José Lino-Neto

Ultrastructural characteristics of the spermatozoa of Scelionidae (Hymenoptera, Platygastroidea) with phylogenetic considerations

The Scelionidae sperm are distinguished from those of all hymenopterans already studied at least by the presence of a single mitochondrial derivative and the absence of a centriolar adjunct. The absence of an acrosome, in Telenomus podisi, is also unique. The helical nucleus and mitochondrial derivative spiralling around a twisted axoneme can be considered as synapomorphies shared with the Chalcidoidea, and the mitochondrial derivative running together with the nucleus for a long distance can be considered as a synapomorphy shared with the Diapriidae. Therefore, from a consideration of these features, it is possible to suppose that the Scelionidae, Chalcidoidea and Diapriidae are more closely related between themselves than are any of them to the Cynipoidea, since the latter does not share any of the above‐mentioned features. This supposition agrees with phylogenetic analyses that supported the inclusion of Platygastroidea (Scelionidae and Platygastridae) and Chalcidoidea within the Proctotrupomorpha lineage, as well as the close relationship of these to the Diapriidae, and the exclusion of the Cynipoidea from this lineage.

Sperm structure and ultrastructure of the fire ant Solenopsis invicta (Buren) (Hymenoptera, Formicidae).

The spermatozoa of Solenopsis invicta are long and slender, measuring about 70 microm in length, of which the head region measures approximately 13 microm. The head consists of an acrosome formed by an acrosomal vesicle covering a perforatorium and a nucleus. This latter measures about 12 microm, has compact chromatin, and many translucent areas. The flagellum consists of an axoneme of 9+9+2 microtubules that begins just below the nuclear base. The two mitochondrial derivatives are of almost equal diameter, and there are two accessory bodies. The nucleus is attached to the flagellum by a centriolar adjunct located between the baso-lateral region of the nucleus and the anterior mitochondrial derivative extremities. Therefore, the nucleus-flagellum transition region of the spermatozoa of S. invicta is very similar to that of other ants and differs from the first description of this species [Ann. Entomol. Soc. Am. 67 (1967) 632]. In relation to other aculeates, it is possible that three characteristics (type of centriolar adjunct, presence of paracrystalline areas in each mitochondrial derivative, and the position of these areas within each derivative) may be autopomorphies for the Formicidae.

Ultrastructure and heteromorphism of spermatozoa in five species of bugs (Pentatomidae: Heteroptera)

Pentatomidae is one of the largest Heteroptera families, comprising about 10% of the species estimated for the suborder. In spite of existing studies, doubts remain regarding the systematics of Pentatomomorpha. In this study, five species of Pentatomidae spermatozoa were examined to achieve characteristics that enable inferences in the phylogeny of the group and in behavioral issues associated with the presence of polymorphisms. Spermatozoa polymorphisms, characterized by two classes of sizes, are found in Podisus nigrispinus, Podisus distinctus, and Brontocoris tabidus, whereas Thynacanta marginata (Dallas) and Supputius cincticeps have single-size spermatozoa. The head region consists of an acrosome, a nucleus, and part of the centriolar adjunct. In the more anterior region, the nucleus is parallel to the centriolar adjunct. In the nucleus-flagellum transition region, the nucleus overlaps the anterior region of the mitochondrial derivatives, just above the axoneme. The mitochondrial derivatives and the axoneme run the entire extent of the flagellum. In species in which the spermatozoa are polymorphic, the larger spermatozoa have derivatives mitochondrial approximately 3-fold larger than the smaller spermatozoa. Characteristics derived from the morphology of spermatozoa indicate synapomorphies and are promising for systematic studies.

Ultrastructural characterization of the spermatozoa of Aethalion reticulatum Linnaeus 1767 (Hemiptera: Auchenorrhyncha: Aethalionidae).

The Hemiptera order is currently divided into four suborders. Among them the Auchenorrhyncha suborder is considered to be paraphyletic. Morphology of insect spermatozoa has provided promising characteristics that can be used for phylogenetic inference. In this study, Aethalion reticulatum (Aethalionidae) spermatozoa were examined by light and electron microscopy. The head of the spermatozoa is composed of an acrosome and a nucleus. The nucleus is linear and filled with compact chromatin but has electron-lucid spaces. The centriole adjunct initiates parallel to the nucleus and terminates in the region anterior to the mitochondrial derivatives. Flagella consist of an axoneme, two mitochondrial derivatives and two accessory bodies and the axoneme has the typical 9+9+2 microtubule pattern. The mitochondrial derivatives are symmetric. The accessory bodies are long and are shaped like a half moon when viewed from a cross-section. The presence of accessory bodies differs from other species of Cicadomorpha previously studied. Spermatozoa morphology of other Auchenorrhyncha families can reveal synapomorphies and contribute to systematics of the suborder.

Sperm ultrastructure of the European hornet Vespa crabro (Linnaeus, 1758) (Hymenoptera: Vespidae)

This study represents the first sperm description of a Vespinae species (Vespa crabro). The acrosome consists of an acrosomal vesicle and a perforatorium. The nucleus has compact chromatin and shows lenticular structures on the nuclear envelope. These structures, which have never been observed in a hymenopteran sperm, could be clusters of nuclear pores. The centriolar adjunct has an asymmetric pattern and shows a structured periphery. The centriole consists of 9 accessory microtubules and 9 doublet microtubules devoid of arms and spokes. The axoneme has a 9+9+2 microtubule pattern and the accessory microtubules have 16 protofilaments. The mitochondrial derivatives differ in length and diameter. The larger one is adjacent to the nuclear base, while the smaller one begins below the centriolar adjunct. They possess three distinct areas and a large paracrystalline region, which occurs only in the large one. The large mitochondrial derivative ends first, followed by the small one. The axoneme gradually disorganizes: first the central microtubules disappear, then the doublets, which show opened B-tubules, and finally the accessory microtubules. The sperm morphology of V. crabro is very similar to that of the polistine wasp, Agelaia vicina. This can indicate that, in Vespidae, sperm morphology is maintained without important variations among subfamilies and/or that this similarity indicates close phylogenetic relationship between these two subfamilies. Although Vespidae phylogenetically related to Formicidae, these data suggest that the former more closely related to Apoidea than to Formicidae.

Structure and ultrastructure of the spermatozoa of Trypoxylon (Trypargilum) albitarse Fabricius 1804 (Hymenoptera: Apoidea: Crabronidae).

The ultrastructure of the spermatozoa of Trypoxylon (Trypargilum) albitarse is described here for the first time within this genus. Testes and seminal vesicles were dissected and processed for transmission electron microscopy. In the testicular follicles, the spermatids are arranged in a maximum number of 32 for each cyst. The spermatozoa are slender and they measure approximately 150 microm in length. The head is about 17 microm long and is formed by the acrosome and the nucleus. The flagellum consists in an axoneme, two mitochondrial derivatives, two accessory bodies and, at the nucleus-flagellum transition, a symmetric centriolar adjunct. The axoneme presents the typical 9+9+2 microtubule pattern. In the terminal region, the central microtubules and nine doublets finish first, followed by the accessory microtubules. Both mitochondrial derivatives begin together and are inserted in the base of the centriolar adjunct. Along the middle region, the larger derivative has almost twice the area of the smaller one and includes a discrete paracrystalline region. At the tip, the smaller derivative ends before the larger one and both before the axoneme. The characters derived from the ultrastructure of the spermatozoa of T. albitarse show synapomorphies shared with the Apoidea and present characters that are probably apomorphic for the Crabroninae subfamily.

Structural and ultrastructural characterization of male reproductive tracts and spermatozoa in fig wasps of the genus Pegoscapus (Hymenoptera, Chalcidoidea)

The three Pegoscapus species present the same internal reproductive tract features comprising testes with a single testicular tubule, seminal vesicles, vasa deferentia, accessory glands and an ejaculatory duct. The seminal vesicle shows two morphologically distinct portions although they do not resemble the separate chambers found in other Chalcidoidea. The anterior portion of the seminal vesicle shows a prominent epithelium and stores the mature spermatozoa, while the posterior region is formed by a thicker muscular sheath that participates on ejaculation. The sexual maturation in Pegoscapus is achieved at emergence, when the testicular degeneration occurs. The spermatozoa of Pegoscapus reveal a basic structure similar to that of other Chalcidoidea. In Pegoscapus sp1. and Pegoscapus sp2. they present the same features, whereas Pegoscapus tonduzi comprises some different characteristics. It measures approximately 160 microm in Pegoscapus sp1. and Pegoscapus sp2., while in P. tonduzi the spermatozoa measure about 360 microm. The extracellular sheath thickness is another difference among the species. While Pegoscapus sp1. and Pegoscapus sp2. show a thick extracellular sheath, in P. tonduzi this sheath is very thin resulting in a large space intervening between the extracellular sheath and the nucleus. Despite these differences, the three species analyzed share some characteristics that allow the establishment of an identity to the spermatozoon of the genus Pegoscapus: the seminal vesicle not divided in chambers; the absence of acrosomal structures in the spermatozoa; the length of the extracellular sheath; the central microtubules being the firsts to terminate in the sequence of microtubular cutoff at the final axonemal portion.

Sexual Success after Stress? Imidacloprid-Induced Hormesis in Males of the Neotropical Stink Bug Euschistus heros

Environmental stress in newly-emerged adult insects can have dramatic consequences on their life traits (e.g., dispersion, survival and reproduction) as adults. For instance, insects sublethally exposed to environmental stressors (e.g., insecticides) can gain fitness benefits as a result of hormesis (i.e., benefits of low doses of compounds that would be toxic at higher doses). Here, we experimentally tested whether sublethal exposure to the insecticide imidacloprid would hormetically affect the sexual fitness of newly-emerged adults of the Neotropical brown stink bug Euschistus heros (Hemiptera: Heteroptera: Pentatomidae), which is the most abundant and prevalent insect pest in Neotropical soybean fields. We evaluated the sexual fitness of four couple combinations: unexposed couples, exposed females, exposed males, and exposed couples. Sublethal exposure to dry residues (i.e., contact) of imidacloprid (at 1% of recommended field rate) did not affect insect survival, but led to higher mating frequencies when at least one member of the couple was exposed. However, the average mating duration was shortened when only females were exposed to imidacloprid. Moreover, exposed males showed higher locomotory (walking) activity, lower respiration rates and induced higher fecundity rates when mated to unexposed females. Although the reproductive tracts of exposed males did not differ morphometrically from unexposed males, their accessory glands exhibited positive reactions for acidic and basic contents. Our findings suggest that males of the Neotropical brown stink bug hormetically increase their sexual fitness when cued by impending insecticidal stress in early adulthood.

Structural and ultrastructural characteristics of male reproductive tract and spermatozoa in two Cryptinae species (Hymenoptera: Ichneumonidae)

This study represents the first characterization of male reproductive tracts and ultrastructural description of sperm of Cryptinae species. In Lymeon dieloceri and Pachysomoides sp., the male reproductive tract is formed by a pair of testis, two deferent ducts, two accessory glands and one ejaculatory duct. The spermatozoa are similar to those described for other Hymenoptera, with: (1) the acrosome formed by the acrosomal vesicle covering the perforatorium, which has its base inserted in a cavity located in the nucleus point; (2) a thin nucleus with electron-dense chromatin; (3) an electron-dense centriolar adjunct located between the nucleus and one of the two mitochondrial derivatives; (4) an axoneme with a 9 + 9 + 2 microtubule arrangement; (5) two long mitochondrial derivatives with peripheral cristae and; (6) two accessory bodies located between the two mitochondrial derivatives and the axoneme. These ichneumonids present structural characteristics similar to other parasitic wasps, such as presence of a single follicle per testis, layer of extracellular material enveloping the acrosome and accessory microtubules ending before others in final portion of the flagellum. However, male reproductive system and the spermatozoa presented morphological characteristics that allowed their differentiation, such as oval shapes accessory glands and the symmetric mitochondrial derivatives in L. dieloceri compared to the spherical accessory glands and asymmetrical derivatives observed in Pachysomoides sp. Taken together, data presented here demonstrates that diversity of morphological characteristic from the male reproductive tract and spermatozoa in Hymenoptera might provide a character system that can be used, in association with other systems, to resolve various uncertainties about the evolutionary relationships of this insect group.

Megaplatypus mutatus (Chapuis) (Coleoptera: Curculionidae: Platypodinae) Attacks Hybrid Eucalyptus L'Héritier De Brutelle Clones In Southern Espírito Santo, Brazil

The cultivation of Eucalyptus L’Héritier de Brutelle in Brazil is directly and indirectly responsible for a great number of jobs and reduces the need for cutting native forest. Species of Hymenoptera (Zanuncio et al. 2002), Lepidoptera (Zanuncio et al. 1993; Pereira et al. 2008) and Coleoptera (Zanuncio et al. 2005, 2009; Garlet et al. 2009) are the main groups of insect pests of Eucalyptus. The important Coleoptera are polyphagous and perforate the sapwood in plantations intended for the production of wood (Wood and Bright 1987; Flechtmann et al. 2001; Kirkendall et al. 2008). Platypodinae were found in two clonal plantations of Eucalyptus (Eucalyptus urophylla S.T. Blake × Eucalyptus camaldulensis Dehn. and E. urophylla × Eucalyptus grandis W. Hill ex Maid. hybrid) in the municipality of Venda Nova do Imigrante, Espírito Santo, Brazil. These areas (Figs. 1A and B) were established in January 2002 and visited and photographed with a Sony Cyber-Shot digital camera (model DSC-W70 with 7.2 megapixels) in January 2009. Some trees with the presence of platypodine tunnels were felled and wood discs of 5 cm × 15–25 cm (height × diameter) were obtained with a chain saw. Discs were sent to the Laboratory of Biological Control of the Department of Animal Biology, Federal University de Viçosa in Viçosa, Minas Gerais, Brazil where they were analyzed. The holes in the wood discs were opened by hammer and chisel to determine the diameter, length, and direction of the galleries, and presence of platypodine adults. Some adult specimens were killed by the sapwood and remained stuck to the Eucalyptus bark without penetrating it. Adults were collected and preserved in 70% alcohol, mounted, photographed in macro mode with a digital camera (Nikon SLR, model D200) coupled to a stereomicroscope (Olympus, model SZH10), and sent to L. R. Kirkendall (University Bergen, Norway) for identification. The species collected was identified as Megaplatypus mutatus (Chapuis, 1865) (Coleoptera: Curculionidae: Platypodinae) (Figs. 1F, G, H, and I). A male and two females were deposited in the Museum of the Department of Biology, University of Bergen, Bergen, Norway. Problems associated with M. mutatus were represented by holes in the stems of Eucalyptus trees up to a height of 3.0 m above the soil (Figs. 1C, D, and E). Adults bored from the Eucalyptus sapwood to the heartwood with galleries in several directions, but always toward the center of the trunk, as done by other ambrosia beetles (Shore et al. 1987; Morales et al. 2000). The linear length of the galleries ranged from 3 to 15 cm and all of them had symbiotic fungal growth. All of the galleries except one had no signs of larvae or adults of M. mutatus. Megaplatypus mutatus is a highly polyphagous species native to South America (Girardi et al. 2006), breeding in a wide variety of native and exotic hosts (Giménez and Etiennot 2003). It is one of only a handful of platypodine or scolytine ambrosia beetle species capable of colonizing living trees (Kirkendall 2006). It attacks living trees with a diameter at breast height greater than 15 cm and forms deep galleries in a transverse plane of the xylem, reducing the commercial value of the wood (Bascialli et al. 1996; Alfaro et al. 2007). This species has also been reported from Argentina, Bolivia, Brazil, Guyana, Paraguay, Peru, Uruguay, and Venezuela in more than 30 genera of woody plants, including some of commercial importance (Giménez and Etiennot 2003; Giménez and Panzardi 2003). Megaplatypus mutatus was reported in the province of Caserta, Naples, Italy, damaging plants of Populus L. spp. (Tremblay et al. 2000), and its populations there are being monitored to prevent the spread of the pest (Alfaro et al. 2007). Megaplatypus mutatus has been reported in Brazil from Acacia meanmsii De Wild. (Santana and Santos 2001), Erythrina crista-galli L. (Giménez and Etiennot 2003), and Caesalpinia echinata Lam (Girardi et al. 2006), but our observation is the first report of this species in clonal Eucalyptus plantations used for sawlogs in Espírito Santo. The economic value of these cultivars and the difficulty of controlling this insect, which penetrates tree stems, requires monitoring with traps made of cotton soaked in 20% ethanol (Audino et al. 2005; Girardi et al. 2006) or sex pheromone (Liguori et al. 2008) combined with search control tactics. Relatively little is known about M. mutatus beyond the damage it causes; knowledge of natural enemies, their efficiency, and possibility of application and methods of controlling this pest have not yet been studied. The occurrence ofM.mutatus in Eucalyptus plantations demonstrates the necessity of studies