Designing a species-selective lure based on microbial volatiles to target Lobesia botrana.

Sustainable, low impact control methods, including mating disruption and microbial insecticides against L. botrana have been available for decades. Yet, successful implementation has been restricted to only a few grapevine districts in the world. A limiting factor is the lack of a female attractant to either monitor or control the damaging sex. Volatile attractants for both female and male insects can be used to assess when L. botrana populations exceed economic thresholds, and to decrease the use of synthetic pesticides within both conventional and pheromone programs. Rather than using host-plant volatiles, which are readily masked by background volatiles released by the main crop, we tested the attractiveness of volatiles that signify microbial breakdown and more likely stand out against the background odour. A two-component blend of 2-phenylethanol (2-PET) and acetic acid (AA) caught significant numbers of both sexes. Catches increased with AA and, to a minimal extent, 2-PET loads. However, a higher load of 2-PET also increased bycatches, especially of Lepidoptera and Neuroptera. Major (ethanol, ethyl acetate, 3-methyl-1-butanol) or minor (esters, aldehydes, alcohols and a ketone) fermentation volatiles, did surprisingly not improve the attraction of L. botrana compared to the binary blend of 2-PET and AA alone, but strongly increased bycatches. The most attractive lure may thus not be the best choice in terms of specificity. We suggest that future research papers always disclose all bycatches to permit evaluation of lures in terms of sustainability.

A pheromone antagonist liberates female sea lamprey from a sensory trap to enable reliable communication.

The evolution of male signals and female preferences remains a central question in the study of animal communication. The sensory trap model suggests males evolve signals that mimic cues used in nonsexual contexts and thus manipulate female behavior to generate mating opportunities. Much evidence supports the sensory trap model, but how females glean reliable information from both mimetic signals and their model cues remains unknown. We discovered a mechanism whereby a manipulative male signal guides reliable communication in sea lamprey (Petromyzon marinus). Migratory sea lamprey follow a larval cue into spawning streams; once sexually mature, males release a pheromone that mimics the larval cue and attracts females. Females conceivably benefit from the mimetic pheromone during mate search but must discriminate against the model cue to avoid orienting toward larvae in nearby nursery habitats. We tested the hypothesis that spawning females respond to petromyzonol sulfate (PZS) as a behavioral antagonist to avoid attraction to the larval cue while tracking the male pheromone despite each containing attractive 3-keto petromyzonol sulfate (3kPZS). We found 1) PZS inhibited electrophysiological responses to 3kPZS and abated preferences for 3kPZS when mixed at the same or greater concentrations, 2) larvae released more PZS than 3kPZS whereas males released more 3kPZS than PZS, and 3) mixtures of 3kPZS and PZS applied at ratios measured in larval and male odorants resulted in the discrimination observed between the natural odors. Our study elucidates how communication systems that arise via deception can facilitate reliable communication.

DREADD-induced silencing of the medial amygdala reduces the preference for male pheromones and the expression of lordosis in estrous female mice.

Sexually naïve estrous female mice seek out male urinary pheromones; however, they initially display little receptive (lordosis) behavior in response to male mounts. Vomeronasal-accessory olfactory bulb inputs to the medial amygdala (Me) regulate courtship in female rodents. We used a reversible inhibitory chemogenetic technique (Designer Receptors Exclusively Activated by Designer Drugs; DREADDs) to assess the contribution of Me signaling to females' preference for male pheromones and improvement in receptivity normally seen with repeated testing. Sexually naïve females received bilateral Me injections of an adeno-associated virus carrying an inhibitory DREADD. Females were later ovariectomized, treated with ovarian hormones, and given behavioral tests following intraperitoneal injections of saline or clozapine-N-oxide (CNO; which hyperpolarizes infected Me neurons). CNO attenuated females' preference to investigate male vs. female urinary odors. Repeated CNO treatment also slowed the increase in lordosis otherwise seen in females given saline. However, when saline was given to females previously treated with CNO, their lordosis quotients were as high as other females repeatedly given saline. No disruptive behavioral effects of CNO were seen in estrous females lacking DREADD infections of the Me. Finally, CNO attenuated the ability of male pheromones to stimulate Fos expression in the Me of DREADD-infected mice but not in non-infected females. Our results affirm the importance of Me signaling in females' chemosensory preferences and in the acute expression of lordosis. However, they provide no indication that Me signaling is required for the increase in receptivity normally seen after repeated hormone priming and testing with a male.

The mechanism for microsporidian parasite suppression of the hindgut bacteria of the migratory locust Locusta migratoria manilensis.

Locusts aggregate into bands of nymphs and swarms of adults that can pose a major threat to crop. Previous studies have shown that infection by the microsporidian parasite Paranosema locustae prevents locust aggregation behavior and we show that gut bacteria, which produce components of locust aggregation pheromones, are substantially reduced in locusts infected with P. locustae. We found that P. locustae could reduce the diversity, abundance and community composition of Locusta migratoria's gut bacteria. The parasite infection was also shown to interrupt the peroxidase activity of locust hindgut. Genome-wide expression analysis showed that the parasite infection suppressed peroxidase mRNA relative expression of locust hindgut, but had no effects on attacin expression and superoxide dismutase at 16 d post-inoculation with 20,000 P. locustae spores. Our findings reveal the mechanisms by which P. locustae impairs bacterial diversity and community structure of Locusta migratoria's gut bacteria.

The TIKI/TraB/PrgY family: a common protease fold for cell signaling from bacteria to metazoa?

We report that the metazoan Wnt protease and signaling inhibitor TIKI shares sequence homology with bacterial TraB/PrgY proteins, inhibitors of pheromone signaling essential for propagation of antibiotic resistance. Our analysis suggests that these proteins represent an ancient metalloprotease clan regulating cellular communications across biological kingdoms.

Antinociception in piauçu fish induced by exposure to the conspecific alarm substance.

Defense and antinociceptive responses can be elicited simultaneously by learned or innate danger signals when an organism is in imminent danger. Antinociception blocks the recuperative behavioral reactions following pain perception that could interfere with defensive efforts. Antinociception associated with fearful experiences involving a confrontation with a predator or a predator being in close proximity is well studied in mammals, but very little is known about how fear affects antinociception responses in fish. Antipredator behavior in Ostariophysan fish may be elicited by exposure to conspecific alarm substance (CAS) that can trigger a fear reaction. During the predator versus prey confrontation, the alarm pheromone system is activated and warns conspecifics about the assessment of predation risk. The purpose of the present study was to examine the possible activation of the endogenous analgesic system in Leporinus macrocephalus fish and to evaluate the modification of swimming activity induced by a nociceptive stimulus (i.e., a subcutaneous injection of 3% formalin) in fish that had previously been exposed to the CAS. The results show that formalin-mediated enhancement in swimming activity was significantly reduced after exposure to the CAS. This enhancement was blocked by naloxone (20mg/kg), which suggests that opioid signaling is involved. Therefore, we hypothesized that antinociceptive processes may occur in fish following exposure to a chemical substance that signals predation.

cGMP modulates responses to queen mandibular pheromone in worker honey bees.

Responses to social cues, such as pheromones, can be modified by genotype, physiology, or environmental context. Honey bee queens produce a pheromone (queen mandibular pheromone; QMP) which regulates aspects of worker bee behavior and physiology. Forager bees are less responsive to QMP than young bees engaged in brood care, suggesting that physiological changes associated with behavioral maturation modulate response to this pheromone. Since 3',5'-cyclic guanosine monophosphate (cGMP) is a major regulator of behavioral maturation in workers, we examined its role in modulating worker responses to QMP. Treatment with a cGMP analog resulted in significant reductions in both behavioral and physiological responses to QMP in young caged workers. Treatment significantly reduced attraction to QMP and inhibited the QMP-mediated increase in vitellogenin RNA levels in the fat bodies of worker bees. Genome-wide analysis of brain gene expression patterns demonstrated that cGMP has a larger effect on expression levels than QMP, and that QMP has specific effects in the presence of cGMP, suggesting that some responses to QMP may be dependent on an individual bees' physiological state. Our data suggest that cGMP-mediated processes play a role in modulating responses to QMP in honey bees at the behavioral, physiological, and molecular levels.

Pretreatment with CP-154526 blocks the modifying effects of alarm pheromone on components of sexual behavior in male, but not in female, rats.

We previously demonstrated that an alarm pheromone released from male donor Wistar rats evoked several physiological and behavioral responses in recipient rats. However, the pheromone effects on social behavior were not analyzed. In the present study, we examined whether the alarm pheromone affects sexual behavior in male or female rats. When a pair of male and female subjects was exposed to the alarm pheromone during sexual behavior, the ejaculation latency was elongated, the number of mounts was increased, and the hit rate (number of intromissions/number of mounts and intromissions) was decreased in the male subject. In contrast, female sexual behavior was not affected by the alarm pheromone. When we exposed only the male or female subject of the pair to the pheromone just before sexual behavior, the results were similar: the pheromone effects were evident in male, but not in female, subjects. In addition, when we pretreated with corticotropin-releasing factor (CRF) antagonist (CP-154526) before exposing the male subject to the alarm pheromone, the pheromone effects were attenuated in a dose-dependent manner. These results indicate that the alarm pheromone modifies male, but not female, components of sexual behavior and that CRF participates in the effects.

New pheromones and insect control strategies.

A survey of the new environmentally safe strategies used for insect control is presented. The survey includes mating disruption, pheromone antagonists as chemical communication inhibitors, pheromones and plant-based volatiles, attractant-and-kill, and push-pull strategies. Important successes have been obtained, particularly in mating disruption with significant reduction in pesticide use in low to moderate pest infestations. One important factor of concern is the high cost of semiochemicals and formulations containing them in comparison to the conventional insecticide treatments, and a combined effort by scientists, producers, and farmers should be made to reduce the cost of application of these semiochemicals.

Agonists and antagonists of antennal responses of gypsy moth (Lymantria dispar) to the pheromone (+)-disparlure and other odorants.

Insects use the sense of smell to guide many behaviors that are important for their survival. The gypsy moth uses a pheromone to bring females and males together over long distances. Male moth antennae are equipped with innervated sensory hairs that selectively respond to pheromone components and other odors. Host plant odors, in particular, are detected by moths and sometimes cause an enhancement of the antennal and behavioral responses of the moths to their pheromone. Inspired by naturally occurring agonists and antagonists of insect pheromone responses, we have screened, by electroantennogram (EAG) recordings, a collection of compound sets and of individual compounds. We have detected interference of some compounds with the EAG responses of male gypsy moth antennae to the pheromone. We describe three activities: (1) short-term inhibition or enhancement of mixed compound + pheromone plumes, (2) long-term inhibition of pure pheromone plumes following a mixed compound + pheromone plume, and (3) inhibition of the recovery phase of mixed compound + pheromone plumes. Long-term inhibition was robust, decayed within 30 s, and correlated with the inhibition of recovery; for both activities clear structure-activity patterns were detected. The commercial repellent N,N-diethyltoluamide (DEET) was included for comparison. The most active and reproducible short-term inhibitor was a mixture of 1-allyl-2,4-dimethoxybenzene and 2-allyl-1,3-dimethoxybenzene. The most active long-term inhibitors were a set of 1-alkoxy-4-propoxybenzenes, DEET, and 1-ethoxy-4-propoxybenzene. DEET was more specific in the olfactory responses it inhibited than 1-ethoxy-4-propoxybenzene, and DEET did not inhibit recovery, whereas 1-ethoxy-4-propoxybenzene did. Target sites for the three activities are discussed.

The alarm pheromone in male rats as a unique anxiety model: psychopharmacological evidence using anxiolytics.

Previously, we demonstrated that an alarm pheromone released from male donor Wistar rats evoked anxiety-related physiological and behavioral responses in recipient rats. Thus, we believe that this pheromone may increase anxiety levels in rats. In the current study, we evaluated the predictive validity of this alarm pheromone-induced anxiogenic effect in detail by investigating whether six types of human anxiolytics, each of which has a different mechanism of action, were efficacious in reducing anxiety, using changes in the acoustic startle reflex (ASR) as an index. The alarm pheromone-enhanced ASR was not affected by vehicle pretreatment but was dose-dependently attenuated by pretreatment with midazolam, phenelzine, propranolol, clonidine, and CP-154,526-although not buspirone. These results may reflect some aspects of the predictive validity of the alarm pheromone-induced anxiety in rats as an animal model of human anxiety.

Development and biological activity of a new antagonist of the pheromone of the codling moth Cydia pomonella.

A new pheromone antagonist of the codling moth Cydia pomonella is reported. Presaturation of the antennae of the insects with vapors of the antagonist (E,E)-8,10-dodecadienyl trifluoromethyl ketone, analogue of the main component of the pheromone (codlemone), resulted in lower electrophysiological responses to the pheromone relative to untreated insects. In the wind tunnel, the compound elicited a remarkable disruptive effect on all types of behavior of males flying toward a source baited with a pheromone/antagonist blend in 1:1, 1:5, and 1:10 ratios. The insects displayed erratic flights in the presence of the antagonist, as shown by their flight parameters in comparison to insects attracted to the pheromone alone. In the field, traps baited with mixtures of 1:10 codlemone/antagonist attracted considerably lower numbers of males than the natural attractant. The antagonist, however, did not inhibit the pheromone-degrading enzymes present in male antennae, suggesting that trifluoromethyl ketones are not sufficiently electrophilic to produce a stable intermediate adduct with a cysteine residue of the enzyme, a mechanism previously proposed for oxidase inhibition in insects. Overall and taking into account our previous reports and, particularly, the reduction in damage induced in maize fields by a trifluoromethyl ketone analogue of the pheromone of Sesamia nonagrioides (Sole, J.; Sans, A.; Riba, M.; Rosa, E.; Bosch, M. P.; Barrot, M.; Palencia, J.; Castella, J.; Guerrero, A. Reduction of damage by the Mediterranean corn borer, Sesamia nonagrioides , and the European corn borer, Ostrinia nubilalis , in maize fields by a trifluoromethyl ketone pheromone analog . Entomol. Exp. Appl. 2008, 126, 28-39), the antagonist might be a new candidate to consider in future strategies to control the codling moth.

Suppression pheromone and cockroach rank formation.

Although agonistic behaviors in the male lobster cockroach (Nauphoeta cinerea) are well known, the formation of an unstable hierarchy has long been a puzzle. In this study, we investigate how the unstable dominance hierarchy in N. cinerea is maintained via a pheromone signaling system. In agonistic interactions, aggressive posture (AP) is an important behavioral index of aggression. This study showed that, during the formation of a governing hierarchy, thousands of nanograms of 3-hydroxy-2-butanone (3H-2B) were released by the AP-adopting dominant in the first encounter fight, then during the early domination period and that this release of 3H-2B was related to rank maintenance, but not to rank establishment. For rank maintenance, 3H-2B functioned as a suppression pheromone, which suppressed the fighting capability of rivals and kept them in a submissive state. During the period of rank maintenance, as the dominant male gradually decreased his 3H-2B release, the fighting ability of the subordinate gradually developed, as shown by the increasing odds of a subordinate adopting an AP (OSAP). The OSAP was negatively correlated with the amount of 3H-2B released by the dominant and positively correlated with the number of domination days. The same OSAP could be achieved earlier by reducing the amount of 3H-2B released by the dominant indicates that whether the subordinate adopts an offensive strategy depends on what the dominant is doing.

Support for (Z)-11-hexadecanal as a pheromone antagonist in Ostrinia nubilalis: flight tunnel and single sensillum studies with a New York population.

The flight-tunnel response of male Z-strain European corn borer moths (ECB), Ostrinia nubilalis, from a population in New York State (USA), was significantly antagonized by addition of 1% (Z)-11-hexadecanal (Z11-16:Ald) to their sex pheromone (a 97:3 mix of (Z)- and (E)-11-tetradecenyl acetate [Z/E11-14:OAc]). The level of antagonism was equivalent to that observed for the previously identified ECB antagonist, (Z)-9-tetradecenyl acetate (Z9-14:OAc), and supports a recent report showing that Z11-16:Ald, a minor pheromone component of the Noctuid moth, Sesamia nonagrioides, caused antagonism of ECB pheromone communication in sympatric populations in the Iberian Peninsula. Single-sensillum recordings from ECB antennae, which included cross-adaptation experiments, showed that the same olfactory receptor neuron processing Z9-14:OAc inputs was responsible for detecting Z11-16:Ald, and that this neuron was not responsive to two other aldehydes, (Z)-9-tetradecanal (Z9-14:Ald) and (Z)-9-hexadecanal (Z9-16:Ald), found in other moth sex pheromones. Our results show that the antagonism is not confined to one geographic region, is specific for Z11-16:Ald, and that antagonist pathways might have the potential for processing a number of structurally similar compounds.

Molecular basis for control of conjugation by bacterial pheromone and inhibitor peptides.

In many bacteria expression of lateral gene transfer and of virulence factors is controlled by cell-cell signalling systems. Molecular interactions of microbial signal molecules with their cognate receptors are not well understood. For the Enterococcus faecalis conjugative plasmid pCF10, the PrgX protein serves as a molecular switch controlling expression of conjugation and virulence genes encoded by the plasmid. The induction state of a pCF10-carrying donor cell is determined by the ratio of two signalling peptides, cCF10 pheromone and iCF10 inhibitor. Recent analysis of PrgX/cCF10 interactions suggests a mechanism for conversion to the induced state. However, the means by which iCF10 peptide antagonizes cCF10 activity is unclear, and it has been suggested that inhibitor peptides block import of pheromone peptides. We now show that both of these peptides interact with the same binding pocket of PrgX, but they differentially alter the conformation of the protein and its oligomerization state, resulting in opposing biological activities.

Antagonism of pheromone response of Ostrinia nubilalis males and implications on behavior in the laboratory and in the field.

The antagonistic effect on the pheromone response and catabolism of male European corn borers, Ostrinia nubilalis, by several trifluoromethyl ketones is reported. (Z)-11-Tetradecenyl trifluoromethyl ketone (Z11-14:TFMK), the most closely related analogue of the main component of the pheromone, elicits a remarkable disruptive effect on close approach and source contact of males flying to a source baited with mixtures of the pheromone and the antagonist in 5:1 and 10:1 ratios. In this experiment, the male displayed an erratic flight track with frequent counter turns and intersections with the plume. In the field, the TFMK significantly lowered the number of males caught when mixed with the pheromone in a 10:1 ratio in comparison with the natural attractant. The compound was also a good inhibitor of the antennal esterase of the insect with a IC(50) value of 0.28 muM. The homologous (Z)-10-tridecenyl trifluoromethyl ketone, with one carbon less in the chain, also elicited an antagonistic effect in the wind tunnel, but in the field, the results were not conclusive. The effect induced was lower than the one displayed by Z11-14:TFMK including the activity as the esterase inhibitor (IC(50) value of 7.55 muM). The saturated tetradecyl trifluoromethyl ketone, tetradecyltrifluoropyruvamide, and (Z)-11-2-thiatetradecenyl trifluoromethyl ketone resulted completely inactive. The results obtained in conjunction to the previously shown low toxicity to mice by related trifluoromethyl ketones provide new important data for the putative utilization of these chemicals as new pest control agents.

Chemical communication: butterfly anti-aphrodisiac lures parasitic wasps.

To locate their hosts, parasitic wasps can 'eavesdrop' on the intraspecific chemical communications of their insect hosts. Here we describe an example in which the information exploited by the parasitic wasp Trichogramma brassicae is a butterfly anti-aphrodisiac that is passed from male to female Pieris brassicae butterflies during mating, to render them less attractive to conspecific males. When the tiny wasp detects the odour of a mated female butterfly, it rides on her (Fig. 1) to her egg-laying sites and then parasitizes the freshly laid eggs. If this fascinating strategy is widespread in nature, it could severely constrain the evolution of sexual communication between hosts.

Biorational approaches for insect control by enzymatic inhibition.

Conventional insecticides are highly toxic to many living organisms as well as to the environment; consequently, new biorational and more specific approaches to pest control have been developed. In this paper, we present an update of those approaches resulting from studies on inhibition of enzymes involved in key processes of insects life, particularly growth, molting and development of larvae and intraspecific communication of adults. The enzymes covered include pheromone degrading enzymes, pheromone biosynthetic enzymes, oxidoreductases, juvenile hormones, juvenile hormone epoxide hydrolases, proteases, molting hormones and phenoloxidases. Although these approaches refer to control of insect pests, many of them can be in principle also considered suitable for medicinal chemistry studies, since the mechanism of action of these inhibitors on related enzymes is quite similar, if not equal, in both fields.

The pheromone production of female Plodia interpunctella is inhibited by tyraminergic antagonists.

Several compounds were found to suppress the calling behavior and in vitro pheromone biosynthesis of the Indian meal moth, Plodia interpunctella. The compounds were screened by means of a calling-behavior bioassay with female P. interpunctella. Five derivatives with activities in the nanomolar range were identified, in order of decreasing pheromonostatic activity: 4-hydroxybenzaldehyde semicarbazone (42) > 5-(4-methoxyphenyl)-1,3-oxazole (38) > 5-[4-(tert-butyl)phenyl]-1,3-oxazole (40) > 5-(3-methoxyphenyl)-1,3-oxazole (35) > 5-(4-cyanophenyl)-1,3-oxazole (36). These compounds also showed in vitro inhibitory activity in intracellular de novo pheromone biosynthesis, as determined with isolated pheromone-gland preparations that incorporated [1-(14)C]sodium acetate in the presence of the so-called pheromone-biosynthesis-activating neuropeptide (PBAN). The non-additive effect of the inhibitor with antagonist (yohimbine) for the tyramine (TA) receptor suggests that it could be a tyraminergic antagonist. Three-dimensional (3D) computer models were built from a set of compounds. Among the common-featured models generated by the program Catalyst/HipHop, aromatic-ring (AR) and H-bond-acceptor-lipophilic (HBAl) features were considered to be essential for inhibitory activity in the calling behavior and in vitro pheromone biosynthesis. Active compounds, including yohimbine, mapped well onto all the AR and HBAl features of the hypothesis. Less-active compounds were shown to be unable to achieve an energetically favorable conformation, consistent with our 3D common-feature pharmacophore models. The present hypothesis demonstrates that calling behavior and PBAN-stimulated incorporation of radioactivity are inhibited by tyraminergic antagonists.