Org Divers Evol (2014) 14:339–348 DOI 10.1007/s13127-014-0180-8 ORIGINAL ARTICLE Floral scent and its correlation with AFLP data in Sorbus Martin Feulner & Stefan Pointner & Lisa Heuss & Gregor Aas & Juraj Paule & Stefan Dötterl Received: 29 October 2013 /Accepted: 24 June 2014 /Published online: 16 August 2014 # Gesellschaft für Biologische Systematik 2014 Abstract Comparisons between floral scent-based and DNA- aromatic compounds, terpenoids, aliphatics, and nitrogen- molecular-based taxonomies are rare, yet such comparisons containing compounds. The scent patterns were taxon- indicate that scent can provide useful taxonomic information. specific, and the number of scent components differed among Here, we correlate the phytochemical differentiation in floral taxa. The correlations with the published AFLP data on pop- scent to the DNA-molecular-based differentiation in the genus ulation and individual level are highly significant, indicating Sorbus. Inflorescence scent patterns of the apomictic and that the scent and AFLP data are highly congruent in the endemic Sorbus latifolia microspecies Sorbus franconica, plants studied. Scent therefore provides useful taxonomic Sorbus adeana,andSorbus cordigastensis originated by hy- characters in Sorbus. bridization as well as their parental taxa Sorbus aria agg. and Sorbus torminalis were investigated with the dynamic head- Keywords AFLP . Apomixis . Correlation analysis . Floral space method. The scent data (presence/absence of com- scent . Rosaceae . Taxonomy . Sorbus pounds) were used to construct an UPGMA tree, to calculate a similarity matrix, and to correlate them with the published amplified fragment length polymorphism (AFLP) data of the Introduction same individuals, populations, and taxa. Flow cytometry was used to estimate the DNA-ploidy level of the taxa. Scent The main function of floral scent is to attract pollinators analyses showed a total of 68 substances, among them (Plepys et al. 2002; Dötterl et al. 2006), which in turn exert selective pressures on its composition (Knudsen and Tollsten 1993; Plepys et al. 2002; Dötterl et al. 2005). * : : : M. Feulner ( ) S. Pointner L. Heuss S. Dötterl Pollinator-mediated selection may lead to the evolution of Department of Plant Systematics, University of Bayreuth, Universitätsstr. 30, 95440 Bayreuth, Germany pollination syndromes, meaning that plant species pollinat- e-mail: [email protected] ed by the same guild of animals have similar phenotypes S. Dötterl of their floral characteristics, including scent (Faegri and e-mail: [email protected] van der Pijl 1979; Fenster et al. 2004; Dobson et al. 2005). Although scent has been shown to be influenced G. Aas by pollinator-mediated selection and coevolution, most Ecological-Botanical Gardens, University of Bayreuth, Universitätsst. 30, 95447 Bayreuth, Germany studies have shown that only a limited number of sub- e-mail: [email protected] stances have key functions in attracting pollinators (Dötterl et al. 2006;Svenssonetal.2010;Burgeretal.2011), J. Paule whereas the occurrence of other substances may be ex- Department of Botany and Molecular Evolution, Senckenberg Research Institute & Biodiversity and Climate Research Centre plained by phylogeny (Steiner et al. 2011). (BiK-F), Senckenberganlage 25, 60325 Frankfurt/Main, Germany There are some examples showing the floral scent data e-mail: [email protected] support phylogenies that are morphology-based (e.g., Raguso et al. 2003; Feulner et al. 2009, 2011) and DNA-based (e.g., S. Dötterl Department of Organismic Biology, University of Salzburg, Levin et al. 2003). However, to the best of our knowledge, a Hellbrunnerstrasse 34, 5020 Salzburg, Austria statistical approach of the taxonomical value of scent entailing 340 M. Feulner et al. detailed correlations between scent data and data from DNA- S. pannonica with affinity to S. aria s.str. [henceforth affine molecular studies using a Mantel test has so far been reported (aff.) aria] were included in the study (see Feulner et al. 2013). only once: In a study of Ophrys (Orchidaceae), no correlation For all taxa, AFLP data were available from the same popu- was found between the scent data and DNA-molecular data lations. In 13 cases, the AFLP data and scent data were (Stökl et al. 2008). collected from the same individuals (Table 1; Feulner et al. In hybrid complexes such as Citrus, Ophrys,and 2013). For further information about the taxonomy, popula- Hieracium, the scent has been shown to consist mainly of a tion structure, and ecology of the taxa investigated, see mixture of the scent components of the parental species and Feulner et al. (2013). only a low number of novel compounds (Gancel et al. 2002; Vereecken et al. 2010; Feulner et al. 2009, 2011). In such Volatile collection cases, scent analyses are a valuable tool for parental species identification (Feulner et al. 2009, 2011). Inflorescence scent was collected in the field using a standard In the genus Sorbus, many taxa are polyploids and derived dynamic headspace method as described in Feulner et al. by hybridization. Such taxa are typically apomictic in that they (2009). For each taxon, three to eight individuals were sam- produce seeds without the use of meiotic recombination pled. The sampling was carried out on newly opened inflo- (Nogler 1984; Jankun and Kovanda 1987; Campbel and rescences (one inflorescence per plant and sample), between Dickinson 1990; Talent 2009). The pollinator-mediated selec- 11 a.m. and 3 p.m., the period with the most intensive scent tion of scent patterns is of minor importance in apomicts emission (as determined by the human nose; Feulner, unpub- (Feulner et al. 2009, 2011). Furthermore, the genotypic vari- lished data). Scent samples of the leaves and the local sur- ability of apomicts is often low (cf., Talent 2009). Therefore, rounding air were collected as a control for each locality and scent patterns may be identical among the individuals and population investigated. populations of the same apomictic taxon and be of high taxonomic value (Feulner et al. 2009, 2011). Here, we investigate the scent of the apomictic Chemical analysis microspecies Sorbus adeana, Sorbus cordigastensis,and Sorbus franconica, belonging to the Sorbus latifolia aggregate The samples were analyzed on a Varian Saturn 2000 mass (Rosaceae) endemic to Northern Bavaria and occupying very spectrometer and a Varian 3800 gas chromatograph with a small parapatric distribution areas (Meyer et al. 2005; Aas and 1079 injector that had been fitted with a ChromatoProbe kit. Kohles 2011). S. latifolia taxa originated from hybridization This kit allows the thermal desorption of small amounts of between Sorbus aria agg. (including Sorbus pannonica)and solids or liquids contained in quartz microvials (Micro-SPE; Sorbus torminalis (Düll 1961;Richetal.2010;Robertson cf., Amirav and Dagan 1997; Dötterl et al. 2005). The injector et al. 2010; Feulner et al. 2013). split vent was opened (1/20) to flush any air from the system In a former study, the hybrid state, the intraspecific vari- and closed after 2 min; the injector was heated at 40 °C for ability, and the genetic structure of taxa were investigated with 2 min, and the temperature was then increased at a rate of amplified fragment length polymorphism (AFLP) analyses 200 °C min−1 to 200 °C; this end temperature was held (Feulner et al. 2013). In the present study, we investigated constant for 4.2 min, after which the split vent opened (1/10) the floral scent composition of the same type of material used and the injector cooled down. in the AFLP study. We (1) investigated the scent composition A ZB-5 column (5 % phenyl polysiloxane) was used for the of the parental species, (2) examined the distribution of com- analyses (60-m long, inner diameter 0.25 mm, film thickness pounds in the putative hybrid taxa, and (3) tested for congru- 0.25 μm, Phenomenex). Electronic flow control was used to ence of the floral scent-based differentiation with the AFLP- maintain a constant helium carrier gas flow of 1.8 ml min−1. based DNA-molecular differentiation (published in Feulner The GC oven temperature was held constant for 7 min at et al. 2013). 40 °C, then increased by 6 °C min−1 to 250 °C and held constant for 1 min. The MS interface was 260 °C and the ion trap worked at 175 °C. The mass spectra were taken at Material and methods 70 eV (in EI mode) with a scanning speed of 1 scan s−1 from m/z 30 to 350. Study plants The GC-MS data were processed using the Saturn Software package 5.2.1. Component identification was carried out We collected scent from S. adeana, S. cordigastensis,and using the NIST 08 mass spectral database or MassFinder 3, S. franconica as well as the parental taxa S. aria s.str., and confirmed by comparing the retention times and retention S. pannonica,andS. torminalis.Somepreviouslytaxonomi- indices with published data (Adams 2007). The identification cally unclassified intermediates between S. aria s.str. and of the individual components was confirmed by comparing Floral scent and its correlation with AFLP data in Table 1 Taxon name, locality, and voucher information of the individuals analyzed Taxon Locality/Gauss Krüger coordinates/voucher Number of individuals Number of AFLP samples from the Number of samples analyzed for scent sampling same population/ndividual as used by flow cytometry a for scent sampling Sorbus Sorbus aria (L.) Crantz Grafenhäusling/4437746/5542273, Feulner 200–207 (UBT) 2 5/0 2 Autobahn Rossdorf/4437013/5539920, Feulner 208 (UBT) 1 2/0 1 S. aff. aria (intermediates) Autobahn Rossdorf/4437013/5539920, Feulner 209–217 (UBT) 3 4/0 1 Grafenhäusling/4437746/5542273, Feulner 209 (UBT) –– 1 S. pannonica Kárpáti Kordigast/4443782/5551720, Feulner 218–227 (UBT) 4 5/1 6 (offspring) Neudorf/4447194/5546549, Feulner 228–234 (UBT) 1 4/0 Brünnberg/4457310/5520910, Feulner 235 (UBT) 1 – S. adeana N. Mey. Neudorf/4447194/5546549, Feulner 236 (UBT) 4 7/0 2 (offspring) S.