Pyrrolizidine Alkaloids Negatively Affect a Generalist Herbivore Feeding on the Chemically Protected Legume Crotalaria pallida.

Plant secondary metabolites can have opposing effects on adapted specialist and non-adapted, generalist herbivores. In this study, we used Heliothis virescens (Fabricius) (Lepidoptera: Noctuidae) as a generalist, non-adapted model herbivore to test the possible effects of Crotalaria pallida (Fabaceae: Papilionoideae) defenses on herbivore performance. Neonate H. virescens larvae were able to consume C. pallida leaves and fruits and grow for a few instars, but none of them survived to pupation. We added isolated pyrrolizidine alkaloids (PAs) to an artificial diet at different concentrations, and PA concentration significantly affected the number of larvae that achieved pupation. Larval survival was not reduced at a PA concentration similar to the concentration on green seeds of C. pallida, but it was significantly reduced at PA concentration 5 and 100 times higher. These results suggest that PAs in isolation are not the defense responsible for the mortality in fresh C. pallida plants, indicating the importance of other possible defenses. The negative effect of PAs on fitness of the non-adapted, generalist herbivore is in agreement with few previous studies, but it is in clear contrast to a previous study on the effects of PAs on the adapted specialist herbivore Utetheisa ornatrix (L.) that were able to sequester PAs with no fitness costs.

Are Aristolochic Acids Responsible for the Chemical Defence of Aposematic Larvae of Battus polydamas (L.) (Lepidoptera: Papilionidae)?

Aristolochic acids (AAs) are thought to be responsible for the chemical protection of the aposematic larvae Battus polydamas (L.) (Papilionidae: Troidini) against predators. These compounds are sequestered by larvae from their Aristolochia (Aristolochiaceae) host plants. Studying the role of the chemical protection of the second and fifth instars of B. polydamas against potential predators, we found that the consumption of larvae by the carpenter ant Camponotus crassus Mayr and young chicks Gallus gallus domesticus was dependent on larval developmental stage. Second instars were more preyed upon than fifth instars; however, the assassin bug Montina confusa Stål was not deterred by chemical defences of the fifth instar B. polydamas. Laboratory bioassays with carpenter ants and young chicks using palatable baits topically treated with a pure commercial mixture of AAs I and AAs II in concentrations up to 100 times those previously found in B. polydamas larvae showed no activity. Similar results were found in field bioassays, where palatable baits treated as above were exposed to the guild of predators that attack B. polydamas larvae and were also consumed irrespective of the commercial AA concentration used. These results suggest that the mixture of AAs I and AAs II have no defensive role against predators, at least against those investigated in the present work. Other compounds present in Aristolochia host plants such as O-glycosylated AAs; benzylisoquinoline alkaloids; and mono-, sesqui-, di-, and triterpenes, which can be sequestered by Troidini, could act as deterrents against predators.

Methyl jasmonate increases the tropane alkaloid scopolamine and reduces natural herbivory in Brugmansia suaveolens: is scopolamine responsible for plant resistance?

The tropane alkaloid (TA) scopolamine is suggested to protect Brugmansia suaveolens (Solanaceae) against herbivorous insects. To test this prediction in a natural environment, scopolamine was induced by methyl jasmonate (MJ) in potted plants which were left 10 days in the field. MJ-treated plants increased their scopolamine concentration in leaves and herbivory decreased. These findings suggest a cause-effect relationship. However, experiments in laboratory showed that scopolamine affect differently the performance of the specialist larvae of the ithomiine butterfly Placidina euryanassa (C. Felder & R. Felder) and the generalist fall armyworm Spodoptera frugiperda (J. E. Smith): the specialist that sequester this TA from B. suaveolens leaves was not negatively affected, but the generalist was. Therefore, scopolamine probably acts only against insects that are not adapted to TAs. Other compounds that are MJ elicited may also play a role in plant resistance against herbivory by generalist and specialist insects, and deserve future investigations.

Colony membership is reflected by variations in cuticular hydrocarbon profile in a Neotropical paper wasp, Polistes satan (Hymenoptera, Vespidae).

Nestmate recognition is one the most important features in social insect colonies. Although epicuticular lipids or cuticular hydrocarbons have both structural and defensive functions in insects, they also seem to be involved in several aspects of communication in wasps, bees and ants. We analyzed and described for the first time the cuticular hydrocarbons of a Neotropical paper wasp, Polistes satan, and found that variation in hydrocarbon profile was sufficiently strong to discriminate individuals according to their colony membership. Therefore, it seems that small differences in the proportion of these compounds can be detected and used as a chemical-based cue by nestmates to detect invaders and avoid usurpation.

Colony membership is reflected by variations in cuticular hydrocarbon profile in a Neotropical paper wasp, Polistes satan (Hymenoptera, Vespidae)

Nestmate recognition is one the most important features in social insect colonies. Although epicuticular lipids or cuticular hydrocarbons have both structural and defensive functions in insects, they also seem to be involved in several aspects of communication in wasps, bees and ants. We analyzed and described for the first time the cuticular hydrocarbons of a Neotropical paper wasp, Polistes satan, and found that variation in hydrocarbon profile was sufficiently strong to discriminate individuals according to their colony membership. Therefore, it seems that small differences in the proportion of these compounds can be detected and used as a chemical-based cue by nestmates to detect invaders and avoid usurpation.

Identification of sex pheromones of Lutzomyia longipalpis (Lutz & Neiva, 1912) populations from the state of São Paulo, Brazil.

In Brazil, four populations of Lutzomyia longipalpis each producing different sex pheromones are recognised. It has been suggested that these chemotype populations represent true sibling species. In this study we present the results of an analysis, by coupled gas chromatography-mass spectrometry, of the pheromones of males L. longipalpis from two different municipalities of the state of São Paulo. Our study showed that L. longipalpis from these two municipalities produced different sex pheromones from each other. This coupled with the remarkable difference between the epidemiological situation in Araçatuba and Espírito Santo do Pinhal, suggests that the (S)-9-methylgermacrene-B and cembrene-1 populations may have different vectorial capacities.

Identification of sex pheromones of Lutzomyia longipalpis (Lutz & Neiva, 1912) populations from the state of São Paulo, Brazil

In Brazil, four populations of Lutzomyia longipalpis each producing different sex pheromones are recognised. It has been suggested that these chemotype populations represent true sibling species. In this study we present the results of an analysis, by coupled gas cromotography - mass spectrometry, of the pheromones of males L. longipalpis from two different municipalities of the state of São Paulo. Our study showed that L. longipalpis from these two municipalities produced different sex pheromones from each other. This coupled with the remarkable difference between the epidemiological situation in Araçatuba and Espírito Santo do Pinhal, suggests that the (S)-9-methylgermacrene-B and cembrene-1 populations may have different vectorial capacities.

Determination of drug levels and the effect of diazepam on the growth of necrophagous flies of forensic importance in southeastern Brazil.

Larvae of Chrysomya albiceps (Wiedemann) and Chrysomya putoria (Wiedemann) (Diptera: Calliphoridae) were reared on tissues from rabbits administered twice the lethal dosage of diazepam in order to study the effects of this drug on the development of these two species. The rabbits were given 50mg of diazepam via ear vein infusion. From 18 to 54h, larvae feeding on tissues containing the drug developed more rapidly than larvae from the control colony for both fly species. The time required for pupariation and adult emergence was significantly greater for colony fed on tissues from diazepam dosed rabbits than for the control ones. These differences are significant for they are large enough to alter the estimate of postmortem interval based on fly development. The presence of diazepam could be detected through gas chromatography-mass spectrometer (GC-MS) in all rabbit samples and in almost all diptera samples in this experiment.

The essential amino acid lysine acts as precursor of glutamate in the mammalian central nervous system.

Lysine has long been recognized as an essential amino acid for humans and the lack or low supply of this compound in the diet may lead to mental and physical handicaps. Since lysine is severely restricted in cereals, the most important staple food in the world, the understanding of its biological roles must be a major concern. Here we show that lysine is an important precursor for de novo synthesis of glutamate, the most significant excitatory neurotransmitter in the mammalian central nervous system. We also show that the synthesis of glutamate from lysine, which is carried out by the saccharopine pathway, is likely to take place in neurons.

Stereochemical inversion of pyrrolizidine alkaloids byMechanitis polymnia (Lepidoptera: Nymphalidae: Ithomiinae): Specificity and evolutionary significance.

Pyrrolizidine alkaloids (PAs), acquired by adults or larvae of Danainae and Ithomiinae butterflies and Arctiidae moths from plants, protect these lepidopterans against predators and are biosynthetic precursors of male sex pheromones. The investigation of PAs in many species of wild-caught adults of Ithomiinae showed lycopsamine (1) [(7R)-OH, (2'S)-OH, (3'S)-OH] as the main alkaloid. In incorporation experiments, PA-free (freshly emerged) adults of the ithomiineMechanitis polymnia were fed seven PAs: lycopsamine and four of its known natural stereoisomers-indicine (2) [(7R)-OH, (2'R)-OH, (3'S)-OH], intermedine (3) [(7R)-OH, (2'S)-OH, (3'R)-OH], rinderine (4) [(7S)-OH, (2'S)-OH, (3'R)-OH], and echinatine (5) [(7S)-OH, (2'S)-OH, (3'S)-OH], and two PAs without the 7-OH: supinine (6) [(2'S)-OH, (3'R)-OH] and amabiline (7) [(2'S)-OH, (3'S)-OH]. Males epimerized PAs 3, 4, and 5 mainly to lycopsamine (1). Females fed these same three PAs changed a smaller proportion to lycopsamine; their lesser capacity to modify PAs corresponds to their normal acquisition of already transformed PAs from males during mating rather than through visits of adults to plant sources of PAs. The alkaloids1 and2, both 7R and 3'S, were incorporated without or with minimum change by males and females. Feeding experiments with6 and7 (males only) showed an inversion at the 3' center of6 and no change in7. The inversion from 7S to 7R (probably via oxyreduction) may be closely related to the evolution of acquisition of PAs by butterflies and moths. Two hypotheses are discussed: (1) The ancestral butterflies are probably adapted to tolerate, assimilate, and use (7R)-PAs (most common in plants; all widespread 1,2-unsaturated macrocyclic PA diesters show this configuration). The development of (7R)-PA receptors in the butterflies could lead to a specialization on this configuration in two ways: to help find PA plants and to utilize these components in sexual chemical communication. A later appearance of (7S)-PAs in plants could have selected an enzymatic system for the inversion of this chiral center in order to continue producing (7R)-PA-derived pheromones. (2) The inversion would be due to the evolution of a enzyme system specialized in the transport of (7R)-PAs to the integument; the failure of this system to carry (7S)-PAs led to an enzymatic system to invert them to transportable (7R)-PAs. In this case, the 7R configuration is an effect and not a cause of (7R)-PA-derived pheromones. In both hypotheses, the partial inversion of the 3'-asymmetric center, when the butterfly was fed intermedine (3), rinderine (4), and supinine (6), could be fortuitous due to the conformation of the molecule and/or the enzymatic system.

Pyrrolizidine alkaloids in the arctiid mothHyalurga syma.

The arctiid mothHyalurga syma (subfamily Pericopinae) sequesters pyrrolizidine alkaloids (PAs) from its larval food plantHeliotropium transalpinum (Boraginaceae). Colorimetric quantification of total PAs in the larvae, pupae, and adults ofHyalurga revealed mean values of about 286-445µg per individual (1.4-2.6% of dry weight). The PA mixtures found in the moth and its larval food plant were evaluated by GC-MS. Food-piant leaves were found to contain the diastereoisomeric retronecine esters indicine (IIIa), intermedine (IIIb), and lycopsamine (IIIc), and the heliotridine ester rinderine (IIId) only as minor constituents, whereas 3'-acetylrinderine (IVc) (68% of total PAs) and the respective 3'-acetyl esters of indicine (IVa) and intermedine (IVb) (both 17%) were the major alkaloids. Supinine (IIa) is detectable in traces only. The PA mixtures in eggs, larvae, pupae, and imagines ofHyalurga were identical: indicine, intermedine, and lycopsamine accompanied by considerable amounts of supinine and amabiline or coromandalinine (IIb/IIc) were the major components. Only larvae were found to store small quantities of a 3'-acetyl derivative. Rinderine and its 3'-acetyl ester were never found in the insects. Low concentrations of the arctiidspecific PA callimorphine (I) were present in larvae, pupae, and imagines. The differences in the PA patterns of the insects and their larval food plant suggest thatHyalurga is capable of modifying plant-derived PAs by inversion of the 7-OH configuration (conversion of the necine base heliotridine into retronecine), and perhaps the inversion of the 3'-OH [conversion of (+)-trachelanthic acid into (-)-viridifloric acid], although the possibility of a selective sequestration of the respective retronecine esters cannot be excluded. Some trials with the orb-weaving spiderNephila clavipes, a common neotropical predator, showed that both freshly emerged and field-caught adults ofHyalurga syma are liberated unharmed by the spider. The liberation could be related to the presence of PAs in the moths.