Floral scent emitted by white and coloured morphs in orchids.

Polymorphism of floral signals, such as colour and odour, is widespread in flowering plants and often considered to be adaptive, reflecting various pollinator preferences for particular floral traits. Several authors have recently hypothesized that particular associations exist between floral colour and scent, which would result from shared biochemistry between these two floral traits. In this study, we compared the chemical composition of floral volatiles emitted by white- and purple-flowered morphs of three different orchid species, including two food-deceptive species (Orchis mascula and Orchis simia) and a food-rewarding species (Anacamptis coriophora fragrans). We found clear interspecific differences in floral odours. As expected from their pollination strategy, the two deceptive orchids showed high inter-individual variation of floral volatiles, whereas the food-rewarding A. c. fragrans showed low variation of floral scent. Floral volatiles did not differ overall between white- and coloured-flowered morphs in O. mascula and A. c. fragrans, while O. simia exhibited different volatile profiles between the two colour morphs. However, a detailed analysis restricted to benzenoid compounds (which are associated with the production of floral anthocyanin pigments) showed that white inflorescences emitted more volatiles of the shikimic pathway than coloured ones, both for O. mascula and O. simia. These results are consistent with the current hypothesis that shared biochemistry creates pleiotropic links between floral colour and scent. Whether intraspecific variation of floral signals actually affects pollinator attraction and influences the reproductive success of these orchids remains to be determined.

Olfactory and behavioural responses of tabanid horseflies to octenol, phenols and aged horse urine.

Electrophysiological and behavioural responses of females of two tabanid species, Tabanus bromius L. and Atylotus quadrifarius (Loew) (Diptera: Tabanidae), to ammonia, octenol (1-octen-3-ol), phenols and aged horse urine were compared. Electroantennogram (EAG) responses in both species to octenol, 4-methylphenol (4MP), 3-propylphenol (3PP) and a phenol mixture (4MP and 3PP at a ratio of 16 : 1) increased in a dose-dependent fashion. The most effective stimulus was 4MP and synergism between the two phenols may exist. Aged horse urine also elicited strong EAG responses in both species. Using gas chromatography-mass spectrometry (GC-MS) analysis, we identified 29 compounds in horse urine, which included, in particular, ketones, fatty alcohols and phenols, among which 4MP was the most abundant component (~ 80%). Trapping experiments were carried out using Nzi traps baited with various odours. Octenol and the phenol mixture in combination with ammonia increased catches of tabanids by 1.8-2.8 times relative to ammonia alone. Aged horse urine increased catches of T. bromius and A. quadrifarius by 2.2 and 4.1 times, respectively. The high attractiveness of aged horse urine, especially for A. quadrifarius, is not likely to derive from 4MP alone, but from the mixture of various active compounds used in host location.

Behavioural and electrophysiological responses of females of two species of tabanid to volatiles in urine of different mammals.

Urine volatiles from different ungulates (cows, horses and sheep) were tested as bait for tabanids in southeastern France using Nzi traps during the early summer of 2011. Tabanus bromius Linnaeus, 1758 and Atylotus quadrifarius (Loew, 1874) (both: Diptera: Tabanidae) were the most captured species, respectively representing 57% and 41% of all tabanids collected (all of which were female). Horse urine significantly increased catches of T. bromius (1.6-fold) and A. quadrifarius (3.5-fold), and sheep urine significantly increased catches of A. quadrifarius (2.5-fold). In parallel, an electroantennogram (EAG) study was conducted for the first time on these two species, in which EAGs were recorded using 1-octen-3-ol and extracts of the same urine samples used in the field. For T. bromius, the EAG response to 1-octen-3-ol increased quasi-sigmoidally with dose, with a maximum response at ≥100 µg on filter paper. For both species of tabanid, cow and horse urine elicited larger EAGs than did sheep urine. The behavioural implications in host-seeking and feeding habits are discussed.

Rare white-flowered morphs increase the reproductive success of common purple morphs in a food-deceptive orchid.

How floral colour polymorphism can be maintained in evolutionary time is still debated. In rewardless orchids, it is unknown whether rare white-flowered morphs differ in scent chemistry from pigmented morphs, and whether such intraspecific variation in floral signals may have an impact on reproductive success. We compared the chemical composition of floral volatiles emitted by white- and purple-flowered morphs of Orchis mascula, and recorded the fruit set of both colour morphs. We also used white ping-pong balls to mimic white-flowered morphs in field bioassays. We found that colour polymorphism was not associated with floral odour polymorphism. Surprisingly, when populations of purple-flowered plants included a few white-flowered individuals, the fruit set of the purple morph increased significantly (from 6 to 27%), while that of the white morph remained low. We obtained the same fourfold increase in fruit set when using ping-pong balls as visual lures, demonstrating the association between colour variation and fruit set, and the key role of visual signals in pollinator attraction. Our results are incompatible with negative frequency-dependent selection, a hypothesis invoked to explain colour polymorphism in other rewardless orchids. We propose several hypotheses to explain the maintenance of white morphs in O. mascula.