Botanical Journal of the Linnean Society, 2008, 156, 491–499. With 2 figures Pollination of the lady’s slipper Cypripedium henryi Rolfe (Orchidaceae) PENG LI2,3, YI-BO LUO1,2*, YIN-XIA DENG2,4 and YONG KOU5 1The National Orchid Conservation Centre, Shenzhen 518114, Guangdong, China 2State Key Laboratory of Systematic and Evolutionary Botany, Institute of Botany, Chinese Academy of Sciences, Beijing 100093, China 3Graduate School of the Chinese Academy of Sciences, Beijing 100049, China 4Jilin Agriculture University, Changchun 130118, Jilin, China 5Huanglong Administration of National Scenic Spots, Huanglong 623300, Sichuan, China Received 2 November 2005; accepted for publication 22 October 2007 The pollination ecology of Cypripedium henryi Rolfe, a slipper orchid endemic to west China, was investigated, and its floral shape, size, colour, and scent were analysed. Examination of the breeding system suggests that the flowers are self-compatible, but need pollen vectors for successful reproduction. The flower is rewardless; over 15 insects belonging to Araneida, Hymenoptera, Diptera, Lepidoptera, and Coleoptera were recorded as flower visitors, but most only alighted or rested on the flower. In the total 32 h of observations over 2 years, female Lasioglossum bees were found to be the most frequent visitors and the only pollinators. They showed a high visitation frequency and, surprisingly, re-visited the same flowers frequently. Cypripedium henryi probably attracts pollinators visiting the flowers through general food deception (odour components, colour, false nectar guides), as well as special structures (slippery labellum, slippery staminode). Although three Lasioglossum species visited the flowers, only L. sauterum Fan et Ebmer was found with pollen. Lasioglossum flavohirtum Ebmer was large and climbed out from the entrance. Morphologically, L. sichuanense Fan et Ebmer could be considered as a potential pollinator, but the collected specimens were found to have no pollen of C. henryi on their bodies. It was speculated that the particular floral scent of C. henryi discouraged the entrance of L. sichuanense bees. Lasioglossum sauterum was matched morphologically to the flower, but not all of the visitations resulted in effective pollinations, as some flowers of C. henryi were frequently re-visited and the pollen mass had been taken away by bees on previous visitations. © 2008 The Linnean Society of London, Botanical Journal of the Linnean Society, 2008, 156, 491–499. ADDITIONAL KEYWORDS: breeding system – deceptive pollination – floral functional morphology – Lasioglossum – re-visitation. INTRODUCTION opening at the base of the labellum. When the polli- nator exits through a basal orifice, it first passes the The lady’s slippers (Cypripedioideae) are well-known stigma and picks up a portion of a pollinium. Thus, orchids of the north temperate and certain tropical one pollination case is completed if the pollinator has zones (Dressler, 1993). This subfamily comprises five already been smeared with pollen during a previous genera with 158 species and has flowers that do not visitation. Although all the slipper orchids have this offer a reward to pollinators (Pridgeon et al., 1999). fixed pollination route, their pollinators and pollina- The slipper orchid flower is a one-way trap flower, and tion mechanisms show a high degree of diversity the pollinator enters into the labellum from the front (Stoutamire, 1967; Nilsson, 1979; Sugiura et al., 2001; entrance, and exits through one of the two basal Bänziger, Sun & Luo, 2005). orifices that are formed by the staminode blocking the One group of lady’s slippers is Cypripedium,a genus of about 45 species widespread in the north *Corresponding author. E-mail: [email protected] temperate zones (Cribb, 1997). Mainly based on the © 2008 The Linnean Society of London, Botanical Journal of the Linnean Society, 2008, 156, 491–499 491 492 P. LI ET AL. labellum morphology, five distinctive types can be Huanglong Nature Reserve, Sichuan, China. The alti- recognized in the genus: ‘arietinum’, ‘guttatum’, tude of the locality is 2000 m and C. henryi grows ‘calceolus’, ‘japonicum’, and ‘margaritaceum’ (Cribb, along a path under sparse low deciduous trees or 1997). To date, most Cypripedium species have been amongst grasses. The main co-blooming species found to be pollinated by various bees (Cribb, 1997; are Rhododendron sp., Fragaria orientalis A. Los., Van der Cingel, 2001), and fly pollination has only Ranunculus sp., Taraxacum sp., and Lysimachia sp.. been reported in C. reginae Walt (Vogt, 1990) and A few other orchids, C. plectrochilum, C. tibeticum hypothesized in C. debile Rchb., C. margaritaceum King ex Rolfe, C. sichuanense Perner, C. fasciolatum Franch., C. lichiangense Cribb & S.C. Chen, and their Franch., and Calanthe tricarinata Lindl., are also allies (Cribb, 1997). The path of the pollinators found in this community, but only C. plectrochilum through the slipper-shaped flower is probably con- was flowering during our study. trolled by a number of features of floral morphology Cypripedium henryi is 30–60 cm tall with one to (for example, inflexed margins, lines of hairs leading four flowers (usually two or three). The pedicel and to the rear exits, false nectar guides, etc.) (Catling & ovary length is 30–60 cm with dense glandular hairs. Catling, 1991). Stoutamire (1967) suggested that the The flowers are entirely greenish yellow or green with sizes of the labellum mouth and basal orifices and of a glossy labellum. The dorsal sepal is similar to the the escape route under the stigma and anthers deter- united lateral sepals, 3–4.5 cm long and 1–1.5 cm mine the possible pollinators. However, it seems that wide. The petals are 3–5 cm long, 1–2 cm wide, and a similar floral morphology does not always mean not twisted. The labellum is ellipsoidal, 1.5–2.5 cm similar pollinators. For example, the European C. long, 1–2 cm wide, and 1–1.5 cm deep, with hairs and calceolus L. and the North American C. parviflorum red veins in the bottom. The staminode is oblong, Salisb. are similar in floral morphology and only differ 6–7 mm long, 3–4 mm wide, and with or without red with regard to staminode shape and colour (Cribb, spots on the surface (Fig. 1A). The stigma is oblong, 1997), but have different pollinators. The most fre- and its surface is retuse groove with papillose quent and regular vectors of the former are female (Fig. 1C). The pollen is aggregated into two yellow Andrena haemorrhoa (F.) (Nilsson, 1979), whereas or yellow–green masses of sticky, friable grains those of the latter are male Ceratina bees (Fig. 1C). (Stoutamire, 1967). By contrast, the East Asian C. plectrochilum Franch. and the North American C. arietinum R. Br. are a vicarious species pair with METHODS similar floral morphology, and are distinguished from FLORAL VISITORS AND THEIR BEHAVIOUR each other by the features of the dorsal sepal and Pollination observations of C. henryi were made for a staminode (Chen, 1983). Both have similar pollina- total of 32 h over 15 days. The behaviour of visitors tors: species of Lasioglossum (Stoutamire, 1967; Li was recorded from the moment when they entered et al., in press). the vicinity of the flowers to the moment when they The species C. henryi Rolfe is endemic to west left. Every insect activity, including approaching, China. It resembles superficially C. calceolus, but alighting, entering, creeping, and climbing, was differs by its usually two- or three-flowered inflores- recorded with a tape recorder. Some of the visiting cence, smaller green or yellow–green flowers, small insects were impossible to identify to the species’ glossy labellum, and anther filaments that are not level with certainty in the field. In such cases, covered by the staminode (Cribb, 1997). It is hard samples were collected in order to cover the diversity to predict whether C. henryi will have pollinators of the visiting insects. In practice, insects were often similar to or different from those of C. calceolus on the only identified to a particular group of species or basis of the floral morphological differences. In this type in the field. For instance, ‘Lasioglossum type’ study, we investigated the pollination ecology of means the bees agreed in all perceptible characters C. henryi to determine whether or not it has similar with this genus. pollinators to C. calceolus. In addition, we investi- gated the pollination mechanism of C. henryi through an analysis of its floral shape, size, colour, and scent. FLORAL FUNCTIONAL MORPHOLOGY AND INSECT MEASUREMENT To assess the relationship between floral structure MATERIAL and pollinators, 20 flowers were randomly chosen, This study was carried out during the anthesis of and flower traits, especially those which are consid- C. henryi from May 15th to 25th, 2004 and from May ered to be related to the success of pollination, includ- 13th to 25th, 2005. A population of about 100 indi- ing the entrance diameter of the labellum (ML), the viduals of C. henryi was found in Danyun Gorge, height between the stigma and the bottom of the © 2008 The Linnean Society of London, Botanical Journal of the Linnean Society, 2008, 156, 491–499 POLLINATION OF CYPRIPEDIUM HENRYI (ORCHIDACEAE) 493 Figure 1. Flowers of Cypripedium henryi and its pollinators. All photographs were taken under natural conditions. Scale bar, 1 cm. A, Close view of an individual flower of C. henryi. B, Longitudinal section of the labellum of C. henryi showing the internal path of the pollinator (AL, height between the anther and the bottom of the labellum; SL, height between the stigma and the bottom of the labellum). C, Stigma and anther of C. henryi.D,Lasioglossum sp. entering the labellum of C. henryi from the entrance. E, Lasioglossum sp. scratching the staminode and sliding into the labellum of C. henryi. F, Lasioglossum sp. in the labellum of C. henryi (ML, mouth diameter of the labellum).