TROPICS Vol. 14 (1) Issued August 31, 2004

Morphological characters and numerical taxonomic study of CapsicumCapsicum frutescensfrutescens in Southeast and East Asia.

Sota YAMAMOTO and Eiji NAWATA Laboratory of Tropical Agriculture, Graduate School of Agriculture, Kyoto University, Kyoto 606-8502, Japan [email protected]

ABSTRACT Morphological characters were studied in 90 accessions of Capsicum frutescens in Southeast and East Asia, and cluster analysis was carried out to qualitatively and quantitatively assess the characters. Four major clusters appeared both in the qualitative characters (XA, XB, XC and XD) and quantitative characters (YA, YA, YC and YD), and 90 accessions were classified into 9 types based on cluster combinations. Four major types (XAYB, XAYC, XBYB and XBYD) including 61 accessions were distributed evenly in Southeast Asia. Five minor types, on the other hand, were distributed unevenly: XAYA in the Ryukyu Islands, Indonesia, northern Thailand; XBYA in Indonesia, southern Thailand, Vietnam; XCYD in northern and southern Thailand; XCYD in Vietnam and Laos; XDYD in southern Thailand. Morphological characters were more diversified in Thailand. This is partly because C. frutescens is used more frequently in Thailand than in other regions. All 17 accessions from the Ryukyu Islands, on the contrary, showed exactly the same qualitative characters and high similarity in quantitative characters. These results agree with the results of biochemical analyses. Accessions from the Ryukyu Islands were both morphologically and biochemically very close to each other, indicating that the Ryukyu Islands is the end of the dispersal route of C. frutescens in Southeast and East Asia.

Key words: cluster analysis, geographical distribution, qualitative characters, quantitative characters, the Ryukyu Islands

INTRODUCTION

Capsicum frutescens is considered to have originated from Amazonia (Eshbaugh, 1970) and is now widely spread in tropical and subtropical regions. Tabasco, used for Tabasco sauce, is the only economical cultivar of C. frutescens. In Southeast and East Asia, C. frutescens is cultivated mainly in home gardens or small fields, and weed forms of C. frutescens are also seen on forest edges, in orchards and along roadsides. This is no doubt the result of dispersal by birds who are attracted to the small and vivid red fruits of C. frutescens (Wheelwright and Janson, 1985). C. frutescens is usually very spicy with a distinctive flavor that adds a much deeper taste to local foods in the tropics. Morphological characters of Capsicum peppers are agronomically very important to breeders and farmers. For example, bigger fruits may be more valued as vegetables or condiments, while small and cute fruits may be useful as ornaments. The consumer may prefer a vivid fruit color. Like other crops, plant height and plant type are also important morphological characters when peppers are cultivated. The morphology of C. frutescens has been studied to some extent as one species of the genus Capsicum. Morphological characters of C. frutescens, according to Andrews (1995), are: Corolla a greenish-white color without diffuse spots at the base of lobes; peduncles erect at anthesis but flowers nodding; anthers violet to blue; calyx of mature fruit without annular constriction at the junction with the peduncle, though often irregularly wrinkled; veins usually not prolonged into teeth; seeds straw-colored. These characters, however, describe mainly about C. frutescens grown in Central and South America, and those of C. frutescens in Southeast and East Asia have not been reported on yet. Earlier, Eshbaugh (1964) carried out a numerical taxonomic analysis of the genus Capsicum, but Pickersgill et al. (1979) pointed out that he produced less than satisfactory results, partly because his use of a large number of size characters might have mislead the numerical taxonomic analysis at the genus level. Capsicum peppers show much parallel variation within the genus, such as gigantism of fruits or other organs, and some domesticated forms clustering with domesticated forms of other species, rather than their presumed wild ancestor. However, at the species level, especially at accessions obtained from limited regional levels, quantitative and qualitative characters are considered to be useful tools to study morphological relationships among accessions. In this paper, to estimate the dispersal routes of C. frutescens into Japan, [1] qualitative and quantitative morphological characters of C. frutescens in Southeast and East Asia were studied, and [2] numerical taxonomic analysis was carried out to investigate morphological relationships among the accessions obtained. 112 Sota YAMAMOTO and Eiji NAWATA

MATERIALS AND METHODS

Morphological characters of 90 accessions were investigated, and they were used for numerical taxonomic analysis. All accessions were obtained from markets, fields, roadsides, and university collections. Accessions obtained from markets were considered to be cultivated in nearby areas, because C. frutescens is usually not A Southeast ● and East Asia ● distributed on a large scale. Collection sites ● ● ● 1,000 km are shown in Fig. 1 and accession details in ●● ● Laos ● ● ● ● Table 1. Seeds of each accession were ● ● ● A ● ● ● B germinated in vermiculite, and about one ● ● ● month later seedlings were transplanted Vietnam into clay pots (18 cm height and 23 cm in ● diameter, 3 seedlings of each accession per ● pot) filled with a mixture of perlite, sand ● ● and organic manure at a ratio of 1:1:1. They C were grown in a greenhouse at Kyoto B University, Japan. Qualitative characters Thailand were investigated with the naked eye, and ● Ryukyu Is. ● quantitative characters were measured ● ●● (number of replications, depending on ● characters, was 3-10). One-year plants ●● ● ● ● 200 km were used for the measurement of plant 500 km ●● height. Normal flowers were chosen to ● ● measure quantitative characters of flower ● organs, because Capsicum peppers Java I. ● Lombok I. sometimes have abnormal flowers that are ● C 500 km Timor I. easily distinguished (regarding abnormal Fig. 1. Collection sites of C. frutescens in Southeast and East Asia. pepper flowers, see Cochran (1934)).

Table 1. Accession number and collection site of 90 accessions of C. frutescens . Accession No. Region Location Accession No. Region Location Accession No. Region Location TACF-1 JPN1 Oshikawa, Kunigami, Okinawa Island TACF-60 THA3 Mae Hong Son, Mae Hong Son TACF-94 THA3 Song, Phrae TACF-3 JPN1 Chatan, Nakagami, Okinawa Island TACF-61 THA3 Mae Hong Son, Mae Hong Son TACF-95 THA3 Mae sot, Tak TACF-4 JPN1 Miyako Island TACF-62 THA3 Mae Hong Son, Mae Hong Son TACF-96 THA3 Mae sot, Tak TACF-5 JPN1 Takano, Miyako Island TACF-63 THA3 Mae Sariang, Mae Hong Son TACF-97 THA3 Mea Ramat, Tak TACF-6 JPN1 Hukuri, Miyako Island TACF-64 THA3 Mae Sariang, Mae Hong Son TACF-106 THA3 Chainat, Chainat TACF-7 JPN1 Sunagawa, Miyako Island TACF-65 THA3 Mae Sariang, Mae Hong Son TACF-107 THA3 Kanchanaburi TACF-8 JPN1 Sawada, Irabu Island TACF-66 THA3 Mae Sariang, Mae Hong Son TACF-108 THA3 Nakon Pathom TACF-9 JPN1 Niikawa, Ishigaki Island TACF-67 THA3 Mae Sam Laep, Mae Hong Son TACF-109 THA3 Bangkok TACF-10 JPN1 Kabira, Ishigaki Island TACF-68 THA3 Mae Sam Laep, Mae Hong Son TACF-116 THA3 Ko Samui, Surat Thani TACF-11 JPN1 Kabira, Ishigaki Island TACF-69 THA3 Mae Sam Laep, Mae Hong Son TACF-117 THA3 Hua Sai, Nakhon Si Thammarat TACF-12 JPN1 Nagura, Ishigaki Island TACF-70 THA3 Pang Ma Pha, Mae Hong Son TACF-118 THA3 Hua Sai, Nakhon Si Thammarat TACF-13 JPN1 Nozoko, Ishigaki Island TACF-71 THA3 Pang Ma Pha, Mae Hong Son TACF-119 THA3 Hua Sai, Nakhon Si Thammarat TACF-14 JPN1 Otomi, Iriomote Island TACF-72 THA3 Rak Phaen Din, Toen, Chiang Rai TACF-120 THA3 Pattani, Pattani TACF-16 JPN1 Toyohara, Iriomote Island TACF-73 THA3 Paug Ung, Mae Chaem, Chiang Mai TACF-124 THA3 Ban Nang Sata, Yala TACF-17 JPN1 Hateruma Island TACF-75 THA3 Pai, Chiang Mai TACF-125 THA3 Ban Nang Sata, Yala TACF-18 JPN1 Tamashiro, Hateruma Island TACF-76 THA3 Pong Maek, Chiang Mai TACF-126 THA3 Narathi Wat, Narathi Wat TACF-19 JPN1 N.D.6 TACF-79 THA3 Nan, Nan TACF-127 THA3 Narathi Wat, Narathi Wat TACF-28 INA2 Timor Island TACF-80 THA3 Nan, Nan TACF-128 THA3 Narathi Wat, Narathi Wat TACF-29 INA2 Kupang, Timor Island TACF-81 THA3 Nan, Nan TACF-129 THA3 Thung kha, Narathi Wat TACF-30 INA2 Kupang, Timor Island TACF-83 THA3 Nan, Nan TACF-131 THA3 N.D.6 TACF-31 INA2 Kupang, Timor Island TACF-84 THA3 Nan, Nan TACF-133 VIE4 Bac Kan TACF-32 INA2 Lombok Island TACF-85 THA3 Nan, Nan TACF-134 VIE4 Bac Kan TACF-33 INA2 Lombok Island TACF-86 THA3 Nan, Nan TACF-135 VIE4 Cao Bang TACF-34 INA2 Jawa Island TACF-87 THA3 Mae Ung, Huai Lee, Nan TACF-136 VIE4 Cao Bang TACF-35 INA2 Jawa Island TACF-88 THA3 Phasing, Nan TACF-137 VIE4 Son la TACF-37 INA2 Jawa Island TACF-89 THA3 Nan, Nan TACF-138 VIE4 Phieng Luong, Mac Chau, Son la TACF-38 INA2 Jawa Island TACF-90 THA3 Pang Poiy, Nan TACF-139 VIE4 Phieng Luong, Mac Chau, Son la TACF-39 INA2 Jawa Island TACF-91 THA3 Nan Khong, Nan TACF-140 VIE4 Na Moung, Mac Chau, Son la TACF-40 INA2 Jawa Island TACF-92 THA3 Phrae, Phrae TACF-143 LAO5 Nan Ngum TACF-41 INA2 Kuningum, Jawa Island TACF-93 THA3 Song, Phrae TACF-144 LAO5 Vientiane JPN1 = Japan, INA2 = the Indonesia, THA3 = Thailand, VIE4 = Vietnam, LAO5 = Laos, N.D.6 = No detailed data. Morphological characters and numerical taxonomic study of Capsicum frutescens in Southeast and East Asia 113

Euclidean distances were calculated to estimate the similarity among accessions and a dendrogram was constructed based on Euclidean distances using the unweighted pair-group method with arithmetic averages (UPGMA). Every qualitative character was considered a yes (scored as 1) or no (scored as 0), and the measured values of quantitative characters were directly used. A cluster analysis was carried out with Aoki’s Excel VBA Macro (http://www006.upp.so- net.ne.jp/dokugaku-club/).

RESULTS AND DISCUSSION

Morphological characters of C. frutescens in Southeast and East Asia Qualitative characters ‘Corolla color’: all accessions were greenish-white. ‘Dark greenish-white spots at the corolla base’: some accessions showed dark greenish-white spots at the corolla base, but not others (Fig. 2-1). These results contradict the taxonomy of Andrews (1995), but 36 among the 90 accessions showed dark greenish-white spots, suggesting that this character is not rare and should be added to the list of morphological characters of C. frutescens. ‘Style color’: green or purple. ‘Anther color’: blue, yellowish-blue, or yellow (Fig. 2-2). It has few yellow anthers, but Yamamoto (1978) also described the presence of yellow anthers in South America. ‘Filament color’: green, light purple, or purple. ‘Peduncle color’: green, light purple, or purple. ‘Immature fruit color’: Dark green, green, greenish-white, or yellowish-white (Fig. 2-3). ‘Mature fruit color’: red or dark orange (Fig. 2-4). ‘Seed color’: seeds of all accessions were yellow to cream. They showed very subtle differences, and were considered to have the same color. ‘Hypocotyl color’: green or purple. ‘Stem color’: all accessions were green.

1(A) 1(B) 3 (A) (B) (C) (D) 5cm

2(A) 2(B) 2(C)

4 (A) (B) 5 (A) (B)

6 (A) (B) (C) 7 Fig. 2. Morphological differences. 1: Dark greenish-white spots at the corolla 5cm base (1a: present, 1b: absent). 2: Anther color (2a: blue, 2b: yellowish-blue, 2c: yellow). 3: Immature fruit color (dark green, green, greenish-white, yellowish-white, from left to right). 4: Mature fruit color (red, dark orange, from left to right). 5: Mature fruit position (5a: erect, 5b: intermediate (horizontal)). 6: Fruit shape at apex (pointed, blunt, sunken, from left to right). 7: Leaf shape (leaf length/leaf width) (1.54 (TACF-119), 2.00 (TACF-1), 2.98 (C. (A) (B) (C) annuum obtained from the Ryukyu Islands, unpublished data), from left to right). 114 Sota YAMAMOTO and Eiji NAWATA

‘Deciduousness of fruits’: present or absent. To distinguish C. frutescens and C. chinense (closely related to C. frutescence), Eshbaugh (1975) adopted a deciduous (C. frutescens) or persistent (C. chinense) characteristic. Some accessions of C. frutescens, however, showed a persistent characteristic, and it is supposed that this criterion is not suitable for distinguishing these two species. ‘Peduncle position at anthesis’: peduncles of all accessions were erect at anthesis. ‘Mature fruit position’: some accessions showed an intermediate (horizontal) position when fruits were maturing, though their peduncles were erect at anthesis (Fig. 2-5). It seems that bigger fruits tend to be horizontal because of their weight, but fruits of some accessions did not show an intermediate position, although they were almost the same size as those with a horizontal trait. It is suggested that there are several factors which determine the fruit position. ‘Fruit shape at apex’: pointed, blunt, or sunken fruits were found (Fig. 2-6). All 17 accessions from the Ryukyu Islands had sunken fruits, although of the accessions from the other regions only two from Indonesian bore sunken fruits. Five accessions with big fruits had blunt fruits, and the others pointed fruits. ‘Calyx margin shape and annular constriction at junction with peduncle’: no typical calyx teeth and annular constriction were found in accessions obtained in Southeast and East Asia. ‘Leaf pubescence’: all accessions showed glabrous to very lightly pubescent.

Table 2. Geographic distribution of 11 qualitative characters which are considered to be useful characters to categorize C. frutescens in Southeast and East Asia. Region Morphological characters Ryukyu Islands Indonesia Thailand Viet Nam Laos Dark greenish-white spots present 0 5 26 3 2 at the corolla base absent 17 8 24 5 0 Style color green 17 13 50 7 0 purple 0 0 0 1 2 Anther color blue 17 8 38 3 2 yellowish blue 0 5 9 5 0 yellow 0 0 3 0 0 Filament color green 17 1 4 0 0 light purple 0 3 22 2 0 dark purple 0 9 24 6 2 Peduncle color green 0 7 24 5 2 light purple 17 6 11 2 0 dark purple 0 0 15 1 0 Immature fruit color dark green 0 1 0 0 0 green 0 8 39 7 0 greenish-white 17 4 10 1 2 yellowish-white 0 0 1 0 0 Mature fruit color red 17 13 49 8 2 dark orange 0 0 1 0 0 Hypocotyl color green 0 7 17 5 2 purple 17 6 33 3 0 Deciduous present 0 3 20 5 0 absent 17 10 30 3 2 Mature fruit position erect 17 10 45 7 0 intermediate (horizontal) 0 3 5 1 2 Fruit shape at apex pointed 0 9 48 8 1 blunt 0 2 2 0 1 sunken 17 2 0 0 0 Morphological characters and numerical taxonomic study of Capsicum frutescens in Southeast and East Asia 115

Eleven qualitative characters are considered as useful characters to distinguish C. frutescens in Southeast and East Asia. The geographical distribution of these characters is shown in Table 2. Characters such as ‘Dark greenish-white spots at the corolla base’, ‘Filament color’, ‘Peduncle color’, ‘Hypocotyl color’, ‘Deciduousness’ and ‘Mature fruit position’ were distributed almost evenly in Southeast and East Asia. The following characters, however, were unevenly distributed: ‘Purple style color’ was distributed in Vietnam and Laos, ‘Yellow anther color’ found only in Thailand, ‘Dark green and yellowish- white immature fruit color’ in Indonesia and Thailand respectively, ‘Dark orange mature fruit color’ only in Thailand, and ‘Sunken fruit shape at apex’ found only in the Ryukyu Islands and Indonesia.

Quantitative characters ‘Plant height and number of branches’: plant height varied from ca. 30 cm to 140 cm. Accessions from Vietnam tended to be short, whereas accessions from Indonesia to be tall. The number of branches was 1 to 7, and smaller accessions were likely to have more branches. There was a negative correlation between plant height and number of branches (p<0.01) (Table. 3). It is indicated that peppers may have two techniques to increase their assimilation products: either they have a higher statue and fewer branches, or, with more branches and a lower statue.

Table 3. Correlation coefficients among 14 quantitative characters. corolla diameter petal corolla diameter ― length peduncle petal length 0.9212** ― length pistil peduncle length 0.5070** 0.4634** ― length stamen pistil length 0.3533** 0.4199** 0.2300* ― length fruit stamen length 0.7570** 0.7765** 0.5827** 0.5701** ― length fruit fruit length 0.6501** 0.6046** 0.6574** 0.1596 0.6515** ― width fruit fruit width 0.4707** 0.4300** 0.4757** 0.3628** 0.7239** 0.6138** ― size fruit fruit size 0.6510** 0.5974** 0.6550** 0.2831** 0.7596** 0.9055** 0.8730** ― shape number seeds fruit shape 0.2601* 0.2597* 0.2852** －0.2105* 0.0340 0.5735** －0.2771** 0.1856 ― per fruit 100 seeds number seeds per fruit 0.3982** 0.4298** 0.5333** 0.3518** 0.6717** 0.5787** 0.8345** 0.7480** －0.1268 ― weight plant 100 seeds weight 0.3592** 0.3230** 0.3788** 0.2000 0.5497** 0.5174** 0.7777** 0.7011** －0.1504 0.5629** ― height number of plant height 0.2005 0.2388* 0.1926 0.0879 0.1741 0.5355** 0.2396* 0.4608** 0.3734** 0.1808 0.1324 ― branches number of branches －0.0831 －0.1287 －0.1145 －0.0770 －0.0774 －0.2073* －0.1584 －0.2149* －0.0845 －0.1210 －0.0888 －0.5053** ― leaf shape －0.1534 －0.1478 0.0001 0.1353 0.1041 0.0725 0.1322 0.0864 －0.0110 0.1611 0.1879 －0.0549 0.1290 * and ** indicate the correlation coefficients at p < 0.05 and p < 0.01 respectively.

‘Corolla diameter and petal length’: corolla diameter was ca. 12.5 mm to 20 mm and petal length was ca. 8.5 mm to 13 mm. There was a clear visible difference between small and big flowers. ‘Pistil length and stamen length’: pistil length was ca. 5 mm to 6.5 mm and stamen length was ca. 3.6 mm to 5 mm. Generally, the pistil was longer than the stamen, and it exserted well enough. This character is considered to play some role in cross-pollination. Capsicum peppers are usually self- pollinating, but it is known that they are also occasionally cross-pollinating (Odland and Porter, 1941), and Tanksley (1984) found in his study that the natural cross-pollination occurrence was 42%. In fact, many bees, butterflies, and other insects were observed visiting flowers of C. frutescens accessions in this study, and also flowers had visible nectar. If the pistil is exserted well enough, cross-pollination will occur easily when bees or other insects visit the flowers. More studies are needed on cross-pollination of this crop. ‘Fruit length, fruit width, fruit size (length×width) and fruit shape (length/width)’: fruit length was ca. 15 mm to over 55 mm, fruit width ca. 4 mm to 12 mm, fruit size ca. 60 mm2 to over 450 mm2 and fruit shape ca. 3 to over 7. ‘Seed number per fruit’: seed number per fruit varied from ca. 10 to over 50. ‘One hundred seeds weight’: one hundred seeds weight varied from ca. 250 mg to 550 mg. ‘Peduncle length’: peduncle length varied from ca. 20 mm to 30 mm. ‘Leaf length, leaf width, and leaf shape (length/width)’: leaf length and width are significantly affected by leaf age, leaf position and environment. Leaf shape, however, is considered to be stable in each accession irrespective of the age, position, and environment (excluding extreme conditions). Leaves of C. frutescens were clearly wider than those of C. annuum (unpublished data), as some researchers 116 Sota YAMAMOTO and Eiji NAWATA reported (Kumazawa et al., 1954; Hirose et al., 1957), and the leaf shape ratio of the widest leaf was 1.54 (Fig. 2-7). There are obvious differences in leaf shape among accessions of C. frutescens, and this character is useful for numerical taxonomic studies. Correlation coefficients among 14 quantitative characters are shown in Table 3. There were highly positive correlations among floral organs, fruit characters and seed characters, indicating that the bigger the flowers, the bigger the fruits and hence more seeds. Moreover, higher plants tend to bear bigger fruits. These results are considered to be typical of gigantism in plants.

Numerical taxonomic studies of C. frutescens in Southeast and East Asia Morphological characters used in numerical taxonomic studies are shown in Table 4, and a UPGMA dendrogram was constructed of qualitative and quantitative characters, from the Euclidean distances among 90 accessions.

Table 4. Morphological characters used in numerical taxonomic studies. Qualitative characters Quantitative characters 1 Spots at the corrola base (present; absent) 12 Corolla diameter (mm)※1 2 Style color (green; purple) 13 Petal length (mm)※1 3 Anther color (blue; yellowish blue; yellow) 14 Peduncle length (mm)※1 4 Filament color (green; light purple; dark purple) 15 Pistil length (mm)※1 5 Peduncle color (green; light purple; dark purple) 16 Stamen length (mm)※1 6 Immature fruit color 17 Fruit length (mm)※1 (dark green; green; greenish-white; yellowish-white) 18 Fruit width (mm)※1 7 Mature fruit color (red; dark orange) 19 Fruit size (mm2) (length x width) 8 Hypocotyle color (green; purple) 20 Fruit shape (length/width) 9 Mature fruit position (erect; intermediate(horizontal)) 21 Number seeds per fruit※1 10 Deciduous (present; absent) 22 One hundred seeds weight (mg) 11 Fruit shape at apex (pointed; blunt; sunken) 23 Plant height (cm)※2 24 Number of branches※2 25 Leaf shape (length/width)※1

※1) n=10, ※2) n=3

Qualitative characters Four major clusters appeared: ‘Cluster XA’, ‘Cluster XB’, ‘Cluster XC’, and ‘Cluster XD’ (Fig. 3-1). All accessions belonging to ‘Cluster XA’ had green styles, blue anthers (except TACF-76), a red mature fruit color, purple hypocotyls, and were erect, whereas, all accessions belonging to ‘Cluster XB’ had green hypocotyls (except TACF-41, 116), and also had green styles, green peduncles (except TACF-67), and a red mature fruit color. ‘Cluster XC’ consisted of the three accessions which showed no spots at the corolla base, had purple styles, blue anthers, a greenish-white immature fruit color and red mature color, an intermediate fruit position, and persistent characteristics. ‘Cluster XD’ consisted of only one accession, TACF-117, which had very unique characters, such as a yellowish-white immature fruit color and a dark orange mature fruit color. Only TACF-117 had a yellowish-white immature fruit color, and turned dark orange when it matured, suggesting that these two characters are linked.

Quantitative characters Four major clusters appeared: ‘Cluster YA’, ‘Cluster YB’, ‘Cluster YC’, and ‘Cluster YD’ (Fig. 3-2). ‘Cluster YA’ (average of fruit length 29.39 ± 0.68 mm) consisted mainly of the accessions from the Ryukyu Islands; and sub-1 group consisted of all 17 accessions from the Ryukyu Islands and one from Indonesia. ‘Cluster YB’ consisted of the medium sized peppers (average of fruit length 28.61 ± 0.72 mm), ‘Cluster YC’ of the smaller size peppers (18.78 ± 0.48 mm) and ‘Cluster YD’ of the bigger peppers (42.38 ± 1.45 mm). Morphological characters and numerical taxonomic study of Capsicum frutescens in Southeast and East Asia 117

Accession No. Cluster and regions (No. of accessions)

TACF-117 Cluster XD Southern Thailand (1) TACF-144 Vietnam (1) TACF-143 Cluster XC TACF-137 Laos (2) TACF-119 TACF-72 TACF-65 TACF-89 TACF-33 TACF-32, -34, -62 TACF-40 TACF-66 TACF-41 TACF-126 Indonesia (8) TACF-127, -133 Northern Thailand (11) TACF-124 Cluster XB Southern Thailand (8) TACF-139, -140 TACF-63, -120, -134 Vietnam (5) TACF-39, -138 TACF-69 TACF-118 TACF-35 TACF-129 TACF-116 TACF-67, -71, -93 TACF-79 TACF-29 TACF-90, -125 TACF-86 TACF-83, -84, -85, -87, -92 TACF-109 TACF-88 TACF-107, -131 TACF-61, -108, -136 TACF-91 TACF-68, -96 Ryukyu Islands (17) TACF-106 TACF-38 Indonesia (5) TACF-95 Northern Thailand (23) Cluster XA TACF-76 Central Thailand (4) TACF-70 Southern Thailand (2) TACF-64, -94, -97 Vietnam (2) TACF-80, -128 TACF-81 TACF-37 TACF-73 TACF-30, -31, -135 TACF-28 TACF-60 TACF-75 TACF-1～TACF-19 3.53 2.5 2 1.5 1 0.5 0

Euclidean distance

Fig. 3-1. Dendrogram construction from the Euclidean distances among 90 accessions according to qualitative characters. 118 Sota YAMAMOTO and Eiji NAWATA

Accession No. Cluster and regions (No. of accessions)

TACF-144 TACF-72 TACF-40 TACF-34 TACF-137 Indonesia (6) TACF-138 TACF-143 Northern Thailand (4) TACF-65 TACF-32 Cluster YD Southern Thailand (2) TACF-33 TACF-119 Vietnam (2) TACF-89 TACF-62 Laos (2) TACF-35 TACF-117 TACF-29 TACF-106 TACF-96 TACF-136 TACF-91 TACF-109 TACF-129 TACF-63 TACF-131 TACF-86 Indonesia (3) TACF-87 Northern Thailand (12) TACF-73 TACF-108 Cluster YC Central Thailand (4) TACF-107 TACF-85 Southern Thailand (1) TACF-80 TACF-61 Vietnam (2) TACF-92 TACF-84 TACF-135 TACF-83 TACF-31 TACF-30 TACF-28 TACF-97 TACF-116 TACF-125 TACF-124 TACF-126 TACF-88 TACF-127 TACF-95 TACF-128 TACF-134 TACF-120 TACF-71 Indonesia (1) TACF-81 TACF-140 Northern Thailand (17) TACF-76 Cluster YB TACF-70 Southern Thailand (7) TACF-94 TACF-79 Vietnam (3) TACF-68 TACF-69 TACF-67 TACF-93 TACF-66 TACF-133 TACF-90 TACF-64 TACF-75 TACF-37 TACF-118 TACF-41 TACF-60 TACF-39 TACF-139 TACF -7 TACF -6 TACF -3 TACF-12 Ryukyu Islands (17) TACF-38 TACF -9 Indonesia (3) TACF-18 Cluster YA TACF-19 Northern Thailand (1) TACF-17 TACF -8 Southern Thailand (1) TACF-14 Sub-1 TACF-16 Vietnam (1) TACF-11 TACF-10 TACF -5 TACF-13 TACF -4 TACF -1 250 200 150 100 50 0 Euclidean distance

Fig. 3-2. Dendrogram construction from the Euclidean distances among 90 accessions according to quantitative characters. Morphological characters and numerical taxonomic study of Capsicum frutescens in Southeast and East Asia 119

Classification of C. frutescens in Southeast and East Asia based on morphological characters Based on the combination of clusters of qualitative and quantitative characters, 16 types (4×4) were defined. For example, accessions belonging to Cluster XA (qualitative) and Cluster YA (quantitative) were classified as XAYA. All 90 accessions were classified into the following 9 types: major types- XAYB, XAYC, XBYB and XBYD, minor types- XAYA, XBYA, XBYC, XCYD and XDYD (Table 5). Accessions belonging to XBYD, XCYD and XDYD (those showing bigger quantitative characters) tended to have a persistent characteristic and an intermediate fruit position, indicating that they are likely to be the more domesticated forms, whereas, accessions belonging to XAYC and XBYC (those showing small quantitative characters) tended to be erect and deciduous, indicating that they are more likely to be spontaneous forms. All accessions belonging to the other types were medium sized and erect, but half of them had a persistent characteristic. They are supposed to be intermediate forms between wild and cultivated forms.

Table 5. Classification of C. frutescens in Southeast and East Asia based on qualitative and quantitative characters. Quantitative Type Qualitative characterscharacters Distribution areas (and belonging accessions) persistence the Nansei Islands, Indonesia, Northern Thailand XAYA medium green style sunken at fruit apex (except TACF-60) 〔TACF-1, ～ -19, -38, -60〕 blue anther the Nansei Islands, Indonesia, Northern Thailand (except TACF-76) green immature fruit color (except TACF-75) XAYB pointed at fruit apex medium TACF-37, -64, -68, -70, -75, -76, -81, -88, -90, -94, -95,-97, red mature fruit color 〔 -125, -128 〕 purple hypocotyl spots at the corolla base (except TACF-80) Indonesia, Northern and Central Thailand, Viet Nam erect small XAYC green immature fruit color (except TACF-73) TACF-28, -30, -31, -61, -73, -80, -83, -84, -85, -86, -87,-91, 〔 -92, -96, -106, -107, -108, -109, -131, -135, -136 〕 erect Indonesia, Southern Thailand, Viet Nam XBYA medium pointed at fruit apex 〔TACF-39, -41, -118, -139〕

green style green immature fruit color (except TACF-69) Northern and Southern Thailand, Viet Nam XBYB persistence (except TACF-116) medium TACF-66, -67, -69, -71, -79, -93, -116, -120, -124, -126,-127, green peduncle erect 〔 -133, -134, -140 〕 (except TACF-67) no spots at the corolla base red mature fruit color dark purple filament green immature fruit color Northern and Southern Thailand XBYC small green hypocotyl deciduousness 〔TACF-63, -129〕 (except TACF-41, -116) erect pointed at fruit apex Indonesia, Northern and Southern Thailand, Viet Nam XBYD persistence big 〔TACF-29, -32, -33, -34, -35, -40, -62, -65, -72, -89, -119, -138〕

no spots at the corolla base red mature color purple style persistence Viet Nam, Laos XCYD big blue anther intermediate fruit position 〔TACF-137, -143, -144〕 greenish-white immature fruit color

yellowish-white persistence immature fruit color Southern Thailand XDYD big dark orange mature 〔TACF-117〕 fruit color intermediate fruit position

The geographical distribution of major type accessions is shown in Fig. 4-1. Major type accessions were almost evenly distributed in Southeast Asia, though there was no accession belonging to XBYB in Indonesia. Minor type accessions, on the other hand, are unevenly distributed in Southeast and East Asia (Fig. 4-2). Accessions belonging to XAYA types were found only on the Ryukyu Islands, Indonesia and Northern Thailand; XBYA types were in Indonesia, Southern Thailand and Vietnam; XBYC types were in Northern and Southern Thailand; XCYD types were only in Vietnam and Laos, or were distributed only in the inland of the continental region of Southeast Asia; XDYD types were only in Southern Thailand. Classification results indicated that morphological characters are more diversified in Thailand. This is partly because the frequency of use of C. frutescens in Thailand is higher than in other regions. In Northern Thailand, for example, there are 120 Sota YAMAMOTO and Eiji NAWATA ethnic groups who show a strong preference A Southeast for C. frutescens, and a lot of C. frutescens ○ and East Asia ▲ ▽ peppers are seen in local markets there. ▲ 1,000 km

▲● ▽ Laos Moreover, C. chinense, which crosses ○ ● ▽ ○ ● ● ▽○ A ▲ ▽● ▲ easily with C. frutescens, is cultivated more ▲ ○ ●▲ B in Thailand than in other regions, which ● ○ may diversify morphological characters Vietnam more pronouncedly in Thailand. ○

In contrast, all 17 accessions from the ○ ○ ○ Ryukyu Islands showed exactly the same C qualitative characters, including a rare morphological character (sunken fruit B Thailand shape at apex), and were grouped together ▲ (Fig. 3-1). Moreover, ‘Cluster YA’ consisted Ryukyu Is. mainly of accessions from the Ryukyu ▽

Islands, and sub-1 consisted of all 17 ▲ ▲▲ accessions from the Ryukyu Islands and ● 500 km 200 km one from Indonesia (Fig. 3-2). It is clear ● that accessions from the Ryukyu Islands ▽ resemble each other morphologically. Java I. ▽ These results agree with the results of Lombok I. ▽○ biochemical analyses, which showed they C 500 km Timor I. have exactly the same isozymes of 8 Fig. 4-1. Geographical distribution of major type accessions. enzymes (unpublished). Now it has been (●:XAYB, ○:XAYC, ▲:XBYB, ▽:XBYD) revealed that accessions from the Ryukyu Islands are both morphologically and biochemically very close to each other and Southeast they are not diversified, indicating that the A and East Asia Ryukyu Islands are the end of the dispersal ▽ ○ 1,000 km route of C. frutescens in Southeast and East ● ▲ Laos Asia. Accessions from the Ryukyu Islands A ▽ B are morphologically close to one accession ▽ from Indonesia and one from Northern Vietnam Thailand (belonging to XAYA). The dispersal routes of C. frutescens into Japan may be more accurately estimated with these results and the results from biochemical C analyses which will be completed soon. B In this research, C. frutescens in Thailand Southeast and East Asia was studied, but Ryukyu Is. ● C. frutescens in Oceania and Africa has ●● not been studied yet. Further research ○ ★ investigating morphological and ecophysiological ▲ ● 500 km 200 km characteristics and applying biochemical ●● analysis to accessions obtained from much ●○ ○ broader areas is necessary. Java I. Lombok I. C 500 km Timor I. Fig. 4-2. Geographical distribution of minor type accessions. (●:XAYA, ○:XBYA, ▲:XBYC, ▽:XCYD, ★:XDYD) Morphological characters and numerical taxonomic study of Capsicum frutescens in Southeast and East Asia 121

ACKNOWLEDGEMENTS We wish to thank Dr. S. Yazawa, of the Laboratory of Vegetable and Ornamental Horticulture, Faculty of Agriculture, Kyoto University, Japan, for providing seeds of many accessions of C. frutescens from Southeast Asia and the Ryukyu Islands. We are also grateful to the National Research Council of Thailand, the Indonesian Institute of Science and the Vietnam Agricultural Science Institute for providing us with opportunities to investigate C. frutescens.

REFERENCES

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Received 4th Feb. 2004 Accepted 11th June 2004