Mate preferences in <Emphasis Type="Italic">Drosophila</Emphasis> infected with <Emphasis Type="Italic">Wolbachia</Emphasis>?

The maternally inherited bacterium Wolbachia pipientis generates strong reproductive incompatibilities between uninfected females and infected males (cytoplasmic incompatibility), significantly reducing both female and male reproductive success. Such fitness costs are thought to place selective pressure on hosts to evolve pre-copulatory preferences for mating with compatible mates, thereby enabling them to avoid the reproductive incompatibilities associated with Wolbachia. Therefore, uninfected females are predicted to prefer mating with uninfected males, whereas infected males are predicted to prefer mating with infected females. Despite these predictions, previous investigations of pre-copulatory mate preferences in Wolbachia-manipulated Drosophila have not found evidence of female preference for uninfected or compatible males. However, none of these studies utilised a design where focal individuals are provided with a simple choice in a relatively non-competitive situation. We examined both female and male pre-copulatory mate preference based on mate infection status in Drosophila simulans and D. melanogaster using simple choice assays involving between 30–50 replicates per treatment. Although we found no evidence of female pre-copulatory mate preferences in either species, male D. simulans exhibited some preference for mating with females of the same infection status. However, this preference was not evident when we repeated the experiment to confirm this finding. Consequently, we conclude that neither male nor female D. melanogaster and D. simulans exhibit significant Wolbachia-associated pre-copulatory mate preferences.     

Chemical characterization of contact semiochemicals for host-recognition and host-acceptance by the specialist parasitoid <Emphasis Type="Italic">Cotesia plutellae</Emphasis> (Kurdjumov)

Summary. Cotesia plutellae is a specialist parasitoid of Plutella xylostella. This specificity is potentially under the control of several factors before and after oviposition. Thereby, the stimuli that lead female parasitoids to host locations and to oviposition, might be at the basis of the specificity. We explore here the response of C. plutellae females exposed to host cuticular lipids. A total cuticular lipid extract of host caterpillars was fractionated into a hydrocarbon fraction and a non-hydrocarbon fraction. Neither fraction alone had any effect on oviposition behaviour in C. plutellae but the hydrocarbon fraction alone did seem to have a positive effect on the rate of antennal contact by the females. To induce oviposition behaviour, both fractions were necessary and reflect cooperation between at least one compound in each fraction. Identification of cuticular lipids shows that hydrocarbons were dominant (77%). Non-hydrocarbon compounds were mainly represented by 15-nonacosanone (18% of the total lipid extract). This ketone is rare in insect cuticle lipids and is thought to originate from the cabbage epicuticle where it is dominant with n-C₂₉ and 14- and 15-nonacosanol also found among the cuticular lipids of the host caterpillar.     

Temporal accumulation of phenylpropanoids in male fruit flies, <Emphasis Type="Italic">Bactrocera dorsalis</Emphasis> and <Emphasis Type="Italic">B. carambolae</Emphasis> (Diptera: Tephritidae) following methyl eugenol consumption

Summary. Males of dacine tephritids, Bactrocera carambolae and B. dorsalis are strongly attracted to, and compulsively feed on methyl eugenol (ME), a potent attractant for many Bactrocera species. While ME was shown to be biotransformed into phenylpropanoids, 2-allyl-4,5-dimethoxyphenol and (E)-coniferyl alcohol, and temporarily stored in the rectal gland of male B. dorsalis prior to release during courtship at dusk, B. carambolae male produces only the latter compound along with its de novo synthesized pheromone components. Both species were also shown to have different age-related response, sensitivity and consumption levels of ME. Here, we monitored and compared temporal changes in the accumulation profiles of these phenylpropanoids by the two sibling species, with male rectal glands being individually excised at different time intervals from 15 minutes to 20 days after initial ME feeding and analysed quantitatively. Results are discussed in light of plant-fruit fly co-evolution relationship.     

<Emphasis Type="Italic">Ceroplastes albolineatus</Emphasis>, the first scale insect shown to sequester pyrrolizidine alkaloids from its host-plant <Emphasis Type="Italic">Pittocaulon praecox</Emphasis>

Summary. The occurrence of pyrrolizidine alkaloids (PAs) in Pittocaulon (ex Senecio) praecox (Asteraceae) a species endemic to Mexico was established. The aboveground plant organs contain the 1,2-saturated monoester 7-angeloyl-dihydroxyheliotridane together with a small proportion of its 9-angeloyl isomer as major alkaloid. The monoesters are accompanied by the macrocyclic otonecine derivative senkirkine. Roots contain only related macrocyclic PAs with senecionine, senkirkine and platyphylline as major components; monoesters are absent. The broom-like succulent stems of P. praecox are infested by the scale insect Ceroplastes albolineatus conspicuously visible by its huge wax cover. All life-history stages, i.e. females, eggs, first instar nymphs (crawlers) and the wax cover were found associated with PAs. The measured PA concentrations clearly indicate sequestration. The highest PA concentrations (mg / g dry weight) reached are: mature females, 0.44; eggs, 0.58; crawlers, 0.37; wax cover, 0.08. The host plant as well as in the infesting scale insect contain the PAs exclusively as free bases. As a phloem-feeder C. albolineatus must acquire the PAs with the ingested phloem sap. This appears plausible since in Senecio species PA are transmitted and circulated through the phloem path. It is suggested that PAs may protect particularly the crawlers as the most endangered stage in the life-cycle of the scale insect.     

Olfactory cues,genetic relatedness and female mate choice in the agile antechinus (<Emphasis Type="Italic">Antechinus agilis</Emphasis>)

Females show mate preferences for males that are genetically dissimilar to themselves in a variety of taxa, but how females choose these males is not clearly understood. In this study, we examined the effects of olfactory stimuli and genetic relatedness on female mate choice in a small carnivorous marsupial, the agile antechinus (Antechinus agilis), during two breeding seasons. Captive female antechinus in oestrus were provided with a combination of male urine and body scent from two novel males, one more genetically similar and one more dissimilar to the females, in a Y-maze olfactometer. Genetic relatedness between females and pairs of males was determined using highly polymorphic, species-specific, microsatellite markers. Females consistently chose to visit the scents of males that were genetically dissimilar to themselves first, spent significantly more time near the source of those scents and showed more sexual and non-exploratory behaviours near those scents. These data demonstrate that chemosensory cues are important in mate choice in the agile antechinus and that females prefer males that are genetically dissimilar to themselves.     

How <Emphasis Type="Italic">Doratomyces stemonitis</Emphasis> copes with Benzoxazolin-2(3<Emphasis Type="Italic">H</Emphasis>)-one (BOA), its derivatives and detoxification products

Summary. Doratomyces stemonitis (Hyphomycetales, Dematiaceae) is a saprotrophic fungus belonging to the mycobiota of the cereal rhizosphere. The fungus is able to metabolize benzoxazolin-2-(3H)-one and a variety of its derivatives including higher plant detoxification products, microbial degradation products and the chemically rather stable 2-amino-(3H)-phenoxazin-3-one. D. stemonitis can use all of these compounds as sole C-sources but their utilization, especially that of microbial degradation products and 2-amino-(3H)-phenoxazin-3-one, seems to be highly energy consuming, resulting in slow mycelium growth and a change of colony morphology. Benzoxazolin-2-(3H)-one derived compounds induce the synthesis of different isoforms of a glycosylated protein with sequence homologies to the endo-1,3-β-glucanase Asp f2, an allergen from Aspergillus fumigatus and other Asp f2-like proteins e.g., from Verticillium dahliae or PRA1 antigen from Candida albicans. The induction of the protein is regarded as a stress response.     

Comparison of the flower scent of the sexually deceptive orchid <Emphasis Type="Italic">Ophrys iricolor</Emphasis> and the female sex pheromone of its pollinator <Emphasis Type="Italic">Andrena morio</Emphasis>

Summary. Ophrys flowers mimic the female produced sex pheromone of their pollinator species to attract males for pollination. The males try to copulate with the putative female and thereby pollinate the flower. Using electrophysiological and chemical analyses, floral volatiles released by O. iricolor as well as the female sex pheromone of its pollinator species, Andrena morio are investigated. Overall, 38 peaks comprising 41 chemical compounds, were found to release reactions in the antennae of male A. morio bees. Analyses using coupled gas chromatography-mass spectrometry revealed the presence of alkanes and alkenes with 20 to 29 carbon atoms, aldehydes (C9 to C24) and two esters. Almost all of those compounds were found in similar proportions in both, the floral extracts of O. iricolor and cuticle surface extracts of A. morio females. The pattern of biologically active volatiles described here is very similar to that used by other Ophrys species pollinated by Andrena males.     

Weaning in the guinea pig (<Emphasis Type="Italic">Cavia aperea</Emphasis> f. <Emphasis Type="Italic">porcellus</Emphasis>): Who decides and by what measure?

Offspring should be selected to influence maternal effort to maximize their own fitness, whereas mothers are selected to limit investment in present progeny. In mammals, this leads to a conflict over the amount of milk provided and the timing of weaning. The intensity and time course of such conflict has so far mostly been investigated experimentally in altricial rodents. However, it is expected that offspring options for conflict will depend on developmental state. We therefore investigated in the highly precocial domestic guinea pig (Cavia aperea f. porcellus) who decides over nursing performance and weaning and how pup state influences these decisions. Specifically, we tested whether a threshold mass of pups predicts weaning time. By exchanging older litters against neonates and vice versa, we produced a situation in which females differed in lactational stage from the cross-fostered pups. Our results indicate that females decide about the timing of weaning, as cross-fostered younger pups were weaned at a much younger age than controls and older pups benefited from continuing lactation of foster mothers. Growth rates did not differ in the treatment groups, and different weaning ages resulted in differing weaning mass refuting the hypothesis that weaning is based on a threshold mass of offspring. This constitutes clear evidence that in a precocial rodent, the guinea pig, decisions about maternal care are primarily determined by maternal state and little influenced by pup state despite the extreme precociality of offspring. We suggest that precocial pups show little resistance to early weaning when food is abundant, as they reach sufficient nutritional independence by the middle of lactation to enable independent survival.     

Worker policing and nest mate recognition in the ant <Emphasis Type="Italic">Formica fusca</Emphasis>

A conflict over male production arises in social insects where workers are able to lay unfertilized male eggs. This happens because each female (queen or worker) is most closely related to her own sons and is thus predicted to reproduce. The conflict is modulated by worker policing where workers prevent each other from reproducing by aggression or egg cannibalism. In this study, we show that in the ant Formica fusca, worker policing occurs by egg cannibalism rather than by overt aggression among workers. Furthermore, we show that, contrary to bees, wasps and other ant species, egg discrimination in F. fusca is not based only on a universal queen signature chemical and that nest mate recognition of eggs occurs.     

Female tree swallows (<Emphasis Type="Italic">Tachycineta bicolor</Emphasis>) increase offspring heterozygosity through extrapair mating

Recent attention has focused on genetic compatibility as an adaptive function for why females engage in extrapair mating. We tested the genetic compatibility hypothesis in tree swallows (Tachycineta bicolor) over five breeding seasons using data from ten microsatellite loci. Tree swallows are socially monogamous passerines exhibiting high levels of extrapair paternity. Overall, we found that 47% of offspring were the result of extrapair fertilizations, and 83% of females produced at least one extrapair offspring. Consistently for all years, extrapair offspring were more heterozygous than their maternal half-siblings, which is in accordance with the genetic compatibility hypothesis. The difference was largely caused by the high heterozygosity of extrapair offspring sired by unknown males, suggesting that females are engaging in extrapair copulations with geographically distant males to increase the likelihood of being inseminated by a more compatible mate. Our findings support the idea that postcopulatory mechanisms are important for females when assessing potential sires for their offspring.     

Paternity and performance of golden-winged warblers (<Emphasis Type="Italic">Vermivora chrysoptera</Emphasis>) and golden-winged X blue-winged warbler (<Emphasis Type="Italic">V. pinus</Emphasis>) hybrids at the leading edge of a hybrid zone

We examined two components of reproduction in a population of golden-winged warblers in the initial stages of hybridization with blue-winged warblers. First, we used genetic analyses of mate choice to determine whether copulations outside the social pairbond (extra-pair copulations; EPCs) occur in these species and their hybrids. Second, we compared several aspects of reproduction (pairing success, clutch size, hatching success, fledging success, and loss of paternity) in nests raised by phenotypic golden-winged warblers to those raised by hybrid individuals. Together, these data provide us with the first quantitative analysis of reproduction within this hybrid system. Our data suggest several reasons why the level of hybridization between these species is likely being underestimated. First, many birds in our population showed only subtle phenotypic signs of introgression indicating that hybrid status can only be determined by close examination. Second, high rates of extra-pair paternity indicate that we cannot base our estimates of hybridization on pairbond data alone. More than 30% of nestlings were the result of EPCs, occurring in 55% of all nests. Third, there was no difference in the number of hybrid or “pure” golden-winged warbler males chosen as social mates, relative to their abundance. Indeed, based on several components of nesting success, it appears as though hybrids are having as great realized reproductive success as are phenotypic golden-winged warblers in our population. Accordingly, we argue that hybrid reproductive success and extra-pair fertilizations are likely playing a major role in the ongoing hybridization between blue-winged and golden-winged warblers. Studies such as this one are important to our understanding of the process and implications of hybridization, and findings may be applicable to other hybrid zones.     

Preservation and loss of the honey bee (<Emphasis Type="Italic">Apis</Emphasis>) egg-marking signal across evolutionary time

Honey bee workers are able to distinguish queen-laid eggs from worker-laid eggs, and remove (‘police’) worker-laid eggs. The cue that police workers use is as yet unidentified but is likely to be a chemical signal. This signal benefits queens for it ensures their reproductive monopoly. It also benefits collective workers because it allows them to raise more closely related queen-laid males than the less-related sons of half sisters. Because both parties benefit from the egg-marking signal, it should be stable over evolutionary time. We show that Apis mellifera workers can distinguish queen-laid from worker-laid eggs of the dwarf honey bee A. florea, a phylogenetically distant species that diverged from the A. mellifera lineage 6–10 mya. However, A. mellifera workers are unable to distinguish worker-laid eggs of A. cerana, a much more recent divergence (2–3 mya). The apparent change in the egg-marking signal used by A. cerana may be associated with the high rates of ovary activation in this species.     

Structure and function of Dufour gland pheromones from the crazy ant <Emphasis Type="Italic">Paratrechina longicornis</Emphasis>

Summary. Dufour glands of Paratrechina longicornis were analyzed chemically using GC-MS techniques. The glands contained twelve alkanes between C₉ and C₂₀ chain length with undecane and tridecane as the main components, six alkenes (1- and 4-undecene, 1-, 4-, and 6- tridecene, and heptadecadiene), two alkyl formates (C₁₁ and C₁₃), and saturated and polyunsaturated acids. Many of the alkanes and alkenes were behaviorally active causing a short lasting attraction of ants with different intensity. Detection of the major gland compound undecane in recruitment trails by in vivo SPME sampling provided evidence for its use as a recruitment pheromone. Both poison gland (formic acid) and Dufour gland (undecane) contents were detected on SPME fibers which had been attacked by the ants.     

Relationship between filtration activity and food availability in the Mediterranean mussel <Emphasis Type="Italic">Mytilus galloprovincialis</Emphasis>

The filtration activity of the Mediterranean mussel, Mytilus galloprovincialis, was assessed under different concentrations and compositions of seston by using a new automated image acquisition and analysis system. This approach allowed for frequent and simultaneous measurements of valve gape and exhalant siphon area. Filtration rates were measured through clearance measurements whereas pumping rates were measured using hot-film probes. The average filtration rate (17.5 l g h⁻¹ DW⁻¹ for a 0.36 g DW mussel) recorded during the present study was higher than those available for Mytilus edulis when standardized to flesh dry weight but almost equivalent (17.5 l h⁻¹ g DW⁻¹ for a 53 mm shell length mussel) to those rates when standardized to shell length. Immediately after the addition of algal cells (Isochrysis galbana; 4.5 μm in size), valve gape, exhalant siphon area and filtration rate increased quickly as mussels reached their maximum filtration activity. These three parameters then gradually decreased until complete closure of the shell. The algal cell concentration inducing this transition was close to 800 cells ml⁻¹ and 0.5 μg Chl a l⁻¹. When algal concentration was maintained above this threshold by successive algal additions, both valve gape and exhalant siphon area remained maximal. Temporal changes in the exhalant siphon area were continuous as opposed to those of valve gape. Therefore, despite the significant correlation between these two parameters, valves and siphon were sometimes dissociated due to a reduction of the area or even a closure of the exhalant siphon while the valves remained open. The velocity of exhaled water tended to be constant irrespective of exhalant siphon area and thus pumping rates were a linear function of exhalant siphon area. Consequently, reductions in exhalant siphon area and pumping rate were almost similar in M. galloprovincialis. Our results thus clearly support the hypothesis that exhalant siphon area constitutes a better proxy of pumping rate than valve gape as already suggested for Mytilus edulis. Finally, the high filtration rates measured during the present study together with the high concentrations of inorganic matter (> 40 mg DW l⁻¹) requested to alter those rates suggest that the studied mussels were well adapted to oligotrophic waters featuring strong hydrodynamism and frequent sediment resuspension events.     

Settlement of the wreckfish (<Emphasis Type="Italic">Polyprion</Emphasis><Emphasis Type="Italic">americanus</Emphasis>)

The wreckfish (Polyprion americanus) is a panoceanic, long-lived, deep-water demersal species, characterized by an extended pelagic juvenile phase associated with floating objects. In the present study morphometric data from 435 specimens collected from Greece (Crete), Italy (Ionian Sea) and the Atlantic coast of France from September 1999 to March 2001 were analyzed to estimate the settlement size of the species and associated changes in morphology. The fishery sample included specimens from both the pelagic and the demersal stage. Length-at-settlement (TL₅₀) and 95% confidence intervals were estimated by fitting a logistic function. Eleven morphometric characters were analyzed, and the existing inflection points, reflecting alterations in body shape, were defined: (1) iteratively, as the transition point, splitting the data set into two groups, for which the reduced major axis functions, between a character and total length, of the successive groups best fit (minimum sum of squares of residuals) the combined data set and (2) where the second derivative of the fitted third-order polynomial functions to morphometric ratios equaled zero. The main size range within which wreckfish settled was 56-65 cm, and the means of the inflection points defined by the two methods were 61 and 64 cm. These results indicate changes in functional morphology associated with settlement. The monitoring of wreckfish juveniles caught in the wild and kept individually in captivity showed changes in growth and food intake at approximately the estimated settlement length and suggested that temperature was the most likely factor triggering settlement.     

Leaf surface compounds of the potato (<Emphasis Type="Italic">Solanum tuberosum</Emphasis>) and their influence on Colorado potato beetle (<Emphasis Type="Italic">Leptinotarsa decemlineata</Emphasis>) feeding

Summary.  The sampling behavior of the Colorado potato beetle Leptinotarsa decemlineata Say (CPB) involves examination of the surface of potato leaves. It has been suggested that leaf surface compounds (volatiles and cuticular waxes) may be involved in host-plant recognition, acceptance or discrimination. Here we report on the effect of leaf surface extracts of six Polish commercial potato varieties on CPB feeding. We tested the influence of potato leaf surface extracts on CPB adult and larval feeding, then separated the extracts with HPLC, and finally tested the effect of the HPLC-separated fractions on CPB feeding. The bioassays were performed using potato leaf discs deprived of their original surface compounds. Applied to test discs at concentrations ten times higher than natural (10 leaf area equivalent), the extracts deterred CPB adults and larvae from feeding. HPLC-separated fractions composed of alkanes, sesquiterpene hydrocarbons, wax esters, benzoic acid esters, fatty acid methyl, ethyl, isopropyl and phenylethyl esters, aldehydes, ketones, methyl ketones, fatty acids, primary alcohols, β-amyrin and sterols did not affect adult CPB feeding. Similarly, alkanes, sesquiterpene hydrocarbons, wax esters, methyl ketones, sesquiterpene alcohols and secondary alcohols had no effect on larval CPB feeding. The sterol fraction (cholesterol, β-sitosterol and stigmasterol) acted as a phagostimulant to CPB larvae. We isolated a fraction demonstrating a phagodeterrent effect on CPB adults and larvae. The qualitative composition of the deterrent fraction was quite similar in all potato extracts, but there were quantitative differences between the varieties. Much further work is needed to identify the compounds that can produce the deterrent effect.     

Herbicide exposure affects the chemical recognition of a non native predator in common toad tadpoles (<Emphasis Type="Italic">Bufo bufo</Emphasis>)

Summary. In amphibians and fishes, evidence is increasing that chemical cues from injured conspecifics can play a role in the chemical labelling and learned recognition of unfamiliar predators. In this laboratory study, we tested the prediction that prior chemical exposure to a non-native predator feeding on conspecific tadpoles will subsequently allow tadpoles of the common toad (Bufo bufo) to recognize the chemical cues specifically released by this starved predator. Furthermore, we investigated the vulnerability of this chemically-mediated process to herbicide contamination. With these aims in view, groups of tadpoles were kept either unexposed or exposed for ten days to chemical cues from Turkish crayfish (Astacus leptodactylus) previously fed on tadpoles, both in uncontaminated water and in the presence of four sublethal concentrations of amitrole (0.01, 0.1, 1 and 10 mg.l⁻¹). We then assessed the effects of the six conditioning treatments on general activity and behavioural response to chemical cues from starved crayfish. Larval treatments did not affect the general activity of the tadpoles. By contrast, the treatments had significant effects on the behavioural response to the test solution prepared form starved crayfish. The only tadpoles to show an antipredator behavioural response to the chemical stimulation from starved crayfish belonged to the groups derived from chemical exposure to tadpole-fed crayfish in uncontaminated water and in contaminated water with the lowest concentration of amitrole (0.01 mg.l⁻¹). Conversely, this chemical stimulation produced no behavioural change in the control group or in the groups derived from exposure to tadpole-fed crayfish in contaminated water containing 0.1, 1 and 10 mg.l⁻¹ of amitrole. This study demonstrates that chemical cues released during the predator’s feeding activity can subsequently be used by common toad tadpoles in the recognition of an unfamiliar predator. In addition, our results show that the presence of sublethal amitrole concentrations can impair this recognition process. Such a pesticide effect might be especially detrimental for amphibian populations threatened by invasive predators.     

Cardiac output during exposure to <Emphasis Type="Italic">Chattonella</Emphasis><Emphasis Type="Italic">marina</Emphasis> and environmental hypoxia in yellowtail (<Emphasis Type="Italic">Seriola</Emphasis><Emphasis Type="Italic">quinqueradiata</Emphasis>)

The experiments were undertaken to measure, for the first time, cardiac output in yellowtail (Seriola quinqueradiata) during exposure to a harmful red tide flagellate (Chattonella marina). The responses were compared with those during exposure to environmental hypoxia to evaluate the significance of the drop of arterial oxygen partial pressure (PaO₂) in the fish-kill mechanisms by C. marina. PaO₂ immediately decreased, whereas heart rate (HR) was maintained until shortly before death during exposure to C. marina. Suffocation developing during the exposure resulted from a decrease in blood oxygen content, but not from lowered blood flow to the tissue. Although exposure to both C. marina and hypoxia immediately decreased PaO₂, arterial oxygen content (CaO₂) and pH (pHa) were significantly lower, whereas HR and cardiac output (&Qdot;) remained significantly higher, for the C. marina-exposed fish than for hypoxia-exposed fish. Although the drop in PaO₂ appears to play a pivotal role in the mechanisms of fish death by C. marina, other physiological response(s) should also be considered.     

Absence of status signalling by structurally based ultraviolet plumage in wintering blue tits (<Emphasis Type="Italic">Cyanistes caeruleus</Emphasis>)

Structurally based ultraviolet (UV)-reflective plumage parts can be important cues in mate choice. However, it remains largely unknown if UV plumage variation can also function as a signal of social status during competitive interactions. In blue tits (Cyanistes caeruleus), the UV-reflective crown plumage functions as a female mate choice cue that probably indicates male quality, as males with higher UV reflectance have been shown to have better chances of over-winter survival. Possibly, the UV crown plumage acts as a status signal in the competition over scarce food sources during winter. To test this idea, we related dominance of individuals at an artificial food source during adverse winter conditions to spectrophotometric measurements of their crown plumage. However, while controlling for the confounding effects of sex, age, and distance from territory, we found no significant effect of crown UV reflectance on dominance. Consistent with this result, we also found no relation between crown UV reflectance and over-winter survival. We conclude that the structurally based UV reflectance of the blue tit crown feathers plays little role in competition between individuals during winter despite its importance as a cue in mate choice.     

Multiple feeding stimulants in <Emphasis Type="Italic">Sinapis alba</Emphasis> for the oligophagous leaf beetle <Emphasis Type="Italic">Phaedon cochleariae</Emphasis>

Summary. In earlier investigations on host plant discrimination of leaf beetles glucosinolates were described as feeding stimulants for the Brassicaceae specialist Phaedon cochleariae F. (Coleoptera: Chrysomelidae). However, since these findings could not be confirmed in later studies offering 2-propenylglucosinolate in concentrations corresponding to those detected in host plant leaf material, the identification of feeding stimulants of this leaf beetle species remained unclear. In order to investigate which compounds of the host plant Sinapis alba (Brassicaceae) are involved in feeding stimulation, leaf extracts of different polarities were tested in bioassays with adults of P. cochleariae. Number of feeding beetles and net consumption rates were highest on pea leaves painted with methanol extracts of S. alba, whereas weak feeding responses were also detectable for hexane extracts. In subsequent bioassay-guided fractionations of methanol extracts with semi-preparative high performance liquid chromatography, two distinct fractions, one containing glucosinolates and another containing flavonoids, were found to stimulate beetles to feed to variable degrees. Other collected fractions had zero activity. The combination of both active fractions evoked significantly higher consumption rates and stimulated more beetles to feed than fractions tested individually. At least one compound of each fraction, among these the main glucosinolate of S. alba, 4-hydroxybenzylglucosinolate, act additively. Effects of two different naturally-occurring ratios of glucosinolates and flavonoids on the strength of feeding responses were investigated by use of extracts of two sets of host plants differently exposed to radiation. One set was outdoors-exposed, whereas the second set was kept in the greenhouse. However, the feeding behaviour of P. cochleariae was not affected by the significantly different relative compositions of both compound classes in the host material. In conclusion, mustard leaf beetles need a combination of distinct plant metabolites acting in concert for feeding stimulation, whereby the mere presence of these stimulants, but probably not the ratio of involved compounds, determines their feeding response.