A Brief Evolutionary Excursion Comes to an End: the Genetic Relationship of British Species of Gentianella Sect
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Plant Syst. Evol. 237: 137–151 (2003) DOI 10.1007/s00606-002-0248-3 A brief evolutionary excursion comes to an end: the genetic relationship of British species of Gentianella sect. Gentianella (Gentianaceae) M. O. Winfield1, P. J. Wilson2, M. Labra3, and J. S. Parker1 1Cory Laboratory, Department of Plant Sciences, University of Cambridge, Cambridge, UK 2Wessex Environmental Associates, Redlynch, Salisbury, UK 3Dipartimento di Biologia, Sezione di Botanica Generale, Universita` di Milano, Milano, Italy Received May 20, 2002; accepted October 28, 2002 Published online: March 31, 2003 Ó Springer-Verlag 2003 Abstract. There is much taxonomic confusion in species were genetically more similar to each other the genus Gentianella section Gentianella, five than they were to individuals of the same species species of which are found in Britain. Gentianella from other populations. It is proposed that G. anglica is a British endemic restricted in its distri- anglica may be an early flowering form of G. ama- bution to the chalk downland of southern England. rella that has been created and maintained as a It is considered to be threatened because of habitat consequence of former grassland management loss, and due to hybridisation with the closely practices. related, widely distributed G. amarella. The Red Data Book species, G. uliginosa, shares morpho- Key words: AFLP, conservation, extinction, logical similarities with the other two species but genetic diversity, Gentianella anglica, Gentianella has a very restricted distribution. Individual plants amarella, Gentianella uliginosa, hybridisation, from populations across the range of these three universal cpSSRs. species were analysed using AFLPs and universal cpSSRs to determine the degree of genetic variabil- Five species of the genus Gentianella section ity within and between them, and to look for Gentianella are recorded in the British Isles. evidence of hybridisation. Samples of the two other According to the Flora Europaea, these species Gentianella section Gentianella species found in fall into three main groups that are essentially Britain, G. campestris and G. germanica, were also well defined morphologically and create few included in the study. The analysis of chloroplast taxonomic problems: the amarella group con- SSRs was not informative for G. anglica, G. ama- tains G. amarella (L.) Bo¨rner, G. anglica rella and G. uliginosa, while G. campestris and G. (Pugsley) E. F. Warb. and G. uliginosa (Willd.) germanica differed from each other and the other Bo¨rner while the other two species, G. campes- three species at a single locus. Principal co-ordinate analysis of the AFLP data revealed only three tris (L.) Bo¨rner and G. germanica (Willd.) distinct groups: one group contained G. campestris Bo¨rner give names to their own groups (Tutin samples, another contained all samples of G. ger- et al. 1972). However, in general, the taxonomy manica, and a third contained all individuals from of section Gentianella is difficult and confused the other three species. In mixed populations of G. in Britain and in the rest of Europe. Problems anglica and G. amarella, individuals of the two arise principally from phenology; variation in 138 M. O. Winfield et al.: Genetic structure of Gentianella spp. in Britain plant size and flowering time may be correlated develops in the absence of regular grazing and with germination time (autumn, winter or cutting: it requires the sward to be kept to a spring). It is also proposed that hybridisation height of about 2–3 cm (Wilson 1999). and backcrossing between species occur fre- G. amarella, the autumn gentian, is a quently so that hybrid swarms are presumed circumpolar species with its centre of distribu- to result. Finally, the characters which differ- tion in southern Europe. It has a wide distri- entiate species and subspecies are mainly bution in Britain and is common and quantitative rather than qualitative, so that widespread in calcareous pastures and on differences such as node number and pedicel dunes; it is found on both limestone and chalk. proportions underlie identification. It is unclear With the exception of a small number of how much of the variation detected in natural Cornish sites, all populations of G. anglica populations of these species is environmental in occur nearby or intermixed with populations origin and how much is due to genetic differ- of G. amarella (Rich et al. 1997). However, the ences since cultivation from seed under con- two species are thought to be completely trolled conditions has proved problematic. sexually isolated because of their distinct and However, it is clear that these Gentianella non-overlapping flowering times; G. anglica species show considerable phenotypic plasticity flowers from May to June, while G. amarella which adds to the taxonomic confusion. flowers from July to October. In recent years, The early gentian, Gentianella anglica, one however, there have been reports of popula- of Britain’s few endemic species, is included on tions containing hybrids (Margetts and David the UK Government Biodiversity Action Plan 1981, Rich et al. 1997). Indeed, G. anglica list of priority species and receives protection subsp. cornubiensis N. M. Pritch, which until under Schedule 8 of the Wildlife and Coun- 1996 was considered a distinct taxonomic unit, tryside Act 1981. It is also protected under is now thought to be the hybrid G. angli- Appendix I of the Bern Convention and ca · G. amarella (G. · davidiana). This has led Annexes IIb and IVb of the European Com- to the additional concern that G. anglica might munity Habitats and Species Directive (Wigg- become extinct by hybridisation with its more ington 1999). This small annual is found abundant relative (Rich et al. 1997). exclusively in closely grazed, calcareous grass- The dune gentian, G. uliginosa, is a species lands and is principally restricted to the chalk with a very restricted range in Britain, being downlands of south eastern and southern found principally at four or five sites in Wales England. It is preferentially found on south where it occurs in dune slacks or in wet and west facing slopes. The species is consid- hollows (Wigginton 1999). There has also been ered of conservation importance because much report that it has been found on Colonsay in of its natural habitat, nutrient poor calcareous Scotland (Gulliver 1998). It is a Red Data grassland, is being destroyed or altered, and in Book species that is considered vulnerable in many places the species has suffered a sharp both Britain and Europe (Holyoak 1999, decline (Darby 2000, Davis and Duckworth Wigginton 1999). The species, which flowers 2000). It is being lost from many of its former from May to October, is always found in close sites following the ploughing of downland, and proximity to populations of G. amarella. the application of nitrogen fertilisers and In order to understand genetic differentia- herbicides to increase productivity – atmo- tion within Gentianella sect. Gentianella in spheric pollution has aggravated this problem Britain, and more specifically the status of because it results in rainfall with a high the three species belonging to the amarella nitrogen content (Bobbink 1998). At other group, a molecular analysis, using AFLPs and sites, G. anglica has been lost as a result of universal cpSSRs, to determine the level of changes in grassland management, since it DNA variation was carried out. This paper cannot survive in tall, dense vegetation that describes research that was aimed to resolve M. O. Winfield et al.: Genetic structure of Gentianella spp. in Britain 139 more clearly the degree of genetic differentia- from plants that were in flower (i.e., late May– tion between the three species G. anglica, June) and from sites where there had been no G. amarella and G. uliginosa, and to look for previous reports of hybridisation with G. amarella. evidence of hybridisation between them that A similar rationale was used for the collection of could be threatening their integrity. We pre- G. amarella samples; collections were made in late sent results obtained from a study of 115 August/September from six populations. Samples were also collected from four hybrid populations; individual plants from 18 separate populations i.e., populations in which G. anglica, G. amarella representing the range of the three species in and their hybrid, G. · davidiana, had been report- Britain. In addition, samples of G. campestris ed. From two of these populations, Eastbourne and G. germanica were included in order to (population 13, Fig. 1) and Stackpole Warren provide outgroups against which to compare (population 16, Fig. 1), samples were collected genetic variability in the other three species. early in the year as representative of G. anglica. From the other two hybrid populations, Bindon Materials and methods Hill (population 14, Fig. 1) and Gear Sands (population 15, Fig. 1), samples were collected at Plant material. Samples of the five species used in three separate times of year (four individuals from the study were collected from a total of 134 each site at each visit): the first collection was made individual plants representing 23 populations to correspond with the flowering time of G. anglica (Fig. 1). Samples of pure G. anglica were collected (late May/early June); the third collection was from six populations covering the range of the made to correspond with that of G. amarella (late species in the U.K. (Fig. 1). Leaves were collected August/early September); the second collection was G. uliginosa 17. Whitford (5) 18. Oxwich (5) G. campestris 19. Holmsley airfield (5) 20. Kit Hill (3) 21. Colonsay (3) G. germanica 12 22. Sands Bank (3) G. anglica 21 23. Turville (5) 1. Elsea Pit (10) 2. Ivinghoe (5) 7 3. Afton Down (5) 4. Winklebury Hill (5) 5. Prescombe Down (5) 6. Aucombe Bottom (9) G. amarella 7. Cow Green Reservoir (5) 1 8. Therfield Heath (7) 11 9.