Revision of Eulophia (Orchidaceae) in Nigeria, Cameroon, Equatorial Guinea, Gabon, and the Republic of the Congo

Background and aims – The genus Eulophia (Orchidaceae) is revised for Nigeria, Cameroon, Equatorial Guinea, Gabon, and the Republic of the Congo. The aims are: to present a morphological characterization of the main vegetative and reproductive features of Eulophia to clarify the delimitation of the taxa within the genus in the studied countries; to provide an updated identification key and an updated checklist of the genus with nomenclatural data, distribution maps, ecological information, and preliminary conservation status. This revision may serve as a basis for future studies of the genus in other regions of Africa. Material and methods – Relevant material kept in BM, BR, FHI, HBG, K, MA, P, and WAG was examined, using standard practices of herbarium taxonomy. Vegetative and reproductive structures were analysed. MapMaker was used to produce the distribution maps. Key results – A total of 24 species are recognised in the study area. The variability of perennating organs, leaves, sepals and petals, lip (including ornamentation), spur, and anther cap are described and depicted, and were found to be informative for species recognition. A taxonomic treatment is given with an identification key, synonymy, distribution maps, preliminary conservation status, and specimen citations. Eulophia galeoloides is neotypified, E. brevipetala, E. leonensis, E. penduliflora, and Lissochilus elatus are lectotypified. According to our study, we consider E. parvula a synonym of E. pyrophila. E. sordida is considered as a doubtful species in the study area.


INTRODUCTION
The genus Eulophia R.Br. ex Lindl. (Orchidaceae, Cymbidieae) is the most diverse in the subtribe Eulophiinae (Chase et al. 2015), including about 164 terrestrial species (Martos et al. 2014;Bone et al. 2015). The genus shows a pantropical distribution, mainly in Africa (Central and Southern), Madagascar, and Asia, with seven species in Australasia and five in America (Pridgeon et al. 2009;Bone et al. 2015;Govaerts et al. 2019).
No complete taxonomic revision has ever been attempted for Eulophia. Thomas (1998) provided a preliminary checklist of the genus. Due to its wide distribution, the high degree of morphological interspecific variation in the vegetative and reproductive characters, and the lack of complete preserved material of many taxa in herbaria, misidentifications occurred, and an infrageneric classification is not firmly settled at present (Cieslicka 2006;Pridgeon et al. 2009;Bone et al. 2015). Based on a molecular phylogenetic analysis of the genus, focused principally on South African taxa (Martos et

MATERIAL AND METHODS
The study area comprises Nigeria, Cameroon, Equatorial Guinea (Bioko and Rio Muni), Gabon, and Republic of the Congo. More than 500 specimens deposited at the herbaria K, MA and WAG were revised. Additional specimens were analysed from the digital collections of the herbaria BM (Natural History Museum 2018), BR (BR Herbarium 2018), HBG (Herbarium Hamburgense 2018), P (MNHN 2018), and the JSTOR Global Plants facility (JSTOR 2018) for type materials and specimens from FHI. All specimens cited have been examined unless indicated by "n.v." after the herbarium acronym. Specimens seen only as digital images are indicated by "web". All herbarium acronyms follow Thiers (2019).
Accepted species appear in alphabetical order, and only synonyms with type specimens recorded in countries of the study area are included. We provide type details for each name, and lectotypes or neotypes were designated when necessary.
Vegetative and reproductive characters were analysed for each species ( Caliper Mitutoyo). Unless otherwise stated, the dimensions mentioned for vegetative structures refer to dry material or relevant literature, and the colours to digital images of live specimens and relevant literature. Some characteristics were captured from labels, especially those which are lost in the process of drying and pressing. Terminology was generally adopted from Dressler (1993), Pridgeon et al. (2009), and Beentje (2016), although a detailed description of the main features of the species studied has been provided. Distribution and ecology were obtained from labels of herbarium specimen and bibliographic resources. Distribution maps were produced using the software Map Maker Pro v.3.5 (Map Maker Limited 2019) from georeferenced specimens. For each species, global distribution is provided followed by the country repartition in the study area.
The conservation assessments followed the criteria and categories of the IUCN Red List (IUCN 2001(IUCN , 2019, based on the regional distribution of each taxon.

RESULTS
After a detailed examination of specimens, the study of vegetative and reproductive characters reveals the occurrence of a set of diagnostic features (tables 2, 3), described below and mainly used for the elaboration of the dichotomous key.
Tubers are thickened ovoid to conical subterranean organs, which are covered by the bases of the leaves ( fig. 1C).
Pseudobulbs are aboveground organs ( fig. 1D), ocassionally with the base underground, with whitish roots. From the pseudobulbs arise the normal leaves on the top and the lateral inflorescence with scale-leaves. Leaves -Among the species studied, leaf shape usually varies from linear to lanceolate. The leaves of E. guineensis are highly variable, from oblong, non-petiolate to elliptic, long-petiolate. The only mycoheterotrophic species is E. galeoloides, which lacks functional leaves.
One of the diagnostic features is the presence of mature leaves at anthesis, distinguishing between coetaneous and non-coetaneous species. Only E. cucullata and E. guineensis have both forms.      4C). The spur is usually parallel to the lip lamina or is slightly curved upwards, except in E. brevipetala and E. stachyodes where it is incurved ( fig. 4D). It varies in length, from 1 mm in E. pyrophila up to 28 mm in E. guineensis, which has the longest spur in the genus. The colour is similar to the lip such as in E. bouliawongo, or dissimilar such as in E. euglossa and E. guineensis.  5A). Habitat and ecology -Swampy areas. Elevation 400-500 m. Preliminary IUCN conservation status -EN (Endangered). The number of locations is less than 5 in the area of study. Besides, a considerable period of time has elapsed since 1860 when Charles Barter collected the only two specimens from Nigeria, so we consider that data on abundance and current distribution in Nigeria are lacking, and future prospections will be appropriate. Other collection examined -Nigeria: Sare, C. Barter 3429 (K). (Raynal 1966 : Boutica, 12 Jul. 1902, O. Debeaux 378 (K); Gamba, near Shell terminal, 2º47′S, 10º02′E, 9 Jul. 1992 Notes -The lectotype specimen of Lissochilus elatus Rolfe, consists of three herbarium sheets, one with the inflorescence (K000078544), one with foliar stem and part of the scape with the bracts (K000078545), and the last one with the basal part of the scape (K000078546). Rolfe (Rolfe 1897: 53). Notes - Cribb (1989) and Lebrun & Stork (2015) considered E. brevipetala as conspecific with E. monile Rchb.f., whereas Pérez Vera (2003) considered it a variety of E. monile. However, both taxa differ in several characters, such as the colour of the petals and lip (rose to whitish in E. brevipetala, yellow-green with purple margin in E. monile), shape and length of the petals (oblong-ovovate and 4.4-5 mm long in E. brevipetala, linear and 7-11 mm long in E. monile) and the number of ridges (5 in E. brevipetala, 3 in E. monile).

Eulophia brevipetala
Two sheets with material collected by G.F. Scott Elliot 5224 from Sierra Leone, were deposited in the herbaria K and BM. Since R.A. Rolfe did not designate the holotype, both sheets can be regarded as syntypes and we can choose a lectotype. The designated lectotype (K000410367) consists of a complete specimen, with a detached flower, measurements of floral structures and drawings from microscopic preparations made by V.S. Summerhayes. The specimen from herbarium BM is incomplete, only contains the scape and three flowers.  Lissochilus ledermannii Kraenzl. (Kraenzlin 1912: 396) -Eulophia ecarinata Butzin (Butzin 1975: 588

Eulophia galeoloides
Kraenzl. (Kraenzlin 1898: 508). Type -Tanzania: Usambara, Mar., E. Heinsen 10 (holotype: B †); Tanzania, Lushoto District, Amani, C. Braun 670 (neotype: K[K000410402, http://specimens.kew.org/herbarium/ K000410402, designated here; isoneotype: EA n.v.). Distribution: Mainly distributed in East Tropical Africa, extending from Ghana to the west. Nigeria ( fig. 6A), Cameroon (?). Habitat and ecology -Shady forests. Elevation 0-100 m. Preliminary IUCN conservation status -CR (Critically Endangered). Although this taxon is widely distributed in Africa, it is only known from one location in the area of study. Notes -Kraenzlin (1898: 508) described this species from material collected by E. Heinsen in Usambara, Tanzania. In the herbaria B, BM, BR, and K where Heinsen's collections could be deposited, no material assigned to E. galeoloides could be traced. Cribb (1989) and Geerinck (1992) mentioned a destroyed sheet from B (Braun 670) as holotype, with an isotype deposited in K, which contains material collected in 1905 by Carl Braun from the seedbeds in Amani (in the East Usambara Mountains, Tanzania). The holotype would have been assigned to a specimen collected by E. Heinsen, but this material was destroyed. So, it is necessary to designate a neotype and we have chosen the specimen from the same region, collected by Braun 670 (K000410402). Summerhayes (1968) revised the sheet FHI20295, with specimens collected by S. Tamajong in Etemi, Ijebu Ode (Nigeria), which is represented in the distribution map.
There is a distribution record for Cameroon in Govaerts et al. (2019), but we have not seen specimens or bibliographic references, which could confirm its presence in this country. Notes -Szlachetko & Olszewski (2001: 650) recorded E. elegans Schltr. in Cameroon, based on the sheet P00365660, collected by T.D. Maitland 1753. After studying this specimen, we consider it belongs to E. stachyodes, so without further evidence, we consider E. elegans to be restricted to Tanzania and Malawi.
Notes - Summerhayes (1953) recognised this taxon as distinct to E. pyrophila and only known from Ghana and Togo. Later, Summerhayes (1968) extended its distribution to Nigeria. Cribb (1989) considered E. sordida as synonym of E. pyrophila. Govaerts et al. (2019) recognize the species, indicating the distribution for Nigeria, Cameroon, and Republic of the Congo. According to the original description (Kraenzlin 1902) and the drawings depicted by Szlachetko & Olszewski (2001), the ornamentation of the lip consists of three decreasing ridges, which acquire a more papillose appearance (pearl-like) on the midlobe. We have not observed any specimens with the characteristics mentioned above for E. sordida in the study area.