The JapaneseSocietyJapanese Society forforPlant Plant Systematics ISSN OOOI-6799 Acta Phytotax, Geobot. 45 (2):95-109 (1994) Distribution of infraspecific taxa of Petunia axillaris (Solanaceae) in Uruguay as revealed analyses by discriminant TOSHIO ANDOi, SHINICHI IIDAi, HISASHI KOKUBUNi, YOSHIHIRO UEDAi and EDUARDO MARCHESI2 iFkeculty 2Flicultad of Hbrticalture, Chiba U}iiversity, 648 Matsudo, Chiba 271; de Agronomia, Uhiversidad de la Republica, Garzon 780, Montevideo, U)uguay Abstract. A survey of native habitats of Petunia species over 5 seasons revealed the co- occurrence of two subspecies of P. axittaris sensu tato in Uruguay. Typical subsp. axiUaris and subsp. parodii were found in south eastern and north western departments, respectively, Using floral measurements (length of corolla tube, corolla limb, pedicel, and calyx lobes, and the length of corolla tube1length of corella limb ratio) of specimens collected in these de- partments, we performed stepwise discriminant analyses. After substituting measurements of 174 specimens collected in the whole of Uruguay into the three discriminant functions thus obtained, we determined them to bc either subsp, c[xittaris or subsp. parodii. Distribution of the two subspecies thus determined by the discriminant functions was roughly delimited by the Rio Negro. A considerable number of doubtfu1 specimens were found in the south west- ern dcpartmcnts, which are on opposite sides of the river de La Plata from the place where secondary intergradation betweell thc two subspecies had been suggested by Wijsman (1982) to have occuiTed, Key words: discriminant function, disnibution, gardcn petunia, geographical taxa, Petunia urillaris, Solanaceae. Received Plebruary 14, i994; accqpted September 30, l994 In 1834, the breeding of garden petunias was started by crossing Petu- nia axitlaris (Lamarck) Britton, Sterns & Poggenburg and P. integrijlotia (Hooker) Schinz & Thellung (Sink, 1984), which had been collected at Montevideo, Uruguay (Lamarck, 1793; Wijsman, 1982), and Rio Negro, Uruguay (Hooker, 1831), respectively. Although these species also grow in Argentina and Brazil (Fries, 1911), the parents of the first Petunia hybrid originated in Uruguay. To know the genetic background of garden petu- nias, detailed information about the species of Petunia native to Uruguay would be usefu1. tours seasons to observe the habitat of Petunia During for five species in Uruguay, we found the co-occurrence of two subspecies of P. axiUaris, that is, subsp. axillaris (short corolla tube with large corolla limb) and subsp. parodii (Steere) Cabrera (long corolla tube with small corolla limb). Subspecies parodii was initially found in Formosa Province in Argentina, NII-Electronic Library Service The JapaneseSocietyJapanese Society forforPlant Plant Systematics . 96 Acta Phytotax. Geobot. Vol, 45 and had not been reported in Uruguay (Wijsman, 1982). In order to develop appropriate methods to distinguish subsp. axiUaris from subsp. parodii and to know the precise distribution of these subspe- cies of P. axillaris in Uruguay, we conducted discriminant analyses using floral characters measurable in dried specimens. Materials and Methods Specimens studied One hundred and thirty-four specimens of Petunia axiUaris sensu lato were collected during five visits to Uruguay (Nov. 15 to Nov. 29, 1988; Nov. 25 to Dec. 4, 1989; Nov. 8 to Nov. 25, 1990; Oct. 31 to Nov. 10, 1991; Dec. 9 to Dec. 19, 1992). Some of the specimens were donated to the MVFA (Laborat6rio de Botanica, Facultad de Agronomia, Monte- video, Uruguay) as indicated in Appendix 2. The distance that we traveled in Uruguay exceeded 20,OOO km. In addition, forty specimens of P. axilla- ris sensu tato collected otherwise and kept in the MVFA were also used for this study. Characters for discriminant analysis Even though the condition of stamens, that is, didynamous (subsp. ax- illaris) or not (subsp. parodii), had been regarded as an important charac- ter to distinguish infraspecific taxa of this species (Steere, 1931), it was dif- ficult to correctly measure this feature in the fragile inner corolla organs without damaging the specimens. Therefore, characters related to stamen condition were not used in this study. For the study of herbarium specimens, Wijsman (1982) had recom- mended using the length of the corolla tube (from the lower end of the calyx to the point separating the tube and limb) to distinguish subspecies of P. axiUaris sensu tato from each other. In addition to this parameter, the length of the corolla limb (from the point separating the tube and limb to the tip of the limb), pedicel, and calyx lobes (from the base of the calyx lobe to the tip of the longest calyx lobe) were also measured in this study. The corolla tube length 1 corolla limb length ratio was also used since a similar ratio (corolla tube length 1 corol]a limb diameter) had been re- garded as a good feature fbr distinguishing live infraspeeific taxa of P. axil- laris sensu tato (Wijsman, 1983). The mean value of all floral measure- ments per specimen was used for subsequent analysis. Discriminant analysis Stepwise discriminant anaiysis was performed by use of the SPSSX software program provided by the Computer Center of Chiba University. The criterion used for determining the selection of characters was the mini- mized Wilks' lambda. The lower limit for selecting or removing a character was based on a partial F-ratio, which was set at F l 1.000 to enter and F < 1.000 to remove. The minimum tolerance level was set at O.OOI. The re- maining conditions for the stepwise discriminant analysis were set to the default values as described in the User's guide of SPSSX (SPSS Inc., NII-Electronic Library Service The JapaneseSocietyJapanese Society forforPlant Plant Systematics December 1994 ANDO et aL : Djstribution of Petunia exitlaris 97 1986). Using floral measurements of specimens collected in the south eastern (Canelones, Maldonado, and Montevideo; 33 specimens of typical subsp. axillaris) and north western departments (Artigas and Salto; 27 specimens of typical subsp. parodii), we performed stepwise discriminant analysis. By substituting measurements of the remaining specimens into discri- minant functions thus calculated, we determined all specimens to be either subsp. axillaris or subsp. parodii. Results and Discussion F7equency distribution of corolla tube length Frequency distribution of corolla tube length measured in all 174 specimens of P. axillaris sensu lato is shown in Fig. 1. We could not con- firm a clear separation of the frequency distribution into two groups (groups of subsp. axillaris and subsp. parodii) as had been shown by Wijs- man (1982). According to his figure, which is sirnilar to Fig. 1, he con- cluded that dried specimens with corolla tubes shorter than 46 mm and those exceeding 46 mm could be identified as subsp. axillaris and subsp. parodii, respectively. 71ypicat subsp. axillaris and subsp. parodii in Uruguay Observations of flowers of live plants in their native habitat revealed that all Petunia axiltaris sensu lato growing in the south eastern depart- ments (Canelones, Maldonado, and Montevideo) were typical subsp. axit- inris, as they had fbur strongly didynamous stamens (2 long, 2 medium, and 1 short), a short corolla tube, and a larger corolla limb. As mentioned earlier, Montevideo is the type locality of this subspecies. 30 :oE'agop6nosEsz 20 10 30 ao 50 6a 70 Length of corolla tube (mm) Ft{/}. 1. Frequency of corolla tubc length in the specimens of Uruguayan Petunia axillan's used in this study. NII-Electronic Library Service The JapaneseSocietyJapanese Society forforPlant Plant Systematics 98 Acta Phytotax. Geobot, Vol, 45 On the other hand, all Petunia axillaris sensu lato growing in the north western departments (Artigas and Salto) were considered to be typical subsp. parodii because of 4 longer stamens ef equal length with 1 short sta- men in addition to a long corolla tube and smaller corolla limb as de- scribed by Steere (1931). This area is close to the type locality of this sub- species, that is, Formosa Province, Argentina. Even though the degrees of the didynamous condition were not re- corded in our field survey, stamens of P. axillaris sensu lato observed by us in the departments south of and north of the Rio Negro were strongly and weakly didynamous, respectively. Variations in stamen condition of live plants grown from seeds will be reported elsewhere. Except for stamen condition, differences in the floral characters men- tioned above were also confirmed in dried specimens (Table 1). According to Wijsman's (1982) criterion as mentioned earlier (critical length of corol- la tube = 46 mm), it is also reasonable to say that the south eastern and north western taxa of P. axillaris sensu lato are subsp. axillaris and subsp. parodii, respectively. As shown in Table 1, differences betwecn five characters in specimens from south eastern departments and those from north western departments were significant. In addition to the corolla tube length, therefore, all other characters measured in this study could be used as additional characters to discriminate these taxa from each other, even though Wijsman (1982) had stated that no character distinguishing subsp. axillaris from subsp. parodii had been found except for corolla tube length. Discriminant analysis At first, the discriminant analysis was performed using all five charac- ters (length of corolla tube, corolla limb, calyx lobes and pedicel, and ratio of corolla tube length to corolla limb length). Pedicel length and corolla tube length ! corolla limb length ratio, however, were removed from the optimum subset for discriminating subspecies since their F values were less than 1.000. The discriminant function thus obtained was as follows: - Zi = O.2168763Xi + 0.4312565X2- O.2530165X3 9.321480 ･･･････････････function 1 TABLE 1. Comparison of floral characters rneasurable in dried specimens of Petunia axiUaris subsp. Maldonado & Montevideo) and subsp. axitlatty from south eastern departments(Caneloncs, paro- north westem alld Salto), diifrom departments(Anigas Length of Ratio Corolla Corolla Calyxlobe Pedicel(mm) Tube1 tube (mm) limb (mm) (mm) limb Subsp.