trifida (6x) and I. batatas (6x), sweet potato, are autoploids derived rather from the doubling of a set of 15 chromosome pairs (genome B) of I. leucantha (2x) than from segmental alloploidy. On the other hand an artificial hexaploid, I. littocantha, synthesized from I. leucantha (2x) and I. littoralis (4x) was proved to have the same genome constitution as I. trifida-(3x)-6x and sweet potato, their Fl hybrids having 45 bivalents or modified configurations. The hybrids were fertile. Another artificial hexaploid, I, lacunocilis, derived from I. lacunosa (2x) and I. gracilis (4x) was self-fertile, but usually did not hybridize either with I. trifida or sweet potato. The phylogenetic relationship was fairly well supported by the pattern of the mating system in interspecific crosses.

A hexaploid wild form of Ipomoea was collected by the author in Mexico in 1955. It was found to represent I. trifida (H.B.K.) Don. Based on the morphological, cytological and genetical evidences, this wild plant was considered to be the direct progenitor of the sweet potato, I. batatas (L.) Lam. (Nishiyama, 1961; Nishiyama et al., 1961a, b; Nishiyama and Teramura, 1962). It goes without saying that it corres- ponded very well to our long known conception of an ancestral type of sweet potato. The close relationship was confirmed by Jones (1967) on the evidence of crossing experiments and cytological observations of F1 hybrids between sweet potato and the wild form. On the other hand, as already reported by some workers, Jones also found that some sweet-potato seedlings exhibited occasionally such wild characteristics as nonedible roots, winding or climbing habit stems and pubescence on stems, petioles, peduncles and leaves. These findings have led him to a different conclusion from ours, namely that I. trifida (K123) is not an ancestor but a segregate of the sweet potato. In recent years a research project of improving the sweet potato by using the germ plasm of a wild species has made much progress due to the co-operation of several institutes. One of the most remarkable advancements has been made in the understanding of the nature of polyploidization in the sweet potato. These new results including those obtained by Jones (1967) seem to support the view of the present author. This paper mainly deals with a brief summary of up-to-date investigations made in Japan contradicting Jones' view. 378 I. NISHIYAMA Vol. 84

Materials The main materials used in this research were sweet potato (Ipomoea batatas (L.) Lam.) and the following closely related species : Ipomoea leucantha (strain no. K221, 2x), I. littoralis (K233, 4x), I. tri fida-3x (K222, 3x), I. trifida (K123, 6x), I. lacunosa (K61, 2x), and I. gracilis (K134, 4x). All of the wild species were collected in wastelands or fields by I. Nishiyama, M. Kobayashi and M. Muramatsu during their tour in Mexico, and by E. Matuda, Biological Institute, University of Mexico. They were then grown inn pots and propagated vegetatively or by self- or crosspollination. I. trifida- 3x represents triploid seedlings obtained from a seed sample collected in Mexico. The plants closely resembled hexaploid I, trifida in general appearance. They gave a number of hexaploid progeny, designated by I. trifida-(3x)-6x, from artificial out- crossing. The hexaploids might have been produced by the union of unreduced male and female gametes. Two amphiploids (6x) were derived from hybrids, I. littoralis (4x) X I. leucantha (2x), and I, lacunosa (2x) X I. gracilis (4x), by treatment with colchicine and were designated I. littocantha and I. lacunocilis, respectively.

Results and Discussion Basic ancestors of I. trifida and sweet potato : Recently Shiotani et al. (1970) reported extensive preliminary observations on the meiotic pairing of chromosomes in PMCs of inter- and intraspecific hybrids within the genus Ipomoea. By cytological evidence it was proved that I. leucantha (2x), I. littoralis (4x), I. trifida-3x and I. trifida (6x) are closely related to I. batatas (6x), the sweet potato. It was further confirmed that synthesized hexaploid I. littocantha and induced I. trifida-(3x)-6x are

Table 1. Meiotic chromosome configurations in interspecific Evolution of the Sweet Potato 379 most probably forms of I. trifida (6x). Table 1 summarizes meiotic chromosome configurations in some off the interspecific hybrids and their parents, variable numbers of bivalents, univalents and multiple chromosome associations were usually observed. The average and range of number of bivalents per FMC was computed by counting tri-, quadri-, penta- and hexavalent complexes as one, two, two and three bivalents, respectively. Fig. 1 indicates only the average and the expected number of bivalents per PMC in certain hybrids. The results give important information in relation to the phylogeny of the sweet potato, namely, A. In all hexaploid hybrids between four species, I. batatas, I. trifida, I. trifida-(3x)-6x and I. littocantha, 45 or nearly 45 bivalents were found, although some meiotic irregularities occasionally occurred. This clearly shows that these hexaploid species have the same genome constitution. Also it is certain that the basic predecessors of the hexaploids were I. leucantha (2x) and I. littoralis (4x), since I. littocantha (6x) has been artificially synthesized from them. I. trifida-3x may be a natural hybrid between them. According to Sakamoto and Miyazaki (1970) 34-36% of hybrid seeds was obtained in their reciprocal crosses. B. Based on the cytological aspects of the triploid hybrid, I. leucantha (2x) X I. littoralis (4x), it can be said with certainty that 15 bivalents were formed by allosyndesis, and a few extra bivalents were sometimes produced by autosyndesis between the remaining univalents. C. It was especially noted that a tetraploid hybrid, I. trifida-(3x)-6x x 1. leucantha (2x), had nearly 30 bivalents or 28.5-29.8 bivalents on the average instead of 15 bivalents and 30 univalents. The hybrid was fertile and gave rise to F2 progeny

hybrids of Ipomoea and parental species'. 380 I. NISHIYAMA Vol. 84

Fig. 1. Average number of bivalents in interspecific hybrids of Ipomoec. The figures in parentheses show the expected numbers of bivalents formed by allosyndesis. * The hybrid had 93 somatic chromosomes (see Table 1). with 60 somatic chromosomes. It is certain that 15 chromosomes of I. trifida-(3x)- 6x paired with those of I. leucantha and the remaining 30 chromosomes of the former formed 15 bivalents. For analysis of the genome constitution, the genomes of I. leucantha, I. littoralis and natural hexaploids involving I. trifida-(3x)-6x are preliminarily designated as B1B1, B1B1B2B2 and B1B1B2B2B3B3,respectively. Based on a careful investigation of chromosome pairings illustrated in Fig. 1 it is certain that chromosomes of B1, B2 and B3 can pair almost normally with each other. However, it seems that a large number of chromosomes of these genomes are similar and a few differ by one or two non- homologous segments, but nevertheless maintain enough homology. On the other hand a high fertility was observed in tetraploid hybrids between I. trifida-(3x)-6x and I. leucantha (2x) and hexaploid hybrids between the amphiploid littocantha and I. batatas. These facts suggest that the tetraploid and hexaploid species arose by doubling of the I. leucantha genome. Accordingly, the genomic formulae BB, BBBB and BBBBBB should be given to I. leucantha, I. littoralis and I. trifida involving I. trifida-(3x)-6x and I. batatas, respectively. The frequency of multivalents at meiosis of interspecific hybrids seems not to be available for determination of the type off polyploidy in the present study, because of the fact that the multivalent frequency differs markedly in autoploids induced from different plant species. Based on the chromosome morphology, Sharma and Datta (1958) concluded that the sweet potato is of alloploid origin. Later Nakajima (1963) also made a karyotype analysis in a number of I pomoea species. From these results it appears that a definite Evolution of the Sweet Potato 381

Fig. 2. I pomoea littoralis (K233), 2n=60. Photo. Aug. 20, 1971. analysis of chromosome morphology of I pomoea, especially I. batatas, was very difficult, because of the small size and the large number of chromosomes. Seed fertility (4.1%) was greatly reduced in autotetraploid I, leucantha, whereas the original diploid produced 86.8% and the natural tetraploid, I. littoralis, 52.5% seeds (Sakamoto et al., 1970). Colchicineinduced autoploids (2n=180?) of the sweet potato became definitely inferior in productivity of edible roots and other traits (Pi and Wang, 1961). These facts strongly indicate that the genomes of I. trifida and I. littoralis might have differentiated to some extent before or after polyploidization, but chromosome homology and genetic harmony in plant life have still well enough maintained. Ways of origin for the sweetpotato : I. trifida or the sweet potato was probably derived through a mechanism shown in Fig. 3A which is the most popular way for the production of polyploids. Fig. 3B indicates another possibility in which no tetraploid takes part. That is, triploids would be directly derived from diploids and maintained vegetatively or by their perenniality. Triploids in Morus (Osawa, 1916; Seki, 1952), Lycoris (Nishiyama, 1928, 1939) and tea plants (Shimura and Inaba, 1953) might also have been derived through the same mechanism, because no tetraploid was found among their relatives. The occurrence of 2n: 3n twin oats suggests another than the usual way by fertilization between reduced and unreduced gametes (Nishiyama et al., 1968). Out of eight plants from a seed sample (K222) obtained in Mexico six (I. trifida- 3x) were cytologically observed to possess45 somatic chromosomes. Their pollen fertility was 22.5-71.4% but the seed set was very poor (0.14%) with artificial pollination between cross-compatiblegroups. In the following generation they gave five plants 382 I. NISHIYAMA Vol. 84

Fig. 3. Two possible origins, A and B, for the sweet potato. Double lines indicate doubling of chromosomes. with 90 somatic chromosomes. In other crossing experiments it was found that viable gametes of I. trifida-3x usually had nearly 45 or 30 chromoomes (Shiotani et al., 1970). The original triploid seed might have been produced by natural crosses between diploids and tetraploids. Another possibility is that they were obtained when triploid plants were crossed with pollen of diploids, because no tetraploid was found as yet in the neighborhood but only diploid I. leucantha. It is interesting to discover various possibilities for the new production of hexaploid forms or I. trifida in the natural conditions of tropical Mexico. Geographical distribution of I. trifida and its basic species : Numerous species of the genus Ipomoea are mainly distributed in the tropics from South-East Asia to Central and South America. Since 1955 I. trifida (6x) was collected at three localities in Mexico, Fortin (strain no. K123), Jalapa (K234) and Oaxaca (K177). I. littoralis (K233, 4x) was found in the Veracruz area. I. leucantha (K221, 2x) and I. trifida- 3x (K222) were collected around Acapulco. It can be briefly said that I. trifida and its relatives inhabited the belt zone reaching from the Pacific Ocean to the Gulf of Mexico at about 17°-20°N latitude. This belt seems to be one of the most probable birthplaces of I. trifida and sweet potato although detailed field surveys of the natural distribution of Ipomoea species in tropical America from Mexico to Peru are needed. Mating system of Ipomoea : The pattern of mating seems to be an important tool for understanding the relationship between sweet potato and wild Ipomoea species. Many workers have already reported interesting investigations into the reproductive Evolutionof the SweetPotato 383 system of a number of Ipomoea species (Nishiyama et al., 1961b; Nishiyama and Teramura, 1962; Sakamoto and Miyazaki, 1970). The results are briefly summarized. Of four diploid species, three, I. triloba, 1. lacunosa and I. trichocarpa were self-fertile and readily crossed with each other. The remaining one, I. leucantha, was observed to be self-sterile and was incompatible with self-fertile diploids, although a small percentage of seeds was found in some cross combinations when it was used as the pollen parent. Wedderburn (1967), however, found that I. trichocarpa (2x) and I. gracilis (2x) were self-sterile. Tetraploid and diploid forms of five species, I. tiliacea, I. gracilis, I. ramoni, I. bilobaand I. arborescens,are mentioned in the literature (King and Bamford, 1937; Ting et al., 1957; Nishiyama et al., 1961a; Jones, 1964; Wedderburn, 1967; Vijaya et al., 1969). As to the first three, however, it is necessary to confirm whether the specimens used for chromosome counts were properly classified. Since that time, a new tetraploid, I. littoralis, has been found and is included in the present study. All three tetraploids, namely I. gracilis (K134), I. tiliacea (K270) and I. littoralis (K233) were self-sterile. The last one set no seed in crosses with the former two but occasionally produced a few seeds (0.6-2.7%) when it was the male parent. The former two species, however, exhibited a normal seed set (about 50%) in reciprocal crosses. Seed fertility (1.7%) of I. gracilis was generally found to be extremely low, but nearly normal in certain intracross-pollinations (T. Teramura's personal communication). Thus the poor fertility appears to depend mostly on genotypic control rather than on the recent autotetraploid origin as discussed by Jones (1970). Three strains of artificial tetraploids have been obtained as follows; (1) Autotetraploid of I. leucantha (K221, 2x), treated with colchicine. (2) I. trifida-(3x)-6x(K222)x l. leucantha (K221, 2x) hybrid. (3) Tetraploid hybrid in (2) x l. littoralis (K233, 4x) hybrid. As mentioned previously, the meiotic division in those tetraploids was nearly normal and 30 bivalents were usually observed. All of them were self-sterile but gave a good seed set in intracross-pollinations and even in crosses between different strains. Only autotetraploids of I. leucantha showed a poor seed set (4.1%) in intracross- pollination. Four self-sterile hexaploid species or strains were found, i.e., two natural I. batatas and I. trifida (K123), an induced I. trifida-(3x)-6x(K222)and a synthesized I. littocantha. They were cross-compatible with each other. In addition, a self- fertile hexaploid I. lacunociliswas artificially derived from self-fertile I. lacunosa (K61, 2x) and self-sterileI. gracilis (K134, 4x). It is especially interesting that I. lacunocilis was highly incompatible with self-sterile hexaploids. Jones and Kobayashi (1969) reported that hexaploids with genome formula AAAAAAderived from doubling the chromosomes of I. ramoni (2x) and I. lacunosa (2x) were almost completely sterile. Thus self-sterility was found to be common to the sweet potato, I. leucantha, I. littoralis, I. trifida and their derivatives. As could be expected, a number of cross- 384 I. NISHIYAMA Vol. 84

Table 2. Seed set(%) in crossing of natural and in duced hexaploids of Ipomoeal ~.

incompatible groups was found in each of them. Briefly it may be concluded that from the stand point of their mating system, the three wild species just mentioned are more closely related to the sweet potato than to the other species. Domestication of the sweet potato : Among all diploid and tetraploid I pomoea species a cultivated type related to sweet potato was never found (Yen, 1961, 1968). No cultivated character occurs in the synthesized I. littocantha and I. trifida-(3x)-6x. These facts suggest that the domestication of the sweet potato could have taken place mainly by successive mutations occurring in I. trifida. The inheritance of autohexa- ploids is very complicated due to random assortment offthe six homologous chromosomes or their chromatids accompanied by crossing over between a kinetochore and a gene locus. In the sweet potato, however, certain chromosomes do not seem to allow completely free assortment because they are to some extent differentiated. The sweet potato or I. trifida may still be in favor of accumulating genes, six doses being expected at the maximum instead of two doses in the corresponding diploid. High doses of desirable genes in a cultivated type would mean a great advancement in the process of domestication. Artificial selection tried by ancient people in the tropics where sweet potatoes and I. trifida bloom and set seeds might have been effective also in the quest for better clones, especially for a higher yield of edible roots. It is also noticeable that the phenotypic expression may be changed owing to different dose rates, the frequency of six-dose recessives or dominants being extremely low in autho- hexaploids. Therefore natural sweet-potato varieties or even modern improved ones may still be heterozygous for domesticated characteristics. In fact it was reported by many workers that a few to about 30% seedlings obtained from crossing certain sweet- potato varieties failed to produce edible roots (Tioutine, 1935; Miyake and Matsunaga, Evolution of the Sweet Potato 385

1939; Jones, 1967). Certain sweet-potato clones or a number of seedlings were also found to exhibit such primitive characteristics as twining stems and pubescence on several plant organs. Should selection be directed toward noneconomic characters, more primitive traits or wild type characteristics close, in the extreme case, to I. trifida may be found. For the same reason the sweet potato could be improved if selection were carried out toward economic characters. Cytotaxonomy of I. batatas and related species : Until recently the classification of I pomoea was almost entirely made from a morphological point of view. However, I pomoea exhibits considerable variability in most of the morphological characters which often do not contribute to easy species identification. With the discovery that the sweet potato arose through repetition of the B genome of I. leucantha a revision became necessary. Table 3 shows a new classification of I. batatas and related species. Though diploid I, leucantha may be the basic species of sweet potato, I. batatas is used for the whole sweet-potato group, because this was the first name applied to any

Table 3. Classification of I pomoea bat atas and its relatives.

member of the group. Little is known about the cytogenetical relationship between I. batatas including its ancestors and other species of the section batatas of the "grupo concavomucronati -sepalas" (Matuda, 1963) in which six species, I. trichocarpa , I. triloba, I. gracilis, I, tiliacea, I, trififa and I. batatas, are placed. Jones and Deonier (1965) already ascertained that I. lacunosa, I. rarnoni, I. trichocarpa and I. triloba have the same genome A. Any cytological difference between genomes A and B is quite unknown. Their differentiation, however, is suggested by the facts that AA and BB species were self-fertile and self-sterile, respectively, and a high cross- incompatibility or a sexual isolation was found between them.

The author wishes to thank Dr. M. Hiroe, Botany Department, Kyoto University, for his valuable suggestions as to cytotaxonomy.

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