J. Jpn. Bot. 93(1): 9–17 (2018)

A Phylogenetic Study of Amphicarpaea with a New Genus Afroamphica (Leguminosae Tribe )

a b, Kazuaki Ohashi and Hiroyoshi Ohashi *

aSchool of Pharmacy, Iwate Medical University, Yahaba, Iwate, 028-3694 JAPAN; bHerbarium, Botanical Garden, Tohoku University, Sendai, 980-0862 JAPAN *Corresponding author: [email protected]

(Accepted on September 13, 2017)

The genus Amphicarpaea was revised by Ohashi and Ohashi (2016) based mainly on pollen morphology in which Shuteria africana was resurrected from Amphicarpaea africana and the taxonomic position of A. ferruginea was settled in Amphicarpaea. In the present study, those results were reexamined in molecular phylogenetic analyses using chloroplast DNA and nuclear DNA (ITS). Our molecular data show that Shuteria africana is sister to the genus Amphicarpaea, and that A. ferruginea is sister to A. edgeworthii and A. bracteata within the genus. For a consistent systematic treatment with the discrepancy between the results of the pollen morphology and the present molecular study, a new monotypic genus Afroamphica is proposed for Shuteria africana.

Key words: Afroamphica, Amphicarpaea, Amphicarpaea africana, Amphicarpaea ferruginea, , Leguminosae, new genus, Phaseoleae, Shuteria, Shuteria africana.

Amphicarpaea was revised by Ohashi (Baker 1876) was placed in Amphicarpaea and Ohashi (2016) based mainly on pollen based on such evidence from pollen morphology morphology. The genus was then circumscribed as above, because the former two genera have to consist of three species: A. edgeworthii Benth. typical 3-colporate pollen grains. To confirm the and A. ferruginea Benth., both distributed in classification of these species based on pollen Asia, and A. bracteata (L.) Fernald in North morphology from our previous work (Ohashi America. Amphicarpaea africana (Hook. f.) and Ohashi 2016), we undertook molecular Harms, which had been widely recognized as analyses of these species. a member of the genus in recent taxonomic This paper aims to provide the results of the works (Hauman 1954, Turner and Fearing phylogenetic findings for our treatment on the 1965, Verdcourt 1971), was transferred to taxonomic position for Shuteria africana and Shuteria africana Hook. f., because of the Amphicarpaea ferruginea. distinct tricolporate pollen grains observed in species of Shuteria in contrast to the triporate Phylogenetic confirmation or tetraporate grains with linear grooves or The material used and the voucher specimens slight colporus-like furrows in Amphicarpaea. for Shuteria africana is ‘ of Uganda, In contrast, A. ferruginea, which was once Mobuku Valley, Ruwenzori. Alt. 6200 ft. Vine, attributed to (Kurz 1874) or Shuteria beside river. Flower purple. 2 January 1939.

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Fig. 1. The 50% majority-rule consensus tree (Bayesian analysis) of Amphicarpaea and Shuteria with species of Phaseoleae, Psoraleeae (Bituminaria, Cullen and Otholobium) and Desmodieae (Campylotropis, Desmodium, Kummerowia and Lespedeza) based on cpDNA dataset (trnK/ matK, trnL-trnF). Support values on the branches are presented as follows: BI-PP/ML-BS. NS indicates the node not supported. Scale bar shows the expected number of substitutions per site in Bayesian analysis.

Mrs. M. V. Loveridge 298 (A).’ The methods BS = 92) (Fig. 1). Shuteria africana was sister used in this study are described at the end*. to all three species of Amphicarpaea (BI-PP = The phylogenetic tree based on chloroplast 1; ML-BS = 100). We obtained the trees with DNA (trnL-trnF and trnK/matK) revealed similar topology in both the BI and ML analyses. a clade composed of Shuteria africana and These results show that Shuteria africana is Amphicarpaea (BI-PP = 1; ML-BS = 100), more closely related to Amphicarpaea than to which formed a sister to the clade consisting of other species of Shuteria: S. vestita Wight & max (L.) Merr. and species of the tribe Arn. and S. involucrata (Wall.) Wight & Arn. ex Psoraleeae: Bituminaria bituminosa Kuntze, Walp. Cullen australasicum (Schltdl.) Hanelt, and To confirm the relationship betweenShuteria Otholobium striatum (Thunb.) C. H. Stirt., and africana and the species of Amphicarpaea, a the clade consisting of uncinatus Sw. phylogenetic analysis based on six chloroplast and T. labialis (L. f.) Spreng. (BI-PP = 1; ML- DNA regions (trnL-trnF, trnK/matK, trnT-trnL, February 2018 The Journal of Japanese Botany Vol. 93 No. 1 11

Fig. 2. The 50% majority-rule consensus tree (Bayesian analysis) of Amphicarpaea with closely related species of Phaseoleae based on six chloroplast DNA regions (trnL-trnF, trnK/matK, trnT-trnL, trnG intron, rpl16 itron and trnL-rpl32). Support values on the branches are presented as follows: BI-PP/ML-BS. Scale bar shows the expected number of substitutions per site in Bayesian analysis.

Fig. 3. The 50% majority-rule consensus tree (Bayesian analysis) of Shuteria africana (Amphicarpaea africana) derived from two specimens with species of Phaseoleae based on ca. 600 bp matK fragment. Support values on the branches are presented as follows: BI-PP/ML-BS. Scale bar shows the expected number of substitutions per site in Bayesian analysis. trnG intron, rpl16 itron and trnL-rpl32) was (the other is a sequence analyzed in this report). performed. The deduced tree (Fig. 2) showed The obtained tree (Fig. 3) shows the close a monophyletic Amphicarpaea clade sister to relationship of these two samples. Shuteria africana. The analysis based on nuclear DNA A short fragment of matK sequence of (ITS) also showed the monophyly of the S. africana obtained from another sample is Amphicarpaea-Shuteria africana clade with accessible in GenBank (Accession number: well supported values (BI-PP = 1; ML-BS = KX213379). Therefore we analyzed the 92) and the first separation of Shuteria africana phylogenetic relationship of the two S. africana (Fig. 4). Analysis of ITS using other models samples with this ca. 600 bp matK fragment (e.g., HYK85+G) obtained a similar clade 12 植物研究雑誌 第 93 巻 第 1 号 2018 年 2 月

Fig. 4. The 50% majority-rule consensus tree (Bayesian analysis) of Amphicarpaea and Shuteria with species of Phaseoleae, Psoraleeae (Bituminaria, Cullen and Otholobium) and Desmodieae (Campylotropis, Desmodium, Kummerowia and Lespedeza) based on nuclear DNA (ITS). Support values on the branches are presented as follows: BI-PP/ML-BS. Scale bar shows the expected number of substitutions per site in Bayesian analysis. containing Amphicarpaea and S. africana (data and Ohashi (2016), but supports the inclusion of not shown). Our molecular phylogenetic data on A. ferruginea in Amphicarpaea as indicated by chloroplast and nuclear DNA shows the close morphological features of the pollen. relationships between S. africana and the species The taxonomic position of Shuteria africana of Amphicarpaea. Additionally, the phylogenetic is problematic. The species is distinct from analysis supported the inclusion of A. ferruginea Amphicarpaea in floral and pollen morphology, in Amphicarpaea. but is close to Amphicarpaea based on molecular evidence. The phylogenetic position is Taxonomic treatment far from other species of Shuteria. We consider Results of the present phylogenetic that creating a new genus for the species is more analyses revealed a Shuteria africana- reasonable than retaining it in Amphicarpaea or Amphicarpaea clade composed of S. africana in Shuteria. In the sense of Ohashi and Ohashi and Amphicarpaea. Shuteria africana is sister (2016), Amphicarpaea in the strict sense is to the genus Amphicarpaea. The results did not more natural in morphology and palynology. support the recognition of Shuteria africana as We, therefore, propose here the distinct genus distinct from Amphicarpaea as treated by Ohashi Afroamphica to accommodate Shuteria africana. February 2018 The Journal of Japanese Botany Vol. 93 No. 1 13

Afroamphica H. Ohashi & K. Ohashi, gen. We thank Dr. David E. Boufford of Harvard nov. University Herbaria for supplying the material of [Diagnosis] Afroamphica is morphologically Shuteria africana and for correction of English intermediate between Amphicarpaea and with valuable comments and Dr. T. Nemoto of Shuteria—similar to Amphicarpaea in Ishinomaki Shenshu University for review of the having chasmogamous and cleistogamous manuscript. flowers, but differs from Shuteria in this characteristic. Whereas Afroamphica differs *Methods for phylogenetic analyses from Amphicarpaea in flowers and pollen Genomic DNA was extracted from leaf tissue using the grains, it resembles Shuteria in these characters. DNeasy Mini Kit (Qiagen, Hilden, Germany). Eight In the flowers of Afroamphica, the standard chloroplast markers and ITS were amplified by PCR: trnK intron including matK (trnK/matK), trnL-trnF (trnL intron has a broadly obovate-round blade, the claw of and trnF-trnL spacer), trnT-trnL spacer, rps16 intron, rpl16 the wings is as long as the blade and the acute intron and trnL-rpl32 spacer. The PCR primers used for keel petals; in Amphicarpaea the blade of the these markers are as follows: trnK1L and trnK2R (Hu et al. standard is oblong-obovate, the claw of the 2000) for trnK/matK, primer ‘c’ and Primer ‘f’ (Taberlet et al. 1991) for trnL-trnF (trnL intron and trnF-trnL spacer), wings is 2–3 times longer than blade, and the trnA2 and trnB (Cronn et al. 2002) for trnT-trnL, trnG(UUC)* keel petals are obtuse. (Shaw et al. 2007) and 5ʹtrnG2G (Shaw et al. 2005) for [Description] Herbs, climbing, perennial. trnG, primer 1 and primer 3 (Lee and Hymowitz 2001) for Stems slender, with reflexed ferruginous hairs. rps16 intron, trnL(UAG) and rpl32-F (Shaw et al. 2007) for Leaves pinnately 3-foliolate, stipulate, petiolate; trnL-rpl32, F71 and R1516 (Jordan et al. 1996) for rpl16 intron and ITS4 and ITS5 (White et al. 1990) for ITS. The leaflets elliptic or obovate; stipelate. Racemes PCR condition and DNA sequencing analysis basically with axillary chasmogamous flowers, many- followed Ohashi et al. (2017). The accession numbers of flowered; peduncle and rachis with appressed newly sequenced data in this analysis are shown in Table hairs; bracts narrowly ovate. Chasmogamous 1. The accession numbers of the sequences obtained from GenBank are also listed (Table 2). For the DNA sequences flowers: calyx tubular, 4-lobed, two adaxial of Glycine max and Pachyrhizus erosus used for the lobes connate; petals long clawed; stamens 10, analysis, corresponding portions were obtained from the adaxial stamen free, others connate; disk present whole chloroplast genome sequence deposited in GenBank at base of pistil; ovules 5 per ovary, stigma long (G. max: NC_007942; P. erosus: KJ468100) with reference papillate. Cleistogamous flowers areal, in axils to the annotation. Sequence alignment was initially performed using of chasmogamous racemes, enclosed in calyx. MUSCLE (Edgar 2004) in Mega version 7.0 (Kumar et Legumes linear-oblong, compressed, 4- or al. 2016) and manually adjusted. Phylogenetic analyses 5-seeded, not septate. For additional description were carried out as previously described, using Bayesian see Turner and Fearing (1965) and Verdcourt inference (BI) and maximum likelihood (ML) approaches. An appropriate model of sequence evolution for (1971). the combined dataset was estimated using the program Type species: Afroamphica africana (Hook. Kakusan4 (Tanabe 2007). The dataset of chloroplast DNA f.) H. Ohashi & K. Ohashi, comb. nov. was divided into data partitions (rps16 intron, trnL intron, Shuteria africana Hook. f. in J. Proc. Linn. trnF-trnL spacer, trnT-trnL spacer, rpl16 intron, trnL- Soc., Bot. 7: 190 (1864) [Holotype (2 sheets): rpl32 spacer, 5ʹtrnK, 3ʹtrnK, and matK, that was treated as protein coding sequence). Models of sequence evolution “Cameroon Mountains, alt. 7000 ft. (Fl. Nov.)”, for each of the partitions and for all combined datasets were Nov. 1974, Y. Mann s.n. (K000087082 [sheet determined. 1/2]), K000087083 [sheet 2/2])]; H. Ohashi & Bayesian inference (BI) analyses were conducted using K. Ohashi in J. Jpn. Bot. 91 Suppl.: 247 (2016). MrBayes version 3.2 (Ronquist et al. 2012). The models applied with each analysis based on AIC were as follow. (1) – Amphicarpaea africana (Hook. f.) Harms in Chloroplast trnK/matK and trnL-trnF: GTR+G for all the Repert. Spec. Nov. Regni Veg. 17: 136 (1921), partitions and Proportional_Codon-proportional model was in adnot. selected based on AIC; (2) six chloroplast DNA regions: 14 植物研究雑誌 第 93 巻 第 1 号 2018 年 2 月

Table 1. Voucher information and GenBank accession numbers for data newly sequenced for this paper Taxon Locality Voucher USA, West Virginia, Roane Co. 14 Aug. 2013. K. Campbell 311 (TUS) Amphicarpaea edgeworthii Japan, Miyagi, Murata-machi. 11 Jul. 2014. K. Yamaki 66 (TUS) Amphicarpaea ferruginea (1) China. Yunnan. Da-li. 3 Aug. 1990. J. Murata & al. 62 (TUS) (2) Nepal. Bagmati Zone, Kathmandu Distr. 13 Sep. 1988. M. Suzuki & al. 8881875 (TUS) (3) Thailand. Chiang Rai. alt. 2000–2350 m. 13 Sep. 1967. K. Iwatsuki & al. 9688 (TUS) Harashuteria hirsuta Origin Myanmar, cultivated in Setsunan University, Osaka. H. Murata s.n. (TUS) Pueraria lobata Japan, Iwate, Yahaba, Nishitokuta. 18 Jul. 2017. K. Ohashi 2971 (TUS) Shuteria africana Uganda, Mobuku Valley, Ruwenzori. Jan. 1939. M. V. Loveridge 298 (A) Shuteria involucrata Nepal. Mt. Phulchoki – Godawari. 14 Nov. 1995. M. Mikage & al. 9558381 (TUS) Shuteria vestita Origin Myanmar, cultivated in Tohoku University, Sendai. T. T. Chen s.n. (TUS) 21 Apr. 1986. Teramnus labialis Taiwan, Kaohsiung City, Shoushan Chaishan. 1 Apr. 1992. S. F. Huang 4656 (TUS)

Table 1. Continued GenBank accession number Taxon trnK/matK trnF-trnL trnT-trnL trnG intron rps16 intron rpl16 intron trnL-rpl32 ITS Amphicarpaea bracteata LC315205 LC315111 LC315115 LC315123 LC315130 LC315100 LC315138 LC315209 Amphicarpaea edgeworthii LC315207 LC315107 LC315116 LC315124 LC315132 LC315101 LC315139 LC315210 Amphicarpaea ferruginea (1) LC315204 LC315109 LC315117 LC315125 LC315133 LC315102 LC315140 LC315211 (2) LC315203 LC315110 LC315118 LC315126 LC315134 LC315103 LC315141 LC315212 (3) – – – – – – – LC315213 Harashuteria hirsuta *LC197941 *LC197931 LC315119 LC315127 LC315135 LC315104 LC315143 LC315215 *LC197935 Pueraria lobata LC315202 LC315108 LC315113 LC315121 LC315129 LC315098 LC315142 LC315214 Shuteria africana LC315206 LC315112 LC315114 LC315122 LC315131 LC315099 LC315137 LC315208 Shuteria involucrata – – – – – – – LC315216 Shuteria vestita – – – – – – – LC315217 Teramnus labialis LC315201 LC315106 LC315120 LC315128 LC315136 LC315105 LC315144 LC315218 (1), (2), (3) correspond to the individuals of Amphicarpaea ferruginea in the phylogenetic tree in Fig. 2, 3 and 4. Missing data are indicated by a dash (–). Asterisk indicates data previously published in Ohashi et al. (2017).

GTR for trnT-trnL, 1st, 2nd and 3rd codon of matK and ML bootstrap (Felsenstein 1985) analyses were performed GTR+G for all the other partitions and Proportional_ using RAxML v.8 (Stamatakis 2014), with the partitioned Codon-proportional model; (3) matK fragment: GTR dataset. Equal rate_Codon-equal rate model was selected and Codon-nonpartitioned; (4) ITS: GTR+G. Markov based on AIC. One thousand replicates for bootstrap test Chain Monte Carlo (MCMC) analysis was executed for were performed for ML. All positions containing gaps and 30 million generations with four chains, and sampled missing data were eliminated for BI and ML analysis. every 500th generation. The average standard deviation of split frequencies was checked below 0.01. Tracer References v.1.6.0 (Rambaut et al., 2014) was used to examine the Baker J. G. 1876. Leguminosae. In: Hooker J. D., The Flora convergence of model parameters and check the values of of British India 2: 56–240 (Shuteria pp. 181–182). L. effective sample size (ESS) were over 200. The first 10% Reeve & Co., London. of the trees from each run were discarded as burn-in from Cronn R. C., Small R. L., Haselkorn T. and Wendel J. the final tree set that was used to determine the posterior F. 2002. Rapid diversification of the cotton genus probability distribution. (Gossypium: Malvaceae) revealed by analysis of Maximum likelihood (ML; Felsenstein 1981) and sixteen nuclear and chloroplast genes. Amer. J. Bot. 89: February 2018 The Journal of Japanese Botany Vol. 93 No. 1 15

Table 2. Sequence data already published and obtained from GenBank GenBank accession number Taxon trnK/matK trnF-trnL spacer/trnL intron ITS Apios americana EU717426 EU717312 KF272978 Bituminaria bituminosa EU717398 EU717349 EF517908 Bolusafra bituminosa EU717413 EU717309 – Cajanus cajan EU717414 EU717310 KR863389 Campylotropis macrocarpa EU717418 EU717298 GU572164 Clitoria ternatea EU717286 EU717355 AF467038 Cologania lemmonii EU717405 EU717319 – Cologania pallida JQ619980 – EF517916 Cullen australasicum JQ619979 – EF517909 Desmodium barbatum EU717420 EU717290 – Desmodium floridanum – – EF517898 Dumasia truncata LC197940 LC197930/LC197934 – Erythrina sousae EU717411 EU717313 KT729508 Galactia striata EU717356 EU717428 AF467049 Glycine max AF142700 EU717321 GMU60551 Hardenbergia violacea EU717425 EU717331 – Kennedia nigricans EU717424 EU717335 – Kummerowia stipulacea EU717417 EU717299 GU572167 Lablab purpureus EU717408 EU717339 AY583516 Lespedeza bicolor EU717415 EU717301 KY174476 Macroptilium atropurpureum EU717409 EU717340 AF115138 Macrotyloma uniflorum EU717410 EU717341 AY583527 Neonotonia wightii U717402 EU717323 – Neustanthus phaseoloides EU717404 EU717327 – Ophrestia radicosa EU717430 EU717359 AF467484 Otholobium striatum EF549949 EF543362 EF517851 Pachyrhizus erosus EU717401 EU717324 AY293846 Pseudovigna argentea EU717403 EU717325 – Psophocarpus tetragonolobus EU717412 EU717343 FJ873693 Pueraria candollei – – KC617872 Pueraria montana – – DQ472517 Pueraria tuberosa – – GQ892046 Shuteria vestita* EU717423 EU717328 – Teramnus uncinatus EU717400 EU717330 – Missing data are indicated by a dash (–). *The sequence data of Shuteria vestita shown in this table were used in the phylogenetic tree in Fig. 1. 16 植物研究雑誌 第 93 巻 第 1 号 2018 年 2 月

707–725. Darling A., Höhna S., Larget B., Liu L., Suchard M. Edgar R. C. 2004. MUSCLE: Multiple sequence alignment A. and Huelsenbeck J. P. 2012. MrBayes 3.2: Efficient with high accuracy and high throughput. Nucl. Acids Bayesian phylogenetic inference and model choice Res. 32: 1792–1797. across a large model space. Syst. Biol. 61: 539–542. Felsenstein J. 1981. Evolutionary trees from DNA Shaw J., Lickey E. B., Beck J. T., Farmer S. B., Liu W., sequences: A maximum likelihood approach. J. Molec. Miller J., Siripun K. C., Winder C. T., Schilling E. Evol. 17: 368–376. E. and Small R. L. 2005. The tortoise and the hare Felsenstein J. 1985. Confidence limits on phylogenies: An II: relative utility of 21 noncoding chloroplast DNA approach using the bootstrap. Evolution 39: 783–791. sequences for phylogenetic analysis. Amer. J. Bot. 92: Hauman L. 1954. Glycininae. Fl. Congo Belge et du 142–166. Ruanda-Urunidi 6: 87–113. L’Institut National pour Shaw J., Lickey E. B., Schilling E. E. and Small R. L. 2007. l’Étude Agronomique du Congo Belge (I. N. É. A. C.), Comparison of whole chloroplast genome sequences Bruxelles. to choose noncoding regions for phylogenetic studies Hu J. M., Lavin M., Wojciechowski M. F. and Sanderson in angiosperms: The tortoise and the hare III. Amer. J. M. J. 2000. Phylogenetic systematics of the tribe Bot. 94 : 275–288 . Millettieae (Leguminosae) based on chloroplast trnK/ Stamatakis A. 2014. RAxML version 8: A tool for matK sequences and its implications for evolutionary phylogenetic analysis and post-analysis of large patterns in Papilionoideae. Amer. J. Bot. 87: 418–430. phylogenies. Bioinformatics 30: 1312–1313. Jordan W. C., Courtney M. W. and Neigel J. E. 1996. Low Taberlet P., Gielly L., Pautou G. and Bouvet J. 1991. levels of intraspecific genetic variation at a rapidly Universal primers for amplification of three non-coding evolving chloroplast DNA locus in North American regions of chloroplast DNA. Plant Mol. Biol. 17: duckweeds (Lemnaceae). Amer. J. Bot. 83: 430–439. 1105–1109. Kumar S., Stecher G. and Tamura K. 2016. MEGA7: Tanabe A. S. 2007. Kakusan: a computer program to Molecular Evolutionary Genetics Analysis version 7.0 automate the selection of a nucleotide substitution for bigger datasets. Molec. Biol. Evol. 33: 1870–1874. model and the configuration of a mixed model on Kurz S. 1874 (dt. 1873). New Burmese plants, part III. J. multilocus data. Mol. Ecol. Notes 7: 962–964. Asiat. Soc. Bengal, Pt. 2, Nat. Hist. 42(4): 204–310. Turner B. L. and Fearing O. S. 1965. A taxonomic study Lee J. and Hymowitz T. 2001. A molecular phylogenetic of the genus Amphicarpaea (Leguminosae). Southw. study of the subtribe Glycininae (Leguminosae) derived Naturalist 9: 207–218. [Different two dates are printed from the chloroplast DNA rps16 intron sequences. on the top: ‘Issued 15 January, 1965’ above, ‘Dec. 25, Amer. J. Bot. 88(11): 2064–2073. 1964’ on the first page at right corner]. Ohashi H. and Ohashi K. 2016. A taxonomic revision Verdcourt B. 1971. Tribe 10. Phaseoleae. In: Gillett J. of Amphicarpaea (Leguminosae) including a pollen B., Polhill R. M. and Verdcourt B., Flora of Tropical morphological comparison with Shuteria. J. Jpn. Bot. East Africa. Leguminosae (Part 4) Subfamily 91 Suppl.: 231–249. Papilionoideae. pp. 503–807. Crown Agents for Ohashi K., Nata K. and Ohashi H. 2017. Harashuteria, Oversea Governments and Administrations, London. a New Genus of Leguminosae (Fabaceae) tribe White T. J., Bruns T., Lee S. and Taylor J. 1990. Phaseoleae. J. Jpn. Bot. 92: 34–43. Amplification and direct sequencing of fungal Rambaut A., Suchard M. A., Xie D. and Drummond A. J. ribosomal RNA genes for phylogenetics. Chapter 38. 2014. Tracer v1.6, Available from http://beast.bio.ed.ac. In: Innis M., Gelfand D., Sninsky J. and White T. (eds.), uk/Tracer. PCR Protocols: a Guide to Methods and Applications. Ronquist F., Teslenko M., Van der Mark P., Ayres D. L., pp. 315–322. Academic Press, San Diego.

a b 大橋一晶 ，大橋広好 ：マメ科ヤブマメ属の分子系統解 析と新属 Afroamphica の設立 マメ科マメ亜科インゲンマメ連に属するヤブマメ属 て，それに所属する属の中で両属は近い位置に置かれて Amphicarpaea はアフリカに 1 種，アジアに 2–3 種，北 きた．また，これまでに調べられた両属の一部の種類 アメリカ 1 種が分布し，形態的には Shuteria に近縁と では花粉形態において明瞭な違いが知られていた．し さ れ， ダ イ ズ 連 (Hutchinson J., Gen. Fl. Pl. 1, 1964) ま かし，分子系統解析ではヤブマメ属は Shuteria と全く たはダイズ亜連 (Lackey J. A. in Polhill R. M. and Raven 離れた位置にあることが明らかにされている (Lee and P. H. (eds.), Adv. Leg. Systematics 1, 1981) に 分 類 さ れ Hymowitz 2001)．このため，ヤブマメ属全種につい February 2018 The Journal of Japanese Botany Vol. 93 No. 1 17

て花粉形態を精査し，Shuteria と比較した (Ohashi and ら れ な か っ た． し か し，Amphicarpaea ferruginea は Ohashi 2016)． そ の 結 果，A. africana (Hook. f.) Harms Amphicarpaea clade に属し，本種をヤブマメ属とする花 の花粉形態は Shuteria と一致することが明らかとなり， 粉形態の証拠と分子系統の証拠は一致した．この点か Shuteria africana に戻された (Ohashi and Ohashi 2016)． らヤブマメ属 (Ohashi and Ohashi 2016) は形態・花粉形 一方，形態的に分類学上の位置について疑問のあった 態・分子系統において矛盾なくまとまった群と考えられ Amphicarpaea ferruginea は花粉形態では Amphicarpaea る．一方，S. africana (= Amphicarpaea africana) につい と一致した (Ohashi and Ohashi 2016)．そのため本研究 ては，花粉形態と分子系統の証拠の矛盾を解決するた では S. africana とヤブマメ属の関係を分子系統解析に め，新属 Afroamphica H. Ohashi & K. Ohashi を設立し， よって再検討した．S. africana はヤブマメ属と Shuteria Afroamphica africana (Hook. f.) H. Ohashi & K. Ohashi と africana-Amphicarpaea clade を形成し，クレード内で することを提案した． S. africana は最初に分岐し，ヤブマメ属の姉妹群であ （a岩手医科大学薬学部， った．S. africana は花粉形態で Shuteria 属との類縁性 b東北大学植物園津田記念館） が示唆されたが，これを支持する分子系統の証拠は得