Hedychiumviridibracteatum X.Hu, a new species from Guangxi Autonomous Region, South China

Abstract Hedychiumviridibracteatum X.Hu, a new species from Guangxi Zhuang Autonomous Region, South China, is described and illustrated. Hedychiumviridibracteatum X.Hu is included in the short-anther group of Hedychium and is most similar to HedychiumvillosumWall.var.tenuiflorum Voigt ex Baker, H.villosumWall.var.villosum Wall., and H.chingmeianum N. Odyuo & D. K. Roy.


Introduction
Hedychium J. Koenig, 1783 (Zingiberaceae, Zingibereae) is a genus with about 50-80 species mainly distributed in tropical Asia (Jain andPrakash 1995, Wu andLarsen 2000). With pleasant fragrances and beautiful flower forms, Hedychium species are used as cut flowers or to decorate landscapes.
Twelve species of Hedychium have been reported in Guangxi Zhuang Autonomous Region, South China (Wu and Larsen 2000). According to Wu and Larsen (2000) Hedychium villosum, has two varieties, H. villosum Wall. with large flowers and H. villosum var. tenuiflorum Wall. ex Baker with small flowers. Yu et al. (2010) proposed that the two varieties should be recognised as distinct species on the basis of their discernible morphological characteristics, different polyploid levels and flowering time, which contribute to complete reproductive isolation. Sanoj et al. (2013), having compared the original specimens and descriptions, concluded that the plant with small flowers was the true H. villosum Wall. var. villosum Wall., and the one with large flowers was H. villosum Wall. var. tenuiflorum Voigt ex Baker. Concurring with Yu et al. (2010), Gao et al. (2014) referred to H. villosum var. tenuiflorum Wall. ex Baker (Wu and Larsen 2000) as an independent species, H. tenuiflorum (Baker) K. Schum. In this paper, the treatment of H. villosum Wall. and its related species follows Sanoj et al. (2013).
In the autumn of 2008 we received a flowering plant of Hedychium from a local person in Napo, Guangxi Autonomous Region, South China. The plant resembled H. villosum Wall. var. tenuiflorum (Wall. ex Voigt) Wall. ex Baker (Sanoj et al. 2013) in having long filaments and sagittate anthers but could be distinguished from the above by its pure white flower, broad leaves, short ligule, short green bracts and bracteoles. After a further field investigation, morphological studies, flow cytometric analysis of nuclear DNA content, and comparison with published protologues and descriptions (Baker 1890(Baker -1892Schumann 1904;Schilling 1982;Ridley 1898;Larsen 1965;Jain and Prakash 1995;Wu and Larsen 2000;Pornpimon 2008;Yu et al. 2010;Sanoj et al. 2013;Odyuo and Roy 2017), the plant in question was confirmed to be a new species of Hedychium.

Morphological comparison and distribution mapping
In October 2011 we carried out a field survey and found the new plant in the wild in the vicinity of the Nongyi village, Napo County, Guangxi Zhuang Autonomous Region. The new plant grows on steep rocks under the forest canopy, at about 600 m a.s.l. Before that field trip, we had collected Hedychium villosum Wall. in the field from different part of China between 2006 and 2010. Based on those field surveys and collections, the affinities between the new species and its relatives, H. villosum Wall., were compared to living plants and specimens. Guided by the collection site of several suspected specimens, we made further field investigations after 2012 to gain a better understanding of the distribution range of the new plant. By combining the geographical distribution data collected from field investigations, specimens (GH, GXMI, HITBC, IBK, IBSC, KUN, PE) and references (Yu et al. 2010), we made a distribution map of the new plant and the related species using DIVA-GIS (version 7.5) (http://www.diva-gis.org/).

Ploidy level analysis of the new plant and its related species
Flow cytometric analysis of nuclear DNA content of the new species and its relatives was performed according to Sakhanokho et al. (2009). Ploidy levels were determined for young leaf tissues. A piece of leaf tissue of about 1 cm 2 was chopped with a sharp razor blade in a petri dish containing 0.4 ml nuclei extraction buffer. After a 3 min incubation with gentle agitation, the extract was poured through a 50 µm mesh sieve. DNA fluorochrome, a nucleus staining buffer, was added to the extract buffer in the ratio of 4:1 and the sample was analysed immediately for the DNA content of the nuclei. Buffers were supplied as part of the Cystain ultraviolet Precise T reagent kit (Partec GmbH, Münster, Germany). The fluorescence of the nuclei was measured using a Partec Cy Flow Space flow cytometer (Partec GmbH, Münster Germany). Sample measurements were replicated three times for each plant. The results were displayed as histograms showing the number of nuclei grouped in peaks of relative fluorescence intensity, which is proportional to the DNA content. To determine the standard peak of diploid cells (2C DNA), leaf tissues were collected from young leaves of diploid Hedychium coronarium plants (Hu et al. 2011;Ramachandran 1969). The instrument gain was adjusted so that the peak of nuclei isolated from diploid (H. coronarium) was set at channel 50 and this calibration was checked periodically to minimise variation resulting from runs. Therefore, peaks representing nuclei from samples with diploid and tetraploid levels were expected at channels 50 and 100, respectively.
Flowering. September-October. Fruiting. Unknown. Etymology. The new species is named after its green bracts and pure white flowers which are highly diagnostic.
Habitat. This species is currently found on limestone rocks in Guangxi Autonomous Region (Napo, Longzhou and Jingxi Counties) mainly growing under forest at altitudes of 600-800 m.
Ploidy level analysis. The results (Fig. 4) show that, when the nuclei of the diploid control (Fig. 4A), Hedychium coronarium (Hu et al. 2011;Ramachandran 1969) were set to channel 50, the new plant, H. viridibracteatum X.Hu (Fig. 4B) resolved at channel 100. This means that the new plant is a tetraploid which shares the same level of ploidy with H. villosum Wall. var. tenuiflorum Voigt ex Baker (Fig. 4C) but differs from H. villosum var. villosum Wall. (Fig. 4D).  Distribution. In addition to the type location, the new species is found in the adjacent area, Longzhou and Jingxi Counties (Fig. 5). Hedychium viridibracteatum can be easily distinguished from related species, even from dried specimens. The following sites were discov-   Although Hedychium viridibracteatum and H. villosum var. tenuiflorum have the same ploidy level the two species differ in their flowering time which ensures that they are reproductively isolated. Hedychium viridibracteatum shares the same flowering time with H. villosum var. villosum, but they can be distinguished by their different flower form, flower size (Table 1), ploidy level (Fig. 4), and are geographically isolated (Fig. 5). The new species is also similar to the newly published H. chingmeianum N. Odyuo &D. K. Roy, 2017 (Odyuo andRoy 2017) but can be distinguished by having green bract (vs. purplish-brown), larger flowers (10.0-11.0 cm long vs. 4-5 cm long), a longer filament (6.0-6.8 cm vs. 2.8-3.0 cm) with white colour (vs. brownish-red). Thus, we confirm that H. viridibracteatum is a new species.