Differential liver histopathological responses to amphibian chytrid infection.

Amphibians have been facing a pandemic caused by the deadly fungus Batrachochytrium dendrobatidis (Bd). Although studies have elucidated cutaneous and homeostatic disturbances, it is still unknown if the hepatic function can be affected or if hepatic effects differ among host species. Thus, we evaluated the effects of an experimental Bd infection on the liver (histopathology and the hepatosomatic index) of 2 anuran species (Xenopus laevis and Physalaemus albonotatus) with different susceptibilities to Bd infection and compared them to uninfected controls. Bd infection increased the melanomacrophage cell area and induced leukocyte infiltration in both species. The effects were more pronounced in the sensitive species, P. albonotatus, which showed severe reduction in glycogen stores and liver atrophy, due to energetic imbalance. Hepatocytes of P. albonotatus also showed ballooning degeneration (vacuolization), which could lead to cell death and liver failure. Our results provide evidence that although the sensitive species showed more severe effects, the tolerant species also had hepatic responses to the infection. These findings indicate that hepatic function can play an important role in detoxification and in immune responses to chytridiomycosis, and that it may be used as a new biomarker of health status in chytrid infections.

A new brilliantly blue-emitting luciferin-luciferase system from Orfelia fultoni and Keroplatinae (Diptera).

Larvae of O. fultoni (Keroplatidae: Keroplatinae), which occur along river banks in the Appalachian Mountains in Eastern United States, produce the bluest bioluminescence among insects from translucent areas associated to black bodies, which are  located mainly in the anterior and posterior parts of the body. Although closely related to Arachnocampa spp (Keroplatidae: Arachnocampininae), O.fultoni has a morphologically and biochemically distinct bioluminescent system which evolved independently, requiring a luciferase enzyme, a luciferin, a substrate binding fraction (SBF) that releases luciferin in the presence of mild reducing agents, molecular oxygen, and no additional cofactors. Similarly, the closely related Neoceroplatus spp, shares the same kind of luciferin-luciferase system of Orfelia fultoni. However, the molecular properties, identities and functions of luciferases, SBF and luciferin of Orfelia fultoni and other  luminescent members of the Keroplatinae subfamily still remain to be fully elucidated. Using O. fultoni as a source of luciferase, and the recently discovered non-luminescent cave worm Neoditomiya sp as the main source of luciferin and SBF, we isolated and initially characterized these compounds. The luciferase of O. fultoni is a stable enzyme active as an apparent trimer (220 kDa) composed of ~70 kDa monomers, with an optimum pH of 7.8. The SBF, which is found in the black bodies in Orfelia fultoni and in smaller dark granules in Neoditomiya sp, consists of a high molecular weight complex of luciferin and proteins, apparently associated to mitochondria. The luciferin, partially purified from hot extracts by a combination of anion exchange chromatography and TLC, is a very polar and weakly fluorescent compound, whereas its oxidized product displays blue fluorescence with an emission spectrum matching the bioluminescence spectrum (~460 nm), indicating that it is oxyluciferin. The widespread occurrence of luciferin and SBF in both luminescent and non-luminescent Keroplatinae larvae indicate an additional important biological function for the substrate, and therefore the name keroplatin.

Optimized Histological Preparation of Ovary for Ovariole Counting in Africanized Honey Bee Queens (Hymenoptera: Apidae).

Techniques for counting ovariole number in virgin and mated Apis mellifera L. queens have been described in previous studies. Having a systematic and fast way to collect this measurement can help accelerate bee breeding programs, because selection decisions can be taken faster. The aim of this work was to develop an efficient histological method to preserve ovaries that allows assessing the number of ovarioles in newly emerged virgin queens, and also in mated queens, in a shorter time than the methods already published. The proposed method resulted in images suitable for ovariole counting in both newly emerged and mated queens, and the total histological process took less than 10 h. This method provides the optimization of the histological procedure for research breeding programs that use ovariole number as selection criteria for improving reproduction and production traits.

Novel findings on the impact of chytridiomycosis on the cardiac function of anurans: sensitive vs. tolerant species.

BACKGROUND: Understanding of the physiological effects of chytridiomycosis is crucial to worldwide amphibian conservation. Therefore, we analyzed the cardiac function of two anuran species (Xenopus laevis and Physalaemus albonotatus) with different susceptibilities to infection by the causative agent of chytridiomycosis, Batrachochytrium dendrobatidis (hereafter Bd). METHODS: We analyzed the in situ heart rate (f H - bpm), relative ventricular mass (RVM -%), and Ca2+ handling in heart of Bd infected animals compared to uninfected controls of both study species. RESULTS: Bd infection resulted in a 78% decrease in contraction force values in P. albonotatus when compared to the less susceptible X. laevis. This negative effect was even more evident (82%) for the cardiac pumping capacity. The time to reach peak tension was 125% longer in P. albonotatus than in X. laevis, and cardiac relaxation was 57% longer. DISCUSSION: These results indicate a delay in the cardiac cycle of P. albonotatus on a beat-to-beat basis, which was corroborated by the bradycardia observed in situ. In summary, Bd-sensitive species present impaired cardiac function, which could be a factor in mortality risk. The more pronounced effects of Bd in P. albonotatus may not only result from electrolyte imbalance, as previously reported, but also could be an effect of toxins produced by Bd. For X. laevis, the ability to promote cardiac adjustments seems to be an important homeostatic feature that allows greater tolerance to chytridiomycosis. This study provides new physiological mechanisms underlying the tolerance or susceptibility of amphibian species to chytridiomycosis, which determine their adaptability to survive in the affected environments.

Impact of an environmental relevant concentration of 17α-ethinylestradiol on the cardiac function of bullfrog tadpoles.

This study evaluated if a concentration of 17α-ethinylestradiol (EE2 - 10 ng L(-1) for 96 h) normally found in Brazilian surface waters exerts any impact on cardiac function of bullfrog tadpoles (25 Gosner stage), Lithobates catesbeianus. During exposure, the animals' activity level (AL -% of active individuals) was monitored twice a day. Then, the in loco heart rate (f(H) - bpm) was determined, as well as the relative ventricular mass (RVM - % of body mass). Afterwards, cardiac ventricles were mounted for isometric force recordings (CS - mN mm(-2)), and determination of the cardiac pumping capacity (CPC - mN mm(-2) min(-1)). EE2 did not affect tadpoles' AL, although it resulted in a tachycardia in animals exposed to EE2 (f(H) = 66 bpm) when compared to controls (f(H) = 52 bpm), suggesting that EE2 acts directly on the cardiac muscle of tadpoles, rather than being a result of an increased cardiac demand due to a higher activity level (i.e., avoidance response). Additionally, EE2 exerted a positive inotropic response, which resulted in a higher CPC, which occurred independently of an increase in the number of myofibrils of EE2-exposed animals, since RVM remained similar between experimental groups. Thus, the increase on cardiac demand induced by the exposure to EE2 elevates considerably the animal energy expenditure, diverting a large amount of energy that tadpoles could use for their growth and development. These alterations can make amphibians more susceptible to predators and reduce the likelihood to reach reproductive stage.

Larval development of Physocephala (Diptera, Conopidae) in the bumble bee Bombus morio (Hymenoptera, Apidae)

Larval development of Physocephala (Diptera, Conopidae) in the bumble bee Bombus morio (Hymenoptera, Apidae). In the summer of 2012, a high incidence of conopid larvae was observed in a sample of female B. morio collected in remaining fragments of semidecidual forest and Cerrado, in the municipality of Sorocaba, state of São Paulo, Brazil. The larval development of conopid flies was studied, beginning at the larval instars (LO to L3) and PUP, until the emergence of the imago under laboratory conditions and inside the host. At the first instar, or LO, the microtype larvae measured less than 1 mm in length. During the transition from L1 to L3, the larvae grew in length. At L3, the larvae doubled their length (4 mm) and then started to develop both in length and width, reaching the PUP stage with 10 mm in length and 7 mm in width. The main characteristic that differentiates L3 from the early instars is the larger body size and the beginning of posterior spiracle development. The development from PUP to puparium took less than 24h. The bees died ten days after the fly oviposition, or just before full PUP development. The early development stages (egg-LO to L1) were critical for larva survival. The pupa was visible between the intersegmental sternites and, 32 days after pupation, a female imago of Physocephala sp. emerged from one bee. The puparium and the fly measured approximately 10 mm in length. In a single day of collection, up to 45% of the bumble bees collected were parasitized by conopid flies.

Guidelines for the use and interpretation of assays for monitoring autophagy.

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field.

Ultrastructure of the intramandibular gland of workers and queens of the stingless bee, Melipona quadrifasciata.

The intramandibular glands of workers and queens of Melipona quadrifasciata Lepeletier (Hymenoptera: Apidae), at different ages and from different functional groups, were studied using light and transmission electron microscopy. The results demonstrated that these glands are composed of two types of secretory structures: 1.A hypertrophied epidermis on the dorsal side of the mandible that is an epithelial gland. 2. Free secretory cells filling the inner spaces of the appendices that constitute a unicellular gland. The epithelial gland is larger in the young (1-2-day-old workers), and the gland becomes involuted during the nurse worker stage. The unicellular glands of the workers posses some secretion during all of the studied phases, but secretory activity is more intensive in the foraging workers. Vesicles of secretion are absent in the unicellular glands of queens. These results demonstrate that these glands show functional adaptations in different castes corresponding to the functions of each caste.

Autophagy and its physiological relevance in arthropods: current knowledge and perspectives.

Autophagic process is one of the best examples of a conserved mechanism of survival in eukaryotes. At the molecular level there are impressive similarities between unicellular and multicellular organisms, but there is increasing evidence that the same process may be used for different ends, i.e., survival or death, at least at cellular levels. Arthropods encompass a wide variety of invertebrates such as insects, crustaceans and spiders, and thus represent the taxon in which most of the investigations on autophagy in non-mammalian models are performed. The present review is focused on the genetic basis and the physiological meaning of the autophagic process on key models of arthropods. The involvement of autophagy in programmed cell death, especially during oogenesis and development, is also discussed.

Occurrence of apocrine secretion in the larval gut epithelial cells of Aedes aegypti L., Anopheles albitarsis Lynch-Arribálzaga and Culex quinquefasciatus say (Diptera: Culicidae): a defense strategy against infection by Bacillus sphaericus Neide?

Cell hypertrophy was the first reaction of the gut epithelial cells of Aedes aegypti (L.), Anopheles albitarsis (Lynch-Arribálzaga) and Culex quinquefasciatus (Say) to the toxins of strains 2362 and S1116 of Bacillus sphaericus, as cells had an increase of intracellular secretory activity. Soon after the cell hypertrophy developed, vesicles were formed at the cell apical portion, which detached with the plasma membrane, characterizing a type of apocrine secretion like. The first pathway of contamination of the mosquito larvae by these bacteria is through the gut, by feeding. Depending on the species of Culicidae and on the bacterial strain used, the hypertrophy of the gut epithelium occurred between 5 and 15 min after exposure to the contaminated environment. The second aspect observed after hypertrophy was the increase in apocrine secretion. The basophilic vesicles that detached from the cells remained between the peritrophic membrane and the gut lumen, such vesicles were filled with material of unknown nature. The gut posterior region showed secretory activity in both control and treated larvae, being much more intense in bacteria-exposed larvae. There were remarkable differences in the epithelial cell reaction according to the toxins of the two bacterial strains, but C.quinquefasciatus was the most susceptible. Differences in the gut cell reactions to the toxins produced by the two bacterial strains are discussed.

Occurrence of apocrine secretion in the larval gut epithelial cells of Aedes aegypti L., Anopheles albitarsis Lynch-Arribálzaga and Culex quinquefasciatus say (Diptera: Culicidae): a defense strategy against infection by Bacillus sphaericus Neide? / Ocorrência de secreção apócrina nas células epiteliais do intestino de larvas de Aedes aegypti L., Anopheles albitarsis Lynch-Arribálzaga e Culex quinquefasciatus say (Diptera: Culicidae): um mecanismo de defesa contra infecção causada por Bacillus sphaericus Neide?

Cell hypertrophy was the first reaction of the gut epithelial cells of Aedes aegypti (L.), Anopheles albitarsis (Lynch-Arribálzaga) and Culex quinquefasciatus (Say) to the toxins of strains 2362 and S1116 of Bacillus sphaericus, as cells had an increase of intracellular secretory activity. Soon after the cell hypertrophy developed, vesicles were formed at the cell apical portion, which detached with the plasma membrane, characterizing a type of apocrine secretion like. The first pathway of contamination of the mosquito larvae by these bacteria is through the gut, by feeding. Depending on the species of Culicidae and on the bacterial strain used, the hypertrophy of the gut epithelium occurred between 5 and 15 min after exposure to the contaminated environment. The second aspect observed after hypertrophy was the increase in apocrine secretion. The basophilic vesicles that detached from the cells remained between the peritrophic membrane and the gut lumen, such vesicles were filled with material of unknown nature. The gut posterior region showed secretory activity in both control and treated larvae, being much more intense in bacteria-exposed larvae. There were remarkable differences in the epithelial cell reaction according to the toxins of the two bacterial strains, but C.quinquefasciatus was the most susceptible. Differences in the gut cell reactions to the toxins produced by the two bacterial strains are discussed.

Observou-se hipertrofia das células do epitélio intestinal de Aedes aegypti (L.), Anopheles albitarsis (Lynch-Arribálzaga) e Culex quinquefasciatus (Say) devido ao aumento da atividade secretora intracelular como primeira reação às toxinas das linhagens 2362 e S1116 de Bacillus sphaericus (Neide). Após a hipertrofia epitelial, formaram-se vesículas na porção apical das células, as quais eram compostas de fragmentos de membrana plasmática contendo material de natureza desconhecida em seu interior, caracterizando um tipo de secreção apócrina. A via de contaminação das larvas dos mosquitos por essas bactérias é pelo intestino, através da alimentação. Dependendo da espécie de Culicidade e da linhagem bacteriana utilizada, a hipertrofia do epitélio ocorreu entre 5 a 15 min após a exposição das larvas ao meio contaminado. O segundo aspecto observado no processo de contaminação foi o aumento da atividade de secreção apócrina. As vesículas basófilas que se desprendiam das células permaneciam entre a membrana peritrófica e o lúmen intestinal. Observou-se atividade secretora tanto no grupo controle como no experimental, porém muito mais intensa no grupo experimental. Os Culicidae estudados apresentaram diferenças marcantes nas respostas às toxinas das bactérias utilizadas, sendo C. quinquefasciatus a espécie mais suscetível. As diferenças de reações das células em relação às toxinas produzidas pelas duas linhagens bacterianas são discutidas.

Tegumentary epithelial glands in the abdomen of virgin and physogastric queens of the stingless bee Scaptotrigona postica Latreille (Meliponini: Trigonina)

In the eusocial worker bees, portions of the epithelium of the abdominal epidermis may become hypertrophied and glandular, and produce the wax used to build the brood alveoli. In meliponine bees, the presence of similar epithelium was already observed in the queens, but its function is still unknown. Would this epithelium have homologous function in the workers? May the queens display other functions, besides reproduction, related to those considered exclusive of workers? In order to contribute to solve these questions, a histological and ultrastructural study of the tegumentary epithelial glands of the third tergite was carried out in virgins and physogastric queens of Scaptotrigona postica Latreille. The glandular tegumentary epithelium was easily discernible in the third tergite of the queens, being much more developed in physogastric than in virgin queens. The ultrastructural differences in the smooth endoplasmic reticulum between the queens, additionally to the histological and morphometrical data, indicate that this epithelium may have functional differences between the queen types. As already known, the virgin queens may produce wax and build brood alveoli, but the role of this epithelium in physogastric queens is totally unknown. A hypothesis considered in the present study is that the epithelium may be involved in production of pheromones, which may play roles in the social interaction of the laying queen.

Nas operárias das abelhas eussociais, regiões do epitélio tegumentar do abdome podem hipertrofiar-se e tornarem-se glandulares, sendo responsáveis pela produção de cera usada na contrução dos alvéolos de cria. Nos meliponíneos, epitélio com essas características também foi observado nas rainhas, porém sua função ainda continua desconhecida. Teria esse epitélio função homóloga nas operárias? As rainhas, além da função reprodutiva, desempenhariam funções na colônia até então consideradas exclusivas das operárias? Para tentar colaborar no esclarecimento dessas questões realizou-se um estudo histológico e ultra-estrutural das glândulas epiteliais tegumentares do terceiro tergito em rainhas virgens e fisogástricas de Scaptotrigona postica Latreille. Os resultados morfológicos mostraram que o epitélio glandular existe e é facilmente discernível no terceiro tergito das rainhas, sendo muito mais desenvolvido nas fisogástricas do que nas virgens. A ultra-estrutura mostrou que há diferenças no tipo de organização do retículo endoplasmático liso entre as rainhas, o que, juntamente com os resultados da histologia e morfometria, indicam poder haver diferenças funcionais desse epitélio entre elas. Como já observado na literatura, nas rainhas virgens tal epitélio pode estar envolvido na produção de cera e confecção de alvéolos de cria, porém sua função nas rainhas fisogástricas é totalmente desconhecida. A hipótese lançada no presente estudo é de que esse epitélio pode estar envolvido na produção de feromônios que irão atuar nas interações sociais da rainha poedeira.

Ocorrência, morfologia e ultra-estrutura da glândula de Dufour de Scaptotrigona postica Latreille (Hymenoptera: Apidae) / Dufour gland occurence, morphology and ultrastructure in Scaptogrigona postica Latreille (Hymenoptera: Apidae)

A glândula de Dufour é uma glândula acessória do aparelho reprodutivo feminino das abelhas. Nas abelhas neotropicais sem ferrão, tem sido pouca estudada sob todos os aspectos: morfológico, ontogenético e bioquímico. Na tentativa de colaborar com o conhecimento dessa glândula em abelhas sem ferrão, foi realizado um estudo da sua ocorrência, morfologia e desenvolvimento em Scaptotrigona postica Latreille. Os resultados mostraram que ela se encontra ausente nas operárias, como ocorre em muitas outras espécies desse grupo. Nas rainhas, as células glandulares parecem mais ativas nas virgens, possuindo uma desenvolvida rede de retículo endoplasmático liso tubular, grânulos de secreção e polirribossomos dispersos no citoplasma, além de apresentarem núcleos maiores do que os das células glandulares das fisogástricas. Nas rainhas fisogástricas há dois tipos de células glandulares, ambas aparentemente inativas sinteticamente. As glândulas das rainhas fisogástricas são claramente capazes de captar substâncias da hemolinfa, provavelmente lipídios, que não penetram nas células, mas passam pelos espaços intercelulares e, através da cutícula, chegam diretamente à luz da glândula. A bem desenvolvida dupla camada de lâmina basal ao redor da glândula pode atuar no processo de captação de substâncias da hemolinfa. A secreção, e conseqüentemente sua função, pode ser diferente nas duas classes de rainhas.

The Dufour gland is an accessory gland of the bee reproductive female apparatus. In neotropical stingless bees, this gland has been poorly investigated, concerning to developmental, morphological or biochemical characteristics. In order to collaborate with the knowledge of the Dufour gland in stingless bees, a study of its occurrence, morphology and development was performed in Scapatotrigona postica Latreille. The results showed that this gland is absent in workers, although it occurs in workers of many stingless bee species. The glandular cells seem to be very active in virgin queens, presenting a well developed tubular network of smooth endoplasmic reticulum, secretion granules and some polyribosomes in the glandular cell cytoplasm, besides larger nuclei than in physogastric queens. In physogastric queen glands there are two types of cells, both of them apparently not synthetically active. Nevertheless, the physogastric queen glands are clearly able to absorb substances from the haemolymph, probably lipids that do not cross the cells, but pass through the intercellular cell spaces and cuticle, and are released directly to the gland lumen. The very developed double-layered gland basal lamina may actuate in the process of substances uptaking. The secretion, and consequently its function, can be different in these two queen types.

Caracterização das glândulas mandibulares nas diferentes classes de adultos de Scaptotrigona postica Latreille (Hymenoptera: Apidae) / Characterization of the mandibular glands in different adult types of Scaptotrigona postica Latreille (Hymenoptera: Apidae)

As glândulas mandibulares de operárias recém-emergidas, nutridoras e campeiras, rainhas virgens e fisogástricas, e machos maduros de Scaptotrigona postica Latreille foram estudadas histológica e morfometricamente. Estas glândulas apresentam diferenças histológicas características para cada idade e/ou função do indivíduo. De acordo com a morfometria conclui-se que as glândulas mandibulares possuem um ciclo secretor determinado, no qual a secreção começa a ser produzida nas operárias recém-emergidas, e acumula-se no citoplasma e reservatório da glândula das operárias nutridoras. Nas operárias campeiras o citoplasma encontra-se bastante vacuolizado, o que foi interpretado como resultado da eliminação da secreção. Rainhas fisogástricas apresentaram as maiores áreas celulares e nucleares, ao oposto dos machos, os quais apresentaram as menores áreas.

The mandibular glands of newly-emerged, nurse and forager workers, virgin and physogastric queens, and mature males of Scaptotrigona postica Latreille were studied under histological and morphometrical analyses. These glands present differences in their histology, according to the age and/or function of the individuals. The morphometric results showed that the mandibular glands have a very markedly secretory cicle, in which the secretion starts to be produced in newly emerged workers, being accumulated into the cytoplasm and reservoir in the nurses. In forager workers the cytoplasm presents many vacuoles, which are remaining of the secretion releasing. Physogastric queens showed the largest cellular and nuclear areas. On the contrary, the males presented the smallest ones.

Comparative anatomy of the male reproductive internal organs of 51 species of bees / Anatomia comparada dos órgãos do aparelho reprodutor masculino de 51 espécies de abelhas

A anatomia dos órgãos internos do aparelho reprodutor de machos (ARM) adultos e pupas foi comparada em 51 espécies de abelhas, incluindo representantes de seis famílias. Foram obtidos quatro tipos diferentes de ARM. O tipo I está presente em machos das famílias mais basais (Colletidae,Andrenidae e Halictidae) e é caracterizado por três túbulos seminíferos por testículo, o qual é quasetotalmente envolvido pela membrana escrotal. O tipo II é um tipo intermediário entre os tipos I e III e estápresente em Mellitidae e Megachilidae, como também em alguns Apidae estudados, sendo caracterizado por possuir dutos deferentes pós-vesiculares fora da membrana escrotal e possuir três ou quatro túbulos seminíferos por testículo, exceto Apis mellifera L., a qual possui secundariamente um número aumentado de túbulos. O tipo III foi achado somente nos Apidae estudados e é caracterizado por apresentar os testículos e dutos genitais (exceto o duto deferente pós-vesicular) encapsulados separadamente, as glândulas acessórias são bem desenvolvidas e o duto ejaculador é calibroso, apresentando fissuras em sua parede externa, as quais podem ocorrer também no tipo II. O tipo IV está presente exclusivamente na tribo Meliponini e é caracterizado pela ausência de glândulas acessórias.

The anatomy of the internal organs of the male reproductive apparatus (MRA) of adults and pupae was compared among 51 species of bees, including representatives of six families. Fourdifferent types of MRA were found. The type I is present in males of the less derived families (Colletidae, Andrenidae, and Halictidae) and is characterized by three seminiferous tubules per testis, which are almost completely enveloped by the scrotal membrane. The type II is an intermediary between types III and I and is present in Melittidae and Megachilidae, as well as in some Apidae studied, being characterized by post-vesicular deferent ducts outside the scrotal membrane and by three or fourseminiferous tubules per testis, except for Apis mellifera L., which has a secondarily increased number of tubules. Type III was only found in the Apidae studied and is characterized by separately encapsulated testes and genital ducts (except for the post-vesicular deferent duct). Accessory glands are well developed, and the ejaculatory duct is thickened with fissures in its wall, which may also occur in the type II. Type IV is present exclusively in the tribe Meliponini, and is characterized by the absence of accessory glands.

Comparative study of the cuticular hydrocarbon composition of Melipona bicolor Lepeletier, 1836 (Hymenoptera, Meliponini) workers and queens.

In social insects, cuticular hydrocarbons are involved in species, kin, caste and nestmate recognition. Gas chromatography and mass spectrometry were used to compare the cuticular hydrocarbon composition of workers, males and queens of Melipona bicolor. The cuticular hydrocarbon composition of this species was found to consist mainly of C23, C25:1, C25, C27:1, C27, C29:1 and C29, which are already present in imagoes that have not yet abandoned the brood cell. This composition varied quantitatively and qualitatively between and within the castes and sexes. The newly emerged workers and young queens (virgins) had similar cuticular hydrocarbon profiles, which were different from those of the males. When the females start executing their tasks in the colony, the cuticular hydrocarbon profile differences appear. The workers have less variety, while the queens conserve or increase the number of cuticular hydrocarbon compounds. The queens have more abdominal tegumentary glands than the workers, which apparently are the source of the new cuticular compounds.