Hannes F. Paulus

Orchid pollination by sexual swindle

The flowers of Ophrys orchids mimic receptive females of usually only one pollinator species. Males of this species are attracted primarily by the odour of a flower and transfer the pollinia during so-called ‘pseudocopulations’ with the flowers. We have found that flowers of O. sphegodes produce the same compounds and in similar relative proportions as are found in the sex pheromone of its pollinator species, the solitary bee Andrena nigroaenea. Common straight-chain saturated and unsaturated hydrocarbons are the key components in this chemical mimicry, which seems to be an economical means of pollinator attraction.

Evolution of reproductive strategies in the sexually deceptive orchid Ophrys sphegodes : How does flower-specific variation of odor signals influence reproductive success?

Abstract The orchid Ophrys sphegodes Miller is pollinated by sexually excited males of the solitary bee Andrena nigroaenea, which are lured to the flowers by visual cues and volatile semiochemicals. In O. sphegodes, visits by pollinators are rare. Because of this low frequency of pollination, one would expect the evolution of strategies that increase the chance that males will visit more than one flower on the same plant; this would increase the number of pollination events on a plant and therefore the number of seeds produced. Using gas chromatography–mass spectrometry (GC-MS) analyses, we identified more than 100 compounds in the odor bouquets of labellum extracts from O. sphegodes; 24 compounds were found to be biologically active in male olfactory receptors based on gas chromatography with electroantennographic detection (GC-EAD). Gas chromatography (GC) analyses of odors from individual flowers showed less intraspecific variation in the odor bouquets of the biologically active compounds as compared to nonactive compounds. This can be explained by a higher selective pressure on the pollinator-attracting communication signal. Furthermore, we found a characteristic variation in the GC-EAD active esters and aldehydes among flowers of different stem positions within an inflorescence and in the n-alkanes and n-alkenes among plants from different populations. In our behavioral field tests, we showed that male bees learn the odor bouquets of individual flowers during mating attempts and recognize them in later encounters. Bees thereby avoid trying to mate with flowers they have visited previously, but do not avoid other flowers either of a different or the same plant. By varying the relative proportions of saturated esters and aldehydes between flowers of different stem positions, we demonstrated that a plant may take advantage of the learning abilities of the pollinators and influence flower visitation behavior. Sixty-seven percent of the males that visited one flower in an inflorescence returned to visit a second flower of the same inflorescence. However, geitonogamy is prevented and the likelihood of cross-fertilization is enhanced by the time required for the pollinium deposited on the pollinator to complete its bending movement, which is necessary for pollination to occur. Cross-fertilization is furthermore enhanced by the high degree of odor variation between plants. This variation minimizes learned avoidance of the flowers and increases the likelihood that a given pollinator would visit several to many different plants within a population. Editor: J. Conner

Pollinator attraction in a sexually deceptive orchid by means of unconventional chemicals

Ophrys flowers mimic virgin females of their pollinators, and thereby attract males for pollination. Stimulated by scent, the males attempt to copulate with flower labella and thereby ensure pollination. Here, we show for the first time, to our knowledge, that pollinator attraction in sexually deceptive orchids may be based on a few specific chemical compounds. Ophrys speculum flowers produce many volatiles, including trace amounts of (ω–1)–hydroxy and (ω–1)–oxo acids, especially 9–hydroxydecanoic acid. These compounds, which are novel in plants, prove to be the major components of the female sex pheromone in the scoliid wasp Campsoscolia ciliata, and stimulate male copulatory behaviour in this pollinator species. The specificity of the signal depends primarily on the structure and enantiomeric composition of the oxygenated acids, which is the same in wasps and in the orchids. The overall composition of the blend differs significantly between the orchid and its pollinator and is of secondary importance. 9–Hydroxydecanoic acid is a rarely occurring compound that until now has been identified only in honeybees. Contrary to the standard hypothesis that Ophrys flowers produce only ‘second–class attractivity compounds’ and are neglected once the pollinator females are present, we show that flowers are more attractive to the males than are their own females.

Variation of Floral Scent Emission and Postpollination Changes in Individual Flowers of Ophrys sphegodes Subsp. sphegodes

We investigated the scent composition of individual flowers of Ophrys sphegodes, its alteration following pollination, and of picked flowers by day and at night. Odor samples were collected by headspace sorption and analyzed by gas chromatography and mass spectrometry. To evaluate the function of postpollination odor changes, we carried out behavioral tests on the pollinator Andrena nigroaenea with pollinated and unpollinated flowers. We identified 27 volatiles in the flower scents. Aldehydes and alkanes were most frequently found. Aldehydes were the most abundant class of compounds (40–50%). When flowers were picked, they emitted significantly lower total amounts of volatiles than unpicked flowers, and their odor bouquets were significantly different. Comparison of scents released by day and at night showed no decrease in scent emission during nighttime, but the odor bouquets were significantly different. Pollinated flowers produced significantly different odor bouquets, and the total amount of scent emitted two to four days after pollination was significantly lower compared with unpollinated flowers. In addition, behavioral tests with A. nigroaenea males showed that flowers were significantly less attractive three days after pollination. This reduced attractiveness is hypothesized to guide pollinators to the unpollinated flowers within an inflorescence, and thus increase the reproductive success of the plant.

Pollinator attracting odour signals in sexually deceptive orchids of the Ophrys fusca group

We investigated patterns of volatiles of several allopatric and sympatric species of the Ophrys fusca group and one species of the O. mammosa/sphegodes group pollinated by either Andrena nigroaenea or A. flavipes, using electrophysiology (gas chromatography coupled with electroantennography; GC-EAD) and chemical analyses. We found 52 GC-EAD active compounds, mainly saturated and unsaturated hydrocarbons with chain lengths of 21 to 31, aldehydes, an ester, and an acid. Based on the relative proportions of all GC-EAD active compounds, the investigated species were compared using various statistical methods (ANOVA, principle component analyses, discriminant function analyses and cluster analyses). Our results show that Ophrys species with the same pollinator – independent of their phylogenetic relationship–use the same volatiles for pollinator attraction. Differences between the species mainly involve different quantitative patterns of volatiles. Our results are in congruence with previous studies that showed different odour bouquets to be responsible for the specific attraction of different pollinators and that alkanes and alkenes are most important for pollinator attraction.

Increase of pollinator attraction by means of a visual signal in the sexually deceptive orchid, Ophrys heldreichii (Orchidaceae)

Orchids of the genus Ophrys are pollinated by males of solitary bees and wasps through sexual deception. Flowers mimic the odor of a receptive female and thus attract males that seek to copulate. Visual stimuli have been assumed so far to play only a minor role in male attraction. We investigated the role of the perigon as a potential visual signal in attracting pollinators in the orchid Ophrys heldreichii and its pollinator, the males of the long-horned bee Tetralonia berlandi (Apidae). In contrast to many other Ophrys species, O. heldreichii exhibits a large and bright pinkish perigon that appears visually conspicuous to a human observer. In a dual choice test we presented two flowers from a single plant and counted visitation rates. We then removed the perigon of one flower and retested the relative attractiveness of both flowers. For 292 male visits in ten trials we found a significant decrease of visitation rate for flowers with the perigon removed. In a second experiment we repeated the dual choice test using photos of the flowers. Males also significantly chose the picture of an intact flower over the picture of a modified flower where the perigon was digitally removed. From our data, we conclude that T. berlandi males respond to and are attracted by the bright pink perigon of the orchid in addition to other stimuli. A bright colorful perigon occurs almost only in the Ophrys holoserica-oestrifera group, a large sub-group of the genus. We hypothesize that this kind of visual signal is adaptive particularly in those Ophrys species where the targeted males patrol resourced-based encounter sites and strongly rely on their visual system while searching for their females.

Ontogenetic shifts in intraguild predation on thrips by phytoseiid mites: the relevance of body size and diet specialization

In greenhouse agroecosystems, a guild of spider mite predators may consist of the oligophagous predatory mite Phytoseiulus persimilis Athias-Henriot, the polyphagous predatory mite Neoseiulus californicus McGregor (both Acari: Phytoseiidae) and the primarily herbivorous but facultatively predatory western flower thrips Frankliniella occidentalis Pergande (Thysanoptera: Thripidae). Diet-specialization and the predator body size relative to prey are crucial factors in predation on F. occidentalis by P. persimilis and N. californicus. Here, it was tested whether the relevance of these factors changes during predator ontogeny. First, the predator (protonymphs and adult females of P. persimilis and N. californicus): prey (F. occidentalis first instars) body size ratios were measured. Second, the aggressiveness of P. persimilis and N. californicus towards F. occidentalis was assessed. Third, survival, development and oviposition of P. persimilis and N. californicus with F. occidentalis prey was determined. The body size ranking was P. persimilis females > N. californicus females > P. persimilis protonymphs > N. californicus protonymphs. Neoseiulus californicus females were the most aggressive predators, followed by highly aggressive N. californicus protonymphs and moderately aggressive P. persimilis protonymphs. Phytoseiulus persimilis females did not attack thrips. Frankliniella occidentalis larvae are an alternative prey for juvenile N. californicus and P. persimilis, enabling them to reach adulthood. Females of N. californicus but not P. persimilis sustained egg production with thrips prey. Within the guild studied here, N. californicus females are the most harmful predators for F. occidentalis larvae, followed by N. californicus and P. persimilis juveniles. Phytoseiulus persimilis females are harmless to F. occidentalis.

Oviposition Behavior of Interacting Predatory Mites: Response to the Presence of Con- and Heterospecific Eggs

Oviposition behavior may be affected by the presence of potential future competitors, mates, or predators of offspring. We examined patch choice, oviposition site preference and egg production in the predaceous mites Phytoseiulus persimilis and Neoseiulus californicus (Acari: Phytoseiidae) when given a choice between paired spider mite patches with and without conspecific eggs, with and without heterospecific eggs, and with conspecific or heterospecific eggs. Neoseiulus californicus females had no patch preference and distributed their eggs randomly in all choice situations. This was also the case with P. persimilis females given a choice between patches with and without conspecific eggs and between patches with either con- or heterospecific eggs. Phytoseiulus persimilis females confronted with patches with and without heterospecific eggs preferentially stayed and oviposited in the predator free patches. We discuss the oviposition strategies of P. persimilis and N. californicus with respect to food competition, cannibalism and intraguild predation.

Reproductive isolation in the Aegean Ophrys omegaifera complex (Orchidaceae)

The orchid genus Ophrys operates a system of sexual deception by which high specificity of pollination is attained. Reproductive isolation in Ophrys mainly rests upon prezygotic isolation mechanisms. The level of genetic separateness of Ophrys taxa with different pollinators is therefore likely determined by the fidelity of pollinators. The present study employs genetic fingerprinting to investigate this in the east Aegean Ophrys omegaifera s.l. complex, also including O. dryis, a west Mediterranean species of this complex. Ophrys fleischmannii, O. basilissa, and the west Mediterranean O. dyris, are found to be well-separated genetic entities whereas O. omegaifera s.str. and the putative hybrid taxon, O. sitiaca, are found to be genetically inseparable across their entire range of co-occurrence. This suggests that specific pollinators have high enough fidelity to act as effective isolating factors in east Aegean O. omegaifera s.l. as a whole, but that the situation in the species pair of O. sitiaca and O. omegaifera is likely to be more complex.

Evidence for progenitor–derivative speciation in sexually deceptive orchids

BACKGROUND AND AIMS Sexually deceptive orchids of the genus Ophrys use mimicry of pollinator females to attract specific pollinators. Pollinator shifts may drive speciation in Ophrys, since novel pollinators may in principle act as isolating factors immediately. It is thus possible that evolution of novel species occurs rapidly and with a progenitor-derivative pattern. The aims of this study are to compare genetic structure and diversity among widespread and geographically restricted Ophrys taxa, to test whether genetic structure is associated with specific pollinators, and to investigate whether any widespread species may have acted as a progenitor for the evolution of more restricted taxa. METHODS Genetic differentiation and diversity were investigated in O. leucadica and O. cinereophila, the two taxa of the Ophrys fusca sensu lato complex widespread in the Aegean, and three geographically restricted taxa from Rhodes, O. attaviria, O. parvula and O. persephonae, all differing in their specific pollinators. This was done using amplified fragment length polymorphism (AFLP) DNA fingerprinting, and sequencing of the low-copy nuclear gene LEAFY (LFY). KEY RESULTS All taxa were found to be separate genetic entities, with O. leucadica forming two geographic groups from the west and east of the Aegean. Genetic structure was significantly shaped by pollinators and geography, and comparison of sequence and AFLP data revealed ancestral polymorphisms shared among several taxa. Among the sampled taxa, O. leucadica harbours the greatest genetic differentiation and geographic structure, and the highest genetic diversity. Part of the genome of O. parvula, endemic to Rhodes, may be derived from O. leucadica. CONCLUSIONS Pollinators probably influence the genetic structure of the investigated Ophrys species. The genetic pattern identified is consistent with O. leucadica being the oldest of the sampled taxa, making O. leucadica a candidate progenitor species from which more restricted taxa such as O. parvula may have evolved.