TAXON 58 (1) • February 2009: 133–140 Stoffelen & al. • Coffea anthonyi, a new coffee species Coffea anthonyi, a new self-compatible Central African coffee species, closely related to an ancestor of Coffea arabica Piet Stoffelen1, Michel Noirot2, Emmanuel Couturon2, Sylvain Bontems3, Petra De Block1 & François Anthony3 1 National Botanic Garden of Belgium, Domein van Bouchout, 1860 Meise, Belgium. piet.stoffelen@br.fgov.be (author for correspondence) 2 Institut de Recherche pour le Développement, Université de la Réunion, Campus du Chaudron, BP 172, 97492 Sainte Clotilde cedex, La Réunion, France 3 Institut de Recherche pour le Développement, BP 64501, 34394 Montpellier cedex 5, France Coffea anthonyi Stoff. & F. Anthony from Cameroon and the Republic of Congo is formally described. Morphological, biochemical and molecular characteristics, isozyme diversity and phylogenetic relationships are discussed. The new diploid species has strong relationships with the East African C. eugenioides and the allotetraploid cultivated C. arabica. As such, it could be related to a progenitor species of C. arabica. The new species is self-compatible, being a rare character within the genus Coffea. This new small-leaved Central African species is of particular interest from a phylogenetic as well as from a breeding point of view. KEYWORDS: Central Africa, Coffea anthonyi, coffee, genetic diversity, self-compatibility INTRODUCTION Coffea belongs to the Rubiaceae and presents a high diversity as shown by the large number of species. Many new taxa have been recently discovered (Stoffelen & al., 1996, 1997a, b, 1999, 2008; Davis, 2001; Davis & Rakotonasolo, 2000, 2001a, b, 2003; Cheek & al., 2002; Davis & Mvungi, 2004; Sonké & Stoffelen, 2004). Up to now, 103 Coffea species have been identified in the forests of continental Africa and Madagascar (Davis & al., 2006). All species are perennial woody shrubs or trees, and differ greatly in morphology, size, and ecological adaptation. They are diploid (2n = 2x = 22) except for C. arabica L., which is allotetraploid (2n = 4x = 44) (Krug & Carvalho, 1951; Lashermes & al., 1999). Commercial coffee production mainly relies on two species, C. arabica and C. canephora Pierre ex A. Froehner, with better quality coffee being associated with C. arabica. Genes from diploid species can be transferred into the C. canephora or C. arabica genomes by interspecific hybridization. Thus, wild coffee represents an important and valuable source of genetic diversity for coffee improvement. However, natural populations of coffee trees have been threatened by massive deforestation occurring in Africa and Madagascar since many decades. Because of the lack of diversity conserved in the existing coffee genebanks, international (FAO and IPGRI) and French organisations (IRD ex-ORSTOM, CIRAD, MNHN) have made a concerted effort to collect coffee germplasm. During the last 40 years the IRD has organized and conducted collection missions in Ethiopia (Guillaumet & Hallé, 1978), Kenya (Berthaud & al., 1980), Tanzania (Anthony & al., 1987), Central African Republic (Berthaud & Guillaumet, 1978), Cameroon (Anthony & al., 1985), Republic of the Congo (Namur & al., 1987), Ivory Coast (Berthaud, 1986) and Guinea (Le Pierrès & al., 1989). A total of 7,800 wild coffee genotypes belonging to ca. 20 taxa were collected and introduced in a field genebank in Ivory Coast (Anthony, 1992). Here, we describe a new coffee species, C. anthonyi, found in southern Cameroon and the border region between Cameroon (Anthony & al., 1985) and the Republic of the Congo (Namur & al., 1987). The discovery of C. anthonyi confirms the presence of an important diversity centre for coffee in West Central Africa. The number of coffee species occurring in Cameroon was underestimated until the recent description of several new taxa (Stoffelen & al. 1996, 1997a, b, 1999, 2008; Cheek & al. 2002; Sonké & Stoffelen 2004; Sonké & al. 2006). In 1995, only five species were known; this number has now increased to fourteen species. Isozyme diversity of C. anthonyi is assessed and compared with that of well-known Central African species, such as C. brevipes Hiern, C. canephora, C. liberica Hiern and C. mayombensis A. Chev. Furthermore, biochemical, genetic and molecular data concerning the new species are reviewed. Many published data exist, since plant material of C. anthonyi has been grown and studied for many years by coffee researchers, mainly from the IRD in Montpellier. Until now, these data have been published using informal taxon names and therefore need to be linked to the accepted name. 133 TAXON 58 (1) • February 2009: 133–140 Stoffelen & al. • Coffea anthonyi, a new coffee species MATERIALS AND METHODS Germplasm collection. — Germplasm collection missions took place in January–March of 1983 in Cameroon (Anthony & al., 1985) and in August-September of 1985 in the Republic of the Congo (Namur & al., 1987). The new species was found in three sites in Cameroon (C27, C29, C30) and in two sites in the Republic of the Congo (Cg15, Cg17) (Fig. 2 below). The collected material comprised 5 cuttings at C27, 1 cutting at C29, 3 cuttings at C30, 2 cuttings at Cg15, and 48 cuttings at Cg17. Seeds were also collected at C27 (2 seeds from one tree), at Cg15 (4 seeds from one tree) and Cg17 (27 seeds from four trees). Locality data of the populations were as follows: C27 (Moloundou-1 population): N2°03 E15°05 ; C29 (Moloundou-2 population): N2°04 E15°00 ; C30 (Moloundou-3 population): N2°06 E14°56 ; Cg15 (Souanké-Gabon-1 population): N2°07 E14°00 ; Cg17 (Souanké population): N2°03 E14°08 . Study of herbarium material. — Six herbarium specimens were studied for the botanical description (BR, BRLU, K, LISC, P, WAG). All other material of Coffea from these herbaria was screened for the new species. Isozyme analysis. — The studied material consisted of young leaves from 31 accessions of C. anthonyi and 35 accessions of C. mayombensis, each species represented by two populations (Table 1) (Anthony & al., 1985; Namur & al., 1987). Electrophoresis data were recorded following the methods described by Berthaud (1986). Enzymes were α-esterase (EST), isocitrate dehydrogenase (ICD), phosphoglucoisomerase (PGI), phosphoglucomutase (PGM), and 6-phosphogluconate dehydrogenase (PGD). A total of 33 allelic frequencies were scored for each sample. Frequencies per population were then calculated and analysed using the joining method of Statistica software (© Statsoft, Inc.) to produce a hierarchical tree. Data of three well-known species (i.e., C. brevipes, C. canephora, C. liberica) native to Central Africa were taken from Berthaud (1986) and included in a comparative analysis (Table 1). Controlled self-pollination. — The method of controlled hybridisations is routinely used in coffee breeding programs using interspecific hybrids (Van der Vossen, 1985), particularly in Ivory Coast (Louarn, 1992). Self-pollination can be controlled by bagging flowering branches before anthesis and pollination. At anthesis, branches are shaken to induce self-pollination. The bags are removed two days later. DESCRIPTION OF THE NEW SPECIES Coffea anthonyi Stoff. & F. Anthony, sp. nov. – Holotype: Anthony F. 20 (BR0000006920555), grown at Divo coffee genebank, Ivory Coast, from cuttings collected in the Republic of the Congo, at Souanké population, 1999 (Fig. 1). Diagnosis: Coffea anthonyi differt ab specie Africae occidentalis et Centralis prope foliis minutis et obovatis. Coffea sp. ‘Moloundou’, informal name in Anthony, 1992: 194; Anthony & al., 1993: 65–70; Cros & al., 1995: 15–18, 1998: 110–115; Cros, 1996: 14–138; Lashermes & al., 1996: 628–630, 1997: 948–954, 1999: 260–164; Stoffelen, 1998: 125; Combes & al., 2000: 1179; Noirot & al., 2003: 710–713; Campa & al., 2004: 40–43, 2005a: 634–636, 2005b: 135–139; Poncet & al., 2004: 1072–1078, 2005: 2–5; Anthony & Lashermes, 2005: 210–214. Coffea sp. ‘Dja Mékas’, informal name in Stoffelen, 1998: 123–124; Sonké & Stoffelen, 2004: 157. Table 1. Material included in the isozyme analysis. No. of accessions Species Country Population C. anthonyi Cameroon Moloundou-1 Republic of the Congo Souanké 26 C. brevipes Cameroon Mbonzi 9 C. canephora Central African Republic Doungba 9 Central African Republic Libengué 45 Central African Republic Ndongue 84 Cameroon Mixeda 16 C. liberica Cameroon a Mixed C. mayombensis Republic of the Congo Dimonika-1 Republic of the Congo Mvouti-2 5 8 31 4 Data of Coffea brevipes, C. canephora and C. liberica were taken from Berthaud (1986). a 134 i.e., from several populations TAXON 58 (1) • February 2009: 133–140 Stoffelen & al. • Coffea anthonyi, a new coffee species Fig. 1. Coffea anthonyi Stoff. & F. Anthony. A, flowering branch; B, single-flowered inflorescence; C, fruit; D, ventral view of seed; E, cross section of seed. Small shrub up to 2 m tall, young branchlets glabrous. Leaf-blades obovate, rarely elliptic, 5–9(–10) cm long, 2.2–4(–5) cm wide; base cuneate; tip acuminate, acumen 5–8(–10) mm long, with rounded tip; minute crypt domatia situated at the junction of midrib and secondary nerves in the basal half of the leaf, visible as ridges with minute openings, glabrous; 5–7 secondary nerves at each side of the midrib. Petioles 5–10 mm long. Stipules deltate, limb ca. 2 mm long, ciliate. Inflorescences shortly pedunculate, 1–2 per axil, each with 1–2 flowers and 2–3 calyculi; peduncle minute; calyculi four-lobed, with two stipular and two small foliar lobes. Flowers 5-merous. Calyx truncate. Corolla white; tube ca. 5 mm long; lobes 6–7 mm long, ca. 3 mm wide. Stamens with anthers 4.5–5 mm long, filaments ca. 0.5 mm long. Gynoecium glabrous. Small annular disc present. Style ca. 10 mm long; stigma bilobed, ca. 2 mm long. Infructescences with peduncle 2.5–3 mm long. Fruits red, 9–10 mm long, 6–8 mm wide. Seeds broadly elliptic, 7–8 mm long, 5.5–6 mm wide, ca. 3 mm thick. Silver skin parenchymatic and provided with parallelly aggregated, elongated sclereids; sclereids with circular or lenticular perforations. 135 Stoffelen & al. • Coffea anthonyi, a new coffee species Material studied. — Cameroon: Breteler F.J. 853 (K, LISC, WAG), 5 km east of Bertoua, Batouri-BetaréOya, N4°35 E13°41; Breteler F.J. 2189 (LISC, P, WAG), Gounté, 27 km on road Bertoua-Bétaré-Oya, N4°42 E13°49 ; Sonké B. 1412 (BR, BRLU): réserve de faune du Dja Mékas, N3°11 E12°33 . Republic of the Congo: Anthony F. 19 (BR), grown in Divo coffee genebank, Ivory Coast from cuttings collected in the Republic of the Congo, at Souanké-Gabon-1 population, N2°07 E14°00 ; Anthony F. 20 (BR), grown in Divo coffee genebank, Ivory Coast from cuttings collected in the Republic of the Congo, at Souanké population, N2°03 E14°08 ; Anthony F. 21 (BR), grown in Divo coffee genebank, Ivory Coast from cuttings collected in the Republic of the Congo, at Souanké population, N2°03 E14°08 . Distribution and habitat. — Coffea anthonyi is endemic in South East Cameroon and North West Republic of the Congo (Fig. 2). Almost all sources mention that this species is found in “primary” rain forest; Breteler 853 states “gallery forest, mostly secondary, marshy soil”. In the wild, C. anthonyi is often associated with other Coffea or Psilanthus species. It was found together with C. canephora (population Cg15), C. liberica var. dewevrei (De Wild. & T. Durand) Lebrun and Psilanthus mannii Fig. 2. Distribution map of Coffea anthonyi Stoff. & F. Anthony. + = collecting locality of C. anthonyi (herbarium specimens or germplasm collection). 136 TAXON 58 (1) • February 2009: 133–140 Hook. f. (population C27), and with P. mannii (populations C29, C30 and Cg17). Population Cg17 was found on ca. 1,000 m², on a slope which was covered by an old forest of Triplochiton scleroxylon K. Schum. on a rocky soil with rocky ravines and sources. History of species recognition. — The new species was identified as Coffea sp. nov. by the IRD collectors because the leaf and fruit morphology did not fit with any described coffee species (Lebrun, 1941; Chevalier, 1947; Bridson & Verdcourt, 1988). In previous publications this species was named Coffea sp. ‘Moloundou’. It was linked by A. Charrier (written note on folder of Breteler 2189 & 853 in WAG) to herbarium material later named as the informal taxon Coffea sp. ‘Dja Mékas’ (Stoffelen, 1998; Stoffelen & al., 1999). Coffea sp. ‘Dja Mékas’ was not formally described, as only fruiting specimens were available and observation of flowers is absolutely necessary to identify Coffea species, because in some cases fruiting individuals of Coffea and Psilanthus Juss. ex M. Roem. are very hard to distinguish. In 1999 herbarium material of the new species collected in Cameroon and the Republic of the Congo and grown in the Divo coffee collection in Ivory Coast was shipped to the IRD in Montpellier. Study of this new material affirmed that the species at hand is new to science and that Coffea sp. ‘Moloundou’ and Coffea sp. ‘Dja Mékas’ are ‘synonyms’. One collection of the Democratic Republic of the Congo (Sapin B2 [BR], Katola, Sandoa) is reminiscent of this new species but material is too poor to be sure about its identity (immature fruits). Comparison with morphologically similar Coffea species. — The new species is one of the few small-leaved coffee species from Central Africa. Only two other smallleaved species have been described for Central and West Africa: C. kapakata (A. Chev.) Bridson (Chevalier, 1947; Bridson, 1994) and C. charrieriana Stoff. & F. Anthony (Stoffelen & al., 2008). Coffea kapakata can be distinguished from C. anthonyi because the former is a savannah species from Angola with typical elliptic leaves, lobed calyces and costate fruits. Coffea charrieriana has elliptic or broadly elliptic, seldom obovate leaves and very small anthers (ca. 3 mm long), while C. anthonyi has obovate, seldom elliptic leaves and anthers of 4.5–5 mm long. The morphology of the new species is reminiscent of the morphology of the East African species C. eugenioides S. Moore, but it differs from the latter by its obovate leaves (vs. elliptic in C. eugenioides), shorter peduncles (only 2–3 mm vs. 5–16 mm in C. eugenioides) and shorter anther filaments (0.5 mm vs. 0.8–3 mm in C. eugenioides). Isozyme diversity and differentiation. — Presence or absence of 33 alleles was recorded in 237 coffee accessions belonging to five species from Central Africa (see Table 1), and allelic frequencies were then calculated per population. TAXON 58 (1) • February 2009: 133–140 Stoffelen & al. • Coffea anthonyi, a new coffee species The resulting hierarchical tree clearly shows several levels of clustering (Fig. 3). As expected, populations belonging to the same species are associated together at the lowest level of clustering: C. mayombensis, C. canephora and C. anthonyi. The C. brevipes population is grouped with the C. canephora populations at the following level. This grouping was also expected because C. brevipes can be considered as an ecotype of C. canephora (Chevalier, 1947; Anthony, 1992; Stoffelen, 1998). The third level of clustering associates the C. mayombensis populations with the C. canephora-C. brevipes group. At the fourth level C. liberica is associated with this multispecific group, while at the final level C. anthonyi is grouped with the other species. This classification confirms the high genetic differentiation (0.74) between C. canephora and C. liberica estimated on the basis of molecular data (N’Diaye & al., 2005). Since C. anthonyi groups with the other species only at the highest level of the hierarchical tree, its differentiation with other species originating from the same region could be even higher than the genetic differentiation between C. canephora and C. liberica. This distinct position of C. anthonyi is supported by two specific alleles (i.e., PGD1-X, ICD-I) that were not detected in populations of other species. Four alleles (i.e., EST-H, PGD2-G, PGI-D, PGM-F) were fixed in the C. anthonyi populations and found in the other populations with variable frequencies. Two alleles (i.e., EST-F, PGM-E) were detected in one or two accessions of C. anthonyi and correspond to heterozygous genotypes. The presence of six fixed alleles, two of which had never been observed in coffee, pointed out the original enzyme patterns of the new species. The other alleles (25) identified in the study were present in at least one accession of other species, but not in C. anthonyi. In another study, a specific pattern (Z4) was also found in the acid phosphatase system (Berthaud, 1986). Biology: self-compatibility, ploidy and phenology. — Self-compatibility of the new species was sus- pected during the visit of the C27 population. Here, a single coffee tree (2 m high) bearing one fruit was found together with two young plants (20–40 cm high). No other adult tree was discovered although an intensive search was conducted over a large area. Such scarceness of reproductive plants is rather rare in coffee populations because selfincompatibility in diploid species requires the presence of two trees for fecundation. Moreover, the harvesting of two seeds from the single fruit indicated the successful fecundation of the two ovules. Again, this is not very frequent in self-incompatible coffee species because of random pollination by insects. Self-compatibility was then confirmed through controlled self-pollination experiments in the field genebank of Ivory Coast (Anthony, 1992), with a fruit set (29.7%) similar to the fruit set of open-pollinations in the self-compatible species C. arabica (Berthaud, 1978). The reproduction cycle was estimated to take about eight months. As a consequence of the self-compatibility in C. anthonyi, heterozygosity is low. Only two heterozygous patterns displayed by three accessions were identified. This confirms previous results comparing heterozygosity in self-compatible and self-incompatible species, based on co-dominant molecular markers. The proportion of heterozygous loci detected by RFLP (Restriction Fragment Length Polymorphism) markers was 4% in C. anthonyi but 23% in C. eugenioides, 27% in C. congensis A. Froehner and 36% in C. canephora (Lashermes & al., 1999). This proportion was higher (10%) using SSR (Simple Sequence Repeat) markers (Poncet & al., 2004). Self-compatibility has been reported so far in two other coffee species: the allotetraploid species C. arabica (Carvalho & al., 1991) and a diploid species from Cameroon, C. heterocalyx Stoff. (Coulibaly & al., 2002). C. mayombensis-1 C. mayombensis-2 C. canephora-1 Fig. 3. Hierarchical tree based on allelic frequencies of five enzyme systems (237 individuals analysed). C. canephora-2 C. canephora-3 C. canephora-4 C. brevipes C. liberica C. anthonyi-1 C. anthonyi-2 3 2 1 0 Aggregating distance 137 Stoffelen & al. • Coffea anthonyi, a new coffee species In self-compatible plants, productivity is less dependent on environmental factors than in incompatible ones, because fecundation success is less or not influenced by field association of at least two cross-compatible plants, synchronous flowering, and the availability of pollinating agents such as wind and insects. Introgression of selfcompatibility from C. anthonyi or C. heterocalyx into C. canephora cultivars would facilitate the selection of homozygous cultivars and their distribution by seeds. Information on ploidy in C. anthonyi can be deduced from molecular studies based on co-dominant markers. A maximum of two alleles per locus has always been observed using isozyme, microsatellite (Combes & al., 2000; Poncet & al., 2004) and RFLP (Lashermes & al., 1999) markers. This strongly supports the hypothesis that C. anthonyi is diploid, like all other Coffea species except C. arabica. The genomic characteristics of C. anthonyi do not differ much from those of the other coffee species. The nuclear DNA content has been estimated using flow cytometry with either external (Cros & al., 1995) or internal standardisation (Noirot & al., 2003). Using internal standardisation, the genome size of C. anthonyi was about 1.44 pg, showing no significant variation between the Moloundou and Souanké populations. In contrast, between species variation exists and C. anthonyi was found to differ from C. eugenioides (1.36 pg) and C. heterocalyx (1.74 pg) (Noirot & al., 2003). Using SSR or AFLP (Amplified Fragment Length Polymorphism) markers, the new species showed amplification products similar to those of well-known species such as C. arabica, C. canephora and C. liberica (Combes & al., 2000; Poncet & al., 2004, 2005). Such markers shared by several species could be used for constructing a consensus genetic map of coffee. Gene flow between related species is often limited by reproductive barriers. The most common reproductive barrier in Coffea is due to chromosome pairing limitation during meiosis (Louarn, 1992) leading to decreasing pollen fertility. In coffee plants, maturing of flowers is triggered by rain with anthesis occurring five to eight days after rainfall and lasting only one day. Different interval lengths from rainfall to flowering limit pollination between species and constitute an important reproductive barrier. In C. anthonyi, the delay is seven days. This is also the case for C. canephora, whereas it is eight days for C. eugenioides and C. arabica (Baranski, unpublished data). Two other phenological differences, but without impact on gene flow, characterize the new species in comparison to C. eugenioides. Indeed, fruit growth begins five weeks after flowering, and fruits are mature after 27 weeks in C. anthonyi, while in C. eugenioides these delays are respectively 7 and 23 weeks (Baranski, unpublished data). 138 TAXON 58 (1) • February 2009: 133–140 Biochemical characterisation of the coffee beans. — The new species does not differ from other Coffea species in the biochemical composition of its beans. The caffeine content of the beans is low, about 0.6% dry matter basis (dmb) (Anthony & al., 1993; Campa & al., 2005a), and does not differ from that of C. eugenioides. Its chlorogenic acid content (5.5% dmb) is intermediate between C. charrieriana (0.8% dmb) and Coffea sp. N’koumbala (11.9% dmb), the two extremes in the subgenus Coffea (Campa & al., 2005b). Its trigonelline content is 1.6% dmb, one of the highest of the sub-genus, but does not differ from C. heterocalyx (1.5% dmb) and C. eugenioides (1.3% dmb) (Campa & al., 2004). Lastly, its sucrose content is 5.7% dmb, statistically similar to that of C. heterocalyx (6.2% dmb), but significantly lower than that of C. eugenioides (7.7% dmb) (Campa & al., 2004). Phylogenetic relationships. — Phylogenetic relationships of coffee species were studied using data from conserved regions of the chloroplast and ribosomal genomes. Variations in cpDNA were assessed through a RFLP analysis on total cpDNA and on the atpB-rbcL intergenic region (Lashermes & al., 1996), and by sequencing the trnL-trnF intergenic spacer (Cros, 1996; Cros & al., 1998). Other data were obtained through an analysis of the internal transcribed spacer (ITS 2) sequences of rDNA (Lashermes & al., 1997). In all these studies, the new species was grouped with C. eugenioides. The rDNA nucleotide-sequence divergence between C. anthonyi and C. eugenioides was estimated to be similar to the divergence between populations of the same species (Lashermes & al., 1997). No difference was detected in the cpDNA sequences of both species (Lashermes & al., 1996; Cros & al., 1998). Furthermore, the cpDNA polymorphism data classified C. anthonyi and C. eugenioides with the allotetraploid species C. arabica (Cros & al., 1998). The relationships between C. arabica, C. anthonyi and C. eugenioides were then confirmed by an analysis of allelic constitution of RFLP loci detected by various single-copy probes in C. arabica and several potential diploid progenitor species (Lashermes & al., 1999; Maurin & al., 2007). Since cpDNA has a strict maternal inheritance in coffee (Lashermes & al., 1996), C. anthonyi and C. eugenioides could be related to the maternal progenitor species of the allotetraploid species C. arabica. The genetic closeness of C. anthonyi and C. eugenioides remains to be investigated in detail since the two species have disjunct distribution areas and are adapted to a different habitat. Coffea anthonyi occurs in low altitude forest (360–650 m) in Cameroon and the Republic of the Congo while C. eugenioides has been reported from montane and submontane forests (1,050– 2,100 m) in the Eastern parts of the Democratic Republic of the Congo, Uganda, Kenya and Tanzania (Bridson & Verdcourt, 1988; Stoffelen, 1998). TAXON 58 (1) • February 2009: 133–140 CONCLUSION Coffea anthonyi can be easily distinguished from other coffee species originating from Central Africa using morphological or isozyme data. Its self-compatibility is a rare trait in Coffea species, and has favoured the homogenisation of its genetic structure. As a consequence, heterozygosity in C. anthonyi is significantly lower than in self-incompatible species. The occurrence of C. anthonyi in the border region of Cameroon and the Republic of the Congo confirms the presence of an important diversity centre for coffee in West Central Africa. Morphological and macromolecular data clearly show that C. anthonyi is closely related to the allotetraploid species C. arabica and to the diploid East African species C. eugenioides. Therefore, C. anthonyi could represent a key species for understanding the origin of C. arabica. ACKNOWLEDGEMENTS We thank the International Plant Genetic Resources Institute (IPGRI) for funding the collection missions of wild coffee trees in Cameroon and the Republic of the Congo. The authors are extremely grateful to the curators of the following herbaria: BM, BR, BRLU, COI, HBG, K, M, MO, LISC, P and WAG. Antonio Fernandez (BR) and Marcel Verhaegen (BR) are acknowledged for preparing the line drawing and the distribution map respectively. Two anonymous reviewers are thanked for their useful comments. LITERATURE CITED Anthony, F. 1992. Les ressources génétiques des caféiers: collecte, gestion d’un conservatoire et évaluation de la diversité génétique. Travaux et Documents Microédités, No. 81. ORSTOM, Paris. Anthony, F., Berthaud, J., Guillaumet, J.L. & Lourd, M. 1987. Collecting wild Coffea species in Kenya and Tanzania. Pl. Genet. Resources Newslett. 69: 23–29. Anthony, F., Clifford, M.N. & Noirot, M. 1993. Biochemical diversity in the genus Coffea: chlorogenic acids, caffeine and mozambioside contents. Genet. Resources Crop Evol. 40: 61–70. Anthony, F., Couturon, E. & Namur, C. de. 1985. Résultats d’une mission de prospection effectuée par l’ORSTOM en 1983. Pp. 495–505 in: Proceedings of 11th International Scientific Colloquium on Coffee. Association Scientifique Internationale du Café, Lausanne. Anthony, F. & Lashermes, P. 2005. The coffee (Coffea arabica L.) genome: diversity and evolution. Pp. 207–228 in: Sharma, A.K. & Sharma, A. (eds.), Plant Genome: Biodiversity and Evolution, vol. 1B, Phanerogams. Science Publishers Inc., New Hampshire. Berthaud, J. 1978. L’hybridation interspécifique entre Coffea Stoffelen & al. • Coffea anthonyi, a new coffee species arabica L. et Coffea canephora Pierre. Obtention et comparaison des hybrides triploïdes, Arabusta et hexaploïdes. Café, Cacao, Thé 22: 3–12. Berthaud, J. 1986. Les ressources génétiques pour l’amélioration génétique des caféiers africains diploïdes. Travaux et documents, No. 188. ORSTOM, Paris. Berthaud, J. & Guillaumet, J.L. 1978. Les caféiers sauvages en Centrafrique: résultats d’une mission de prospection (janvier-février 1975). Café, Cacao, Thé 3: 171–186. Berthaud, J., Guillaumet, J.L., Le Pierrès, D. & Lourd, M. 1980. Les caféiers sauvages du Kenya: prospection et mise en culture. Café, Cacao, Thé 24: 101–112. Bridson, D. 1994. Additionnal notes on Coffea (Rubiaceae) from Tropical East Africa. Kew Bull. 49: 331–342. Bridson, D. & Verdcourt, B. 1988. Coffea. Pp. 703–723 in: Polhill, R.M. (ed.), Flora of Tropical East Africa, part 2. A.A. Balkema, Rotterdam. Campa, C., Ballester, J.M., Doulbeau, S., Dussert, S., Hamon, S. & Noirot, M. 2004. Trigonelline and sucrose diversity in wild Coffea species. Food Chem. 88: 39–43. Campa, C., Doulbeau, S., Dussert, S., Hamon, S. & Noirot, M. 2005a. Diversity in bean caffeine content among wild Coffea species: evidence of a discontinuous distribution. Food Chem. 91: 633–637. Campa, C., Doulbeau, S., Dussert, S., Hamon, S. & Noirot, M. 2005b. Qualitative relationship between caffeine and chlorogenic acid contents among wild Coffea species. Food Chem. 93: 135–139. Carvalho, A., Medina Filho, H.P., Fazuoli, L.C., Guerreiro Filho, O. & Lima, M.M.A. 1991. Aspectos genéticos do cafeeiro. Revista Brasil. Genet. 14: 135–183. Cheek, M., Csiba, L. & Bridson, D. 2002. A new species of Coffea (Rubiaceae) from western Cameroon. Kew Bull. 57: 675–680. Chevalier, A. 1947. Les vrais et les faux caféiers. Nomenclature et systématique. Encycl. Biol. 28: 115–263. Combes, M.C., Andrzejewski, S., Anthony, F., Bertrand, B., Rovelli, P., Graziosi, G. & Lashermes, P. 2000. Characterization of microsatellite loci in Coffea arabica and related coffee species. Molec. Ecol. 9: 1178–1180. Coulibaly, I., Noirot, M., Lorieux, M., Charrier, A., Hamon, S. & Louarn, J. 2002. Introgression of self-compatibility from Coffea heterocalyx to the cultivated species Coffea canephora. Theor. Appl. Genet. 105: 994–999. Cros, J. 1996. Implications phylogénétiques des variations de l’ADN chloroplastique chez les caféiers (genres Coffea L. et Psilanthus Hook. f.). Doctoral thesis, Thèses et Documents Microfichès, No. 147. ORSTOM, Paris. Cros, J., Combes, M.C., Chabrillange, N., Duperray, C., Monnot des Angles, A. & Hamon, S. 1995. Nuclear content in the subgenus Coffea (Rubiaceae): inter- and intra-specific variation in African species. Canad. J. Bot. 73: 14–20. Cros, J., Combes, M.C., Trouslot, P., Anthony, F., Hamon, S., Charrier, A. & Lashermes, P. 1998. Phylogenetic relationships of Coffea species: new evidence based on the chloroplast DNA variation analysis. Molec. Phylog. Evol. 9: 109–117. Davis, A.P. 2001. Two new species of Coffea L. (Rubiaceae) from eastern Madagascar. Kew Bull. 56: 479–489. Davis, A.P., Govaerts, R., Bridson, D.M. & Stoffelen, P. 2006. An annotated taxonomic conspectus of the genus Coffea (Rubiaceae). Bot. J. Linn. Soc. 152: 465–512. 139 Stoffelen & al. • Coffea anthonyi, a new coffee species Davis, A.P. & Mvungi, E.F. 2004. Two new endangered species of Coffea (Rubiaceae) from the Eastern Arc Mountains (Tanzania) and notes on associated conservation issues. Bot. J. Linn. Soc. 146: 237–245. Davis, A.P. & Rakotonasolo, F. 2000. Three new species of Coffea L. (Rubiaceae) from Madagascar. Kew Bull. 55: 405–416. Davis, A.P. & Rakotonasolo, F. 2001a. Two new species of Coffea L. (Rubiaceae) from northern Madagascar. Adansonia 23: 337–345. Davis, A.P. & Rakotonasolo, F. 2001b. Three new species of Coffea L. (Rubiaceae) from NE Madagascar. Adansonia 23: 137–146. Davis, A.P. & Rakotonasolo, F. 2003. New species of Coffea L. (Rubiaceae) from Madagascar. Bot. J. Linn. Soc. 142: 111–118. Guillaumet, J.L. & Hallé, F. 1978. Echantillonnage du matériel récolté en Ethiopie. Bull. Inst. Franc. Café Cacao 14: 13–18. Krug, C.A. & Carvalho, A. 1951. The genetics of Coffea. Advances Genet. 4: 127–158. Lashermes, P., Combes, M.C., Robert, J., Trouslot, P., D’Hont, A., Anthony, F. & Charrier, A. 1999. Molecular characterisation and origin of the Coffea arabica L. genome. Molec. Gen. Genet. 261: 259–266. Lashermes, P., Combes, M.C., Trouslot, P. & Charrier, A. 1997. Phylogenetic relationships of coffee tree species (Coffea L.) as inferred from ITS sequences of nuclear ribosomal DNA. Theor. Appl. Genet. 94: 947–955. Lashermes, P., Cros, J., Combes, M.C., Trouslot, P., Anthony, F., Hamon, S. & Charrier, A. 1996. Inheritance and restriction fragment length polymorphism of chloroplast DNA in the genus Coffea L. Theor. Appl. Genet. 93: 626–632. Lebrun, J. 1941. Recherches morphologiques et systématiques sur les caféiers du Congo. Publ. Inst. Natl. Etude Agron. Congo Belge, Ser. Sci. 11: 1–186. Le Pierrès, D., Charmetant, P., Yapo, A., Leroy, T., Couturon, E., Bontems, S. & Tehe, H. 1989. Les caféiers sauvages de Côte d’Ivoire et de Guinée: bilan des missions de prospection effectuées de 1984 à 1987. Pp. 420–428 in: Proceedings of 13th International Scientific Colloquium on Coffee. Association Scientifique Internationale du Café, Lausanne. Louarn J. 1992. La fertilité des hybrides interspécifiques et les relations génomiques entre caféiers diploïdes d’origine africaine (genre Coffea sous-genre Coffea). Doctoral thesis, Université Paris-Sud 11, Orsay, France. Maurin, O., Davis, A.P., Chester, M., Mvungi, E.F., Jaufeerally-Fakim, Y. & Fay, M.F. 2007. Towards a phylogeny for Coffea (Rubiaceae): identifying well-supported lineages based on nuclear and plastid DNA sequences. Ann. Bot. 100: 1565–1583. 140 TAXON 58 (1) • February 2009: 133–140 Namur, C. de, Couturon, E., Sita, P. & Anthony, F. 1987. Résultats d’une mission de prospection des caféiers sauvages du Congo. Pp. 397–404 in: Proceedings of 11th International Scientific Colloquium on Coffee. Association Scientifique Internationale du Café, Lausanne. N’Diaye, A., Poncet, V., Louarn, J., Hamon, S. & Noirot, M. 2005. Genetic differentiation between C. liberica var. liberica and C. liberica var. Dewevrei and comparison with C. canephora. Pl. Syst. Evol. 253: 95–104. Noirot, M., Poncet, V., Barre, P., Hamon, P., Hamon, S. & Kochko, A. de. 2003. Genome size variations in diploid African Coffea species. Ann. Bot. 92: 709–714. Poncet, V., Hamon, P., Minier, J., Carasco, C., Hamon, S. & Noirot, M. 2004. SSR cross-amplification and variation within coffee trees (Coffea spp.). Genome 47: 1071–1081. Poncet, V., Hamon, P., Sauvage de Saint Marc, M.B., Bernard, T., Hamon, S. & Noirot, M. 2005. Base composition of Coffea AFLP sequences and their conservation within the genus. J. Heredity 96: 1–7. Sonké, B., Ngeumba, C.K. & Davis, A.P. 2006. A new dwarf Coffea (Rubiaceae) from southern Cameroon. Bot. J. Linn. Soc. 151: 425–430. Sonké, B. & Stoffelen, P. 2004. Une nouvelle espèce de Coffea L. (Rubiaceae) du Cameroon, avec quelques notes sur ses affinités avec les espèces voisines. Adansonia 26: 153–160. Stoffelen, P. 1998. Coffea and Psilanthus (Rubiaceae) in Tropical Africa: A Systematic and Palynological Study, Including a Revision of the West and Central African Species. PhD-thesis, Catholic University Leuven, Leuven, Belgium. Stoffelen, P., Cheek, M., Bridson, D. & Robbrecht, E. 1997a. A new species of Coffea and notes on mount Kupe. Kew Bull. 52: 989–994. Stoffelen, P., Noirot, M., Couturon, E. & Anthony, F. 2008. A new caffeine-free coffee species of Cameroon. Bot. J. Linn. Soc. 158: 67–72. Stoffelen, P., Robbrecht, E. & Smets, E. 1996. Coffea (Rubiaceae) in Cameroon: a new species and a nomen recognized as species. Belg. J. Bot. 129: 71–76. Stoffelen, P., Robbrecht, E. & Smets, E. 1997b. Adapted to the rain forest floor: a remarkable new dwarf Coffea (Rubiaceae) from lower Guinea (tropical Africa). Taxon 46: 37–47. Stoffelen, P., Robbrecht, E. & Smets, E. 1999. A new species of Coffea (Rubiaceae) from central Africa, with notes on tentative other taxa. Syst. Geogr. Pl. 69: 119–124. Van der Vossen, H.A.M. 1985. Coffee selection and breeding. Pp. 48–96 in: Clifford, M.N. & Willson, K.C. (eds.), Coffee Botany, Biochemistry and Production of Beans and Beverage. Croom Helm, London.