Cecropia Research Articles

Avaliação do efeito mutagênico do extrato hidroalcoólico bruto, por meio de bioensaios in vivo e prospecção fitoquímica de Cecropia glaziovii Sneth (embaúba), Cecropiaceae

O gênero Cecropia é conhecido popularmente por "embaúba" e apresenta várias espécies medicinais, sendo que estudos com a espécie C. glaziovii Sneth indicam que o extrato aquoso apresenta efeitos broncodilatador, anti-hipertensivo e antidepressivo, provavelmente, atribuídos às catequinas, procianidinas e flavonóides. O presente estudo objetivou avaliar os potenciais efeitos tóxico, citotóxicos, clastogênicos e aneugênicos do extrato bruto hidroalcoólico de folhas de C. glaziovii, por meio da dose letal média (DL50), da análise de micronúcleo em células de medula óssea de roedores e do teste com Allium cepa e realizar a prospecção fitoquímica. Os resultados demonstraram que as concentrações testadas não causaram toxicidade aguda, nem apresentaram atividade clastogênica e aneugênica, sugerindo que o extrato de C. glaziovii não interferem na divisão celular. A prospecção fitoquímica indicou a presença de taninos, flavonóides, fenóis, antraquinonas, cumarinas, catequinas, proteínas, açúcares redutores, depsídeos/depsidonas e triterpenos. Mesmo assim, esses resultados não prescrevem o consumo terapêutico da espécie, sem continuidade da sua avaliação e do cumprimento de todas as etapas pré-clínicas e clínicas.

Welcome Letter from Dr. Chikindas.

Dear Colleagues, It is with great pleasure that I introduce you to our new journal, Probiotics and Antimicrobial Proteins. The overall trend of the last few decades is obvious: there are fewer and fewer novel antibiotics being launched, and the resistance of human pathogens to commercially available drugs is on the rise. This remains a serious issue, with many cases of multiple drug resistance reported for Mycobacterium tuberculosis, Enterococcus spp., Salmonella, Escherichia coli, Campylobacter, etc.—not to mention several articles reporting on the ease of raising resistance to the drugs of choice in Bacillus anthracis. All of these factors have forced the international scientific community to search for new, effective antimicrobials. An increased understanding and acknowledgment of the role of healthy human probiota has placed the focus not on broad-spectrum antimicrobials with minimal side effects on the eukaryotic host but on antimicrobials that are specifically targeted against nondesired microorganisms that leave both healthy bacteria and the host alive and unharmed. These substances are ideally biodegradable, preferably nonimmunogenic, and available through cost-effective manufacturing processes. Many of the aforementioned features can be used to describe antimicrobial proteins which are produced by virtually all living creatures: histatins that are present in human saliva, defensins found in blood, plant defensins, cecropins from the Cecropia moth, magainins that were discovered in an African frog’s skin, and, of course, bacteriocins that are produced by microorganisms. These substances of a proteinaceous nature are ribosomally synthesized and serve as the first line of the host’s defense against intrusion into either its body or its ecological environment. This group of antimicrobials can be synergistically ‘‘joined’’ with secondary metabolites of a proteinaceous nature, such as e-poly-L-lysine that is produced by Streptomyces albulus, substances of a ‘‘mixed nature’’, such as lipopeptides. Common features among all of these molecules are that they are all naturally produced, and they all have protein (peptides) in the core of their structures. Most importantly, while their antimicrobial mechanisms of action are different from commercially available antibiotics, many of these proteins are also active against antibiotic-resistant bacteria. Taking into consideration all of the above, it is clear why there is a constant increase in papers published on antimicrobial proteins, and why the interest of the scientific community in these kinds of molecules is progressively growing.

Urticaceae Juss. do Parque Estadual das Fontes do Ipiranga, São Paulo, SP, Brasil

Urticaceae está representada por sete gêneros e 14 espécies na área do Parque. Destas, 11 são nativas e três cultivadas (Pilea cadierei Gagnep. & Guillaumin, Pilea nummulariifolia (Sw.) Wedd. e Boehmeria nivea Jacq.). Foram incluídos os gêneros Cecropia com três espécies e Coussapoa com uma espécie.

Structure and function of a custom anticancer peptide, CB1a

Several natural antimicrobial peptides including cecropins, magainins and melittins have been found to kill cancer cells. However, their efficacy may not be adequate for their development as anticancer agents. In this study, we used a natural antimicrobial peptide, cecropin B (CB), as a template to generate a novel anticancer peptide. Cecropin B is an amphipathic and polycationic peptide derived from the hemolymph of Hyalophora cecropia with well-known antimicrobial and cytolytic properties. The signature pattern of cecropins is W-x-(0,2)-[KDN]-x-{L}-K-[KRE]-[LI]-E-[RKN] (PROSITE: PS00268), and this signature sequence is located at N-terminus of CB. CB1a was constructed by repeating the N-terminal ten amino acids of CB three times and including a hinge near C-terminus. The circular dichroism spectra showed that CB1a is unstructured in aqueous solution, but adopt a helical conformation in membrane-like environment. The solution structure of CB1a in a polar solvent was also studied by NMR. CB1a formed a helix–hinge–helix in 20% HFIP solution, and it was found the bent angle between two helical segments was induced ranging from 60° to 110°. A heparin-binding motif is located in the central part of helix 1. Isothermal titration calorimetry reveals the association constant of CB1a bound to low molecular weight heparin is 1.66 × 10 5 M −1 at physiological ionic strength at 25 °C. Binding of CB1a to heparin produces a large conformational change toward a more structural state. CB1a demonstrated promising activity against several cancer cells but low toxicity against non-cancer cells. The IC 50 of CB1a on leukemia and stomach carcinoma cells were in the range of 2–8-fold lower than those of CB. Besides, CB1a exhibited low hemolytic activity against human red blood cells. Due to these properties, CB1a has the potential to become a promising anticancer agent.

Quantitative determination of catecholic degradation products from insect sclerotized cuticles

Acid hydrolysates of cuticle from various insect species were quantitatively analyzed for five catecholic amino acid adducts. Four of the adducts are ketocatechols; in three of them the amino acid moiety, either lysine, glycine or β-alanine, is connected via its amino group to the α-carbon atom of 3,4-dihydroxyacetophenone, in the fourth a tyrosine residue is connected to the same position via its phenolic group. The fifth adduct contains histidine linked via its imidazole-ring to the β-position of the dopamine sidechain. The three ketocatecholic adducts containing α-amino acids were obtained in significant yields from adult cuticles of the locust Schistocerca gregaria, the cockroaches Blaberus craniifer and Periplaneta americana, and the beetles Pachynoda sinuata and Tenebrio molitor, but only in trace amounts from larval and pupal cuticles of T. molitor, pupal cuticles of the moths Manduca sexta and Hyalophora cecropia, and puparia of the blowfly Calliphora vicina. The β-alanine-containing ketocatechol was not obtained from cuticle of locusts and T. molitor larvae and pupae, but it was present in the hydrolysates of the other cuticles. The β-histidine–dopamine adduct was obtained from all the cuticles, the highest yield was obtained from adult P. sinuata and the lowest yield was from adult S. gregaria. The β-histidine–dopamine adduct is derived from the product formed by reaction of p-quinone methides of N-acetyldopamine (NADA) or N-β-alanyldopamine (NBAD) with histidine residues in the cuticular proteins. The ketocatecholic adducts are assumed to be degradation products of crosslinks formed when oxidized dehydro-NADA reacts with the cuticular proteins. The insect species investigated appear to use both pathways for sclerotization, but to widely differing extents; the dehydro-NADA pathway dominates in cuticles which are exposed to strong deforming forces, such as those of adult locusts and cockroaches, and the p-quinone methide pathway dominates in cuticle of lepidopteran pupae and blowfly puparia, which are not exposed to strong mechanical forces but have to be effectively protected against microbial and fungal attacks.

Diet and Trophic Structure in a Community of Fruit-Eating Bats in Lacandon Forest, México

Assemblages of neotropical frugivorous bats display a trophic structure composed of groups of species reflecting diet preferences. The structure is hypothesized to be an ancestral trait, suggesting that very similar diets would be observed throughout a species' range. We examined the frequency of occurrence of seeds in feces of a community of frugivorous bats in Lacandon Forest, Mexico. Using metric multidimensional scaling, we found 3 groups of species, similar to those found in other regions and congruent with phylogenetic groupings, lending support to a historical origin of this structure. However, the diets of some species differed from those observed in other regions, in particular Barro Colorado Island (BCI), Panama. Here, we found species of the tribe Ectophyllini to be specialized on plants of the genus Cecropia, rather than Ficus-specialists as on BCI. This discrepancy can be related to differences in plant composition or in disturbance regimes, and we suggest that Ectophyllini (including Artibeus) are facultative specialists of the genera Ficus and Cecropia rather than strict Ficus-specialists.

Evaluation of the uniformity and stability of T-DNA integration and gene expression in transgenic apple plants

The generation of transgenic apple plants relies on the molecular analysis of transgene integration and expression based on polymerase chain reaction (PCR) analysis, blotting techniques and enzymatic assays on vitro leaves of putative transgenic regenerates. In order to assess the uniformity and the stability of transfer DNA (T-DNA) integration and gene expression, we studied 26 transgenic apple lines carrying the attacin E gene from Hyalophora cecropia , the β-glucuronidase gene, and the nptII gene. Plants were evaluated using standard molecular techniques, such as PCR, Southern blot, reverse transcription PCR (RT-PCR) and Enzyme Linked Immunosorbent Assay (ELISA), and propagated in vitro on non-selective antibiotic-free media for four years to mimic natural conditions in the field. In some T-lines transgene integration and expression did not remain stable; differences were also found between distinct plants of a single T-line. Individual plants with partially or completely silenced transgenes were identified as well as plants with non-detectable T-DNA. Several lines appeared chimeric or partially silenced. Although most molecular techniques can reliably detect the presence of transgenic cells, they often fail to detect mixtures of transformed and non-transformed cells, or cells with silenced transgenes. This should be taken into consideration, especially in the case of vegetatively propagated trees, where non-transformed or silenced plant parts could mistakenly be used as propagation material.

Two New Species of Lissoderes Champion, 1906 (Coleoptera: Curculionidae: Conoderinae) with Comments on the Ecology of the Genus

Lissoderes longinoi, new species, and L. masneri, new species are described from specimens collected in Costa Rica and Ecuador, respectively. Lissoderes championi Hespenheide is reported for the first time to be associated with Cecropia angustifolia Trécul, and L. cecropiae Hespenheide reported for the first time with C. obtusifolia Bertoloni. The distribution of members of the genus along an elevational transect in Costa Rica is described, and the ecology of their association with the plant genus Cecropia (Cecropiaceae) is reviewed.

Effect of Bat Exclusion on Patterns of Seed Rain in Tropical Rain Forest in French Guiana

ABSTRACTTo assess the impact of bats on seed dispersal in a tropical mature forest (Nouragues, French Guiana), we conducted a bat exclusion experiment and tested the hypotheses that an artificial reduction in the abundance of bats would result in: (1) a decrease in seed species diversity, at the community level; and (2) an increase in seed limitation (a failure of seeds to reach all suitable sites for germination) at the species level. Seed rain was sampled in two contiguous forest plots for a total of 120 d, using 49 seed traps (1 m2) arranged in 7 × 7 grids and spaced at 5‐m intervals. Using mist nets, bat activity was reduced in one forest plot for a total of 60 nights. Thirty‐nine plant species, or species groups, likely to be consumed and dispersed by bats, were collected within a total sample of 50,063 seeds. The overall seed rain was dominated by epiphytic Araceae and Cyclanthaceae (83.3%) and tree species within the genera Cecropia and Ficus (16.0%). Seeds from bat‐dispersed shrubs and treelets (Piper, Solanum, and Vismia) were relatively rare (0.7%). The bat exclusion resulted in a 30.5 percent reduction in seed species richness and increased seed limitation for most of the species sampled. Seed limitation was caused mainly by a reduced seed rain (seed source limitation) rather than a decrease in seed dispersal uniformity (seed dispersal limitation). Therefore, bat‐dispersed plants with low seed production are likely to be particularly affected by a decline in bat abundance, as a result of anthropogenic change.

RNA interference of Hemolin causes depletion of phenoloxidase activity in Hyalophora cecropia

Melanization is regulated by the prophenoloxidase cascade and functions as a response to intruding microorganisms in invertebrates. When injecting dsRNA of the lepidopteran immune protein hemolin in pupae of Hyalophora cecropia (Lepidoptera: Saturniidae), we observed a significant reduction in phenoloxidase activity after 24 h, but not after 72 h. The link between hemolin and the prophenoloxidase system suggests that hemolin is a pattern recognition protein important for the triggering of the prophenoloxidase cascade in the defence against bacterial infections.

Steroid hormone secretion in insects comes of age.

The control of molting and metamorphosis in insects first captured the attention of physiologists in the 1920s, when the Polish physiologist Stephan Kopec showed that the brain stimulated molting in larvae of the gypsy moth (1). However, 70 years elapsed before the hormone from the brain, prothoracicotropic hormone (PTTH), was finally sequenced by groups led by H. Ishizaki and A. Suzuki (2). During the interim, it was shown that the brain’s role in causing molting was not direct but was mediated through its action on paired endocrine glands in the thorax, the prothoracic glands (PGs), that then secreted a steroid molting hormone, ecdysone. Circulating PTTH acts on these glands to stimulate steroidogenesis via calcium entry and cAMP generation (3) in a manner similar to that seen in some vertebrate steroidogenic systems. Although the control of ecdysone secretion by a circulating tropic hormone has been a central concept in comparative endocrinology for >40 years, a series of recent articles have revealed that negative factors are also at play (4–6). The article by Yamanaka et al . (7) in a recent issue of PNAS described some novel peptides that exert inhibitory control on the PGs and showed that the pathway by which they exert their effects is through the direct innervation of the glands. The simple picture of PG control in insects arose from the simplicity of the system first used to elucidate the brain–PG axis. Pioneering work by Carroll Williams (8) exploited diapausing pupae of the giant silkmoth, Hyalophora cecropia , whose development shuts down early in metamorphosis to deal with the rigors of winter. To then activate adult differentiation in the spring, the dormant PGs required PTTH released from the brain, and diapausing pupae whose brains were surgically removed never resumed development, although they survived for months (8 … *E-mail: jwt{at}u.washington.edu

A Case of Natural Hybridization Between Hyalophora cecropia and Hyalophora columbia (Lepidoptera: Saturniidae) in Nova Scotia

Abstract Progeny of a wild female of Hyalophora cecropia (Linnaeus) (Lepidoptera: Saturniidae) were reared and appear to be natural hybrids resulting from a mating with Hyalophora columbia (Smith).

Enhanced resistance to the rice blast fungus Magnaporthe grisea conferred by expression of a cecropin A gene in transgenic rice

Cecropins are a family of antimicrobial peptides, which constitute an important key component of the immune response in insects. Here, we demonstrate that transgenic rice (Oryza sativa L.) plants expressing the cecropin A gene from the giant silk moth Hyalophora cecropia show enhanced resistance to Magnaporthe grisea, the causal agent of the rice blast disease. Two plant codon-optimized synthetic cecropin A genes, which were designed either to retain the cecropin A peptide in the endoplasmic reticulum, the ER-CecA gene, or to secrete cecropin A to the extracellular space, the Ap-CecA gene, were prepared. Both cecropin A genes were efficiently expressed in transgenic rice. The inhibitory activity of protein extracts prepared from leaves of cecropin A-expressing plants on the in vitro growth of M. grisea indicated that the cecropin A protein produced by the transgenic rice plants was biologically active. Whereas no effect on plant phenotype was observed in ER-CecA plants, most of the rice lines expressing the Ap-CecA gene were non-fertile. Cecropin A rice plants exhibited resistance to rice blast at various levels. Transgene expression of cecropin A genes was not accompanied by an induction of pathogenesis-related (PR) gene expression supporting that the transgene product itself is directly active against the pathogen. Taken together, the results presented in this study suggest that the cecropin A gene, when designed for retention of cecropin A into the endoplasmic reticulum, could be a useful candidate for protection of rice plants against the rice blast fungus M. grisea.

20‐Hydroxyecdysone indirectly regulates Hemolin gene expression in Hyalophora cecropia

Development and innate immune defence are two vital processes that have been demonstrated to use the same or similar molecules and signalling pathways in insects. Hemolin is a moth haemolymph protein belonging to the immunoglobulin superfamily. It is strongly induced upon bacterial infection. However, recent studies indicate a developmental regulation of hemolin. We show that the steroid hormone 20-hydroxyecdysone (20E) can activate the expression of Hyalophora cecropia Hemolin (HcHemolin) in the fat body of diapausing pupae. Using the protein synthesis inhibitor cycloheximide we demonstrate that Hemolin up-regulation by 20E requires ongoing protein synthesis. Moreover, 20E enhances transcription of the Hemolin gene in response to bacteria. Comparing the upstream regions of Manduca sexta Hemolin (MsHemolin) and HcHemolin, we identified four putative regulatory sites. Two are putative hormone response elements (HREs), one with an imperfect inverted repeat (HRE-IR) and one with a monomeric site (HRE-M). An additional monomeric hormone receptor site (MRE) is present only in HcHemolin. The third conserved motif is similar to the interferon (IFN) regulatory factor binding element (IRF-E) and IFN-stimulated response element (ISRE). The fourth conserved element is a kappaB motif situated between the Cap-site and the TATA-box. Finally, by electrophoresis mobility shift assay we demonstrate that the HRE-IR forms specific complexes with nuclear extract proteins of normal pupae that increase after 20E stimulation.

Solution structures of stomoxyn and spinigerin, two insect antimicrobial peptides with an α‐helical conformation

Stomoxyn and spinigerin belong to the class of linear cysteine-free insect antimicrobial peptides that kill a range of microorganisms, parasites, and some viruses but without any lytic activity against mammalian erythrocytes. Stomoxyn is localized in the gut epithelium of the nonvector stable fly that is sympatric with the trypanosome vector tsetse fly. Spinigerin is stored and secreted by hemocytes from the fungus-growing termite. The structure of synthetic stomoxyn and spinigerin in aqueous solution and in TFE/water mixtures was analyzed by CD and NMR spectroscopy combined with molecular modeling calculations. Stomoxyn and spinigerin adopt a flexible random coil structure in water while both assume a stable helical structure in the presence of TFE. In 50% TFE, the structure of stomoxyn is typical of cecropins, including an amphipathic helix at the N-terminus and a hydrophobic C-terminus with helical features that probably fold in a helical conformation at higher TFE concentration. In contrast to stomoxyn, spinigerin acquires very rapidly a helical conformation. In 10% TFE the helix is highly bent and the structure is poorly defined. In 50% TFE, the helical structure is well defined all along its sequence, and the slightly bent alpha-helix displays an amphiphilic character, as observed for magainin 2. The structural similarities between stomoxyn and cecropin A from Hyalophora cecropia and between spinigerin and magainin 2 suggest a similar mode of action on the bacterial membranes of both pairs of peptides. Our results also confirm that TFE induces helix formation and propagation for amino acids showing helical propensity in water but also enhances the helix propagation propensity of nonpolar beta-branched residues.

Molecular characterization and expression of prothoracicotropic hormone during development and pupal diapause in the cotton bollworm, Helicoverpa armigera

Using a strategy of rapid amplification of cDNA ends, the cDNA encoding prothoracicotropic hormone (PTTH) was cloned from the brain of Helicoverpa armigera (Hearm). The Hearm-PTTH cDNA contains an open reading frame encoding a 226-amino acid preprohormone, which shows high identity with the closely related noctuid moths, Helicoverpa zea (98%) and Heliothis virescens (94%), and low identity with five species of Bombycoidea: Bombyx mori (57%), Manduca sexta (55%), Hyalophora cecropia (52%), Samia cynthia ricini (49%) and Antheraea peryni (48%). Hearm-PTTH cDNA shares important structural characterization known from other PTTHs, such as seven cysteine residues, proteolytic cleavage site, glycosylation site, and hydrophobic regions within the mature peptides. Northern blot analysis indicated a 0.9 kb transcript present only in the brain. Using the more sensitive technique of RT-PCR, PTTH mRNA was also detected in the subesophageal ganglion, thoracic ganglion, abdominal ganglion, midgut and fat body. During the pupal stage, PTTH mRNA in the brain remained at a constant high level in nondiapausing individuals, was low in diapausing pupae, but increased again at diapause termination. The PTTH protein was detected only in the brain by Western blot analysis. Immunocytochemical results revealed that Hearm-PTTH is localized in two pairs of dorsolateral neurosecretory cells within the brain. Recombinant Hearm-PTTH was successfully expressed in E. coli, and purified recombinant-PTTH was effective in breaking pupal diapause. The results are consistent with a role for PTTH in the regulation of diapause in this species.

Chlorinated tyrosine derivatives in insect cuticle

A method for quantitative measurement of 3-monochlorotyrosine and 3,5-dichlorotyrosine in insect cuticles is described, and it is used for determination of their distribution in various cuticular regions in nymphs and adults of the desert locust, Schistocerca gregaria. The two chlorinated tyrosine derivatives were present in all analyzed regions in mature adult locusts, the highest concentrations were found in the sclerotized cuticle of femur and tibia, but significant amounts were also present in the unsclerotized arthrodial membranes. Small amounts of the two amino acids were obtained from pharate, not-yet sclerotized cuticle of adult femur and tibia, the amounts increased rapidly during the first 24 h after ecdysis and more slowly during the next two weeks. Control analyses using stable isotope dilution mass spectrometry have confirmed that the chlorinated tyrosines are not artifacts formed during sample hydrolysis. Mono- and dichlorotyrosine are also present in cuticular samples from other insect species, such as the beetle, Tenebrio molitor, the moth Hyalophora cecropia, the cockroach Blaberus craniifer, and the bug Rhodnius prolixus, but not in the sclerotized puparial cuticle of the blowfly, Calliphora vicina, or in sclerotized ootheca from the cockroach, Periplaneta americana. Cuticular sclerotization and formation of chlorotyrosines occur simultaneously in locust legs; sclerotized cuticles tend to have a higher content of chlorotyrosines than unsclerotized cuticles, but it is concluded that the chlorotyrosines are not just a by-product from the sclerotization process.

Characterization of antimicrobial peptides against a US strain of the rice pathogen Rhizoctonia solani.

To identify antimicrobial peptides with high lytic activity against Rhizoctonia solani strain LR172, causal agent of rice sheath blight and aerial blight of soyabeans in the US. Among 12 natural and synthetic antimicrobial peptides tested in vitro, the wheat-seed peptide, purothionin, showed the strongest inhibitory activity that was similar to the antifungal antibiotics, nystatin and nikkomycin Z. Cecropin B, a natural peptide from cecropia moth, and synthetic peptide D4E1 produced the highest inhibitory activity against R. solani among linear peptides. Membrane permeabilization levels strongly correlated with antifungal activity of the peptides. Noticeable changes in membrane integrity were observed at concentrations of >/=0.5 micromol l(-1) for purothionin, 2 micromol l(-1) for cecropin B, D4E1, D2A21, melittin, and phor21, and 8 micromol l(-1) for magainin II and phor14. An increase of nuclear membrane permeabilization was observed in fungal cells treated with cecropin B, but not with purothionin. Diffusion of nuclear content was observed by fluorescent microscopy 10 min after adding a lethal concentration of cecropin B. Evaluation by electron microscopy confirmed severe cytoplasmic degradation and plasma membrane vesiculation. Purothionin and cecropin B were the most stable against proteolytic degradation when added to liquid cultures of R. solani. Purothionin, cecropin B, D4E1 and phor21 were shown to exhibit high in vitro lytic activity against R. solani strain LR172 for rice and soyabean. These peptides are greater than 16 amino acids long and rapidly increase fungal membrane permeabilization. Resistance to proteolysis is important for sufficient antifungal activity of antimicrobial peptides. Selected antimicrobial peptides offer an attractive alternative to traditional chemicals that could be utilized in molecular breeding to develop crops resistant to rice sheath blight and aerial blight of soyabean.

Influences of Fruit Diversity and Abundance on Bird Use of Two Shaded Coffee Plantations1

We studied avian foraging at two shaded coffee plantations in Ciales, Puerto Rico. Both coffee plantations contained patches of second-growth forest but differed in shade types; one was a rustic plantation with a species-diverse shade including many fruiting plant species and the other was a commercial polyculture shaded almost solely by Inga vera. We quantified foraging activity of five fruit-eating bird species (Euphonia musica, Loxigilla portoricensis, Nesospingus speculiferus, Spindalis portoricensis, and Vireo altiloquous) and monthly fruit abundance in the coffee plantation and adjacent second-growth forest habitats at each site. Fruits comprised more than 50 percent of the diets for four of five focal bird species. We found a significant difference in the number of foraging records for focal bird species between coffee and forest habitats in the commercial polyculture but found few differences between these habitats in the rustic coffee farm. Overall, foraging activity was positively correlated with the abundance of fruits across study sites. Bird foraging was concentrated on plant species in the genera Cecropia, Miconia, Schefflera, Phoradendron, and Guarea, which together accounted for over 50 percent of frugivory records. Plant species in such genera fruited over prolonged time periods and provided birds with a fairly constant fruit supply. Our findings underscore the importance of fruiting plant species in making coffee plantations suitable habitat for birds and suggest that native fruiting plants be incorporated in coffee farms for avian conservation.

Distribution of cuticular protein mRNAs in silk moth integument and imaginal discs

The distributions of mRNAs for two cuticular proteins of Hyalophora cecropia were examined with RT-PCR and in situ hybridization. For major regions of larval and pupal cuticle, there was a strong correspondence between the type of cuticle and the predominant cuticular protein message found. Epidermal cells underlying soft cuticle had mRNA for HCCP12, with a RR-1 consensus attributed to soft cuticle, while the epidermal cells associated with hard cuticle had predominantly mRNA for HCCP66, a protein with the RR-2 consensus attributed to hard cuticle. Both messages were found in all areas of the pupal fore- and hind-wings, with modest area-specific difference in concentration being much less than differences in the relative abundance of these cuticular proteins. mRNA for HCCP12 was present in imaginal discs of feeding larvae of H cecropia. Data from Bombyx mori available at SilkBase ( http://www.ab.a.u-tokyo.ac.jp/silkbase/) revealed that imaginal discs from feeding larvae had abundant mRNA for RR-1 cuticular proteins, representing six distinct gene products. Only discs from spinning larvae had mRNAs that coded for RR-2 proteins arising from 10 distinct genes. Thus, lepidopteran wing imaginal discs can no longer be regarded as inactive in larval cuticle production.