Sewage sludge toxicity in edaphic organism: analysis of midgut responses in the diplopod Rhinocricus padbergi

Diplopods are agile saprophagous invertebrates with the ability to colonize several soil layers. They feed on waste and decaying organic matter, making the soil in which they are present better aerated and mineral‐enriched. Because of their habits, diplopods have been suggested to be used in studies to analyze possible toxic effects of impacted soils in edafic fauna. In that respect, this study aimed at checking the feasibility of using morphological analysis from millipede's midgut as a biomarker for soils exposed to sewage sludge from sewage treatment plants. Histological and histochemical techniques, applied to the species Rhinocricus padbergi's midgut, were used. Two sewage sludge samples were used; they were collected in two small towns in the countryside of São Paulo State. Both cities are part of the Piracicaba–Capivari–Jundiaí basin (PCJ‐1 and PCJ‐3). For 7 and 15‐day periods, the animals were exposed to 10% concentrations and crude sludge for the PCJ‐1sample and to 1, 10, and 50% concentrations and sludge for the PCJ‐3 sample. The material exposed to the PCJ‐1 sample showed an increase in the presence of cytoplasmic granules in both concentrations in all periods and a slight increase in the rate of epithelial renewal. The material of PCJ‐3 sample presented the same alterations observed in PCJ‐1 sample in higher frequence and over that high quantity of neutral polysaccharides in the hepatic cells layer. We can conclude that with the increase of sewage sludge concentrations tested, there is an increase in morphophysiologicals alterations frequencies found in the diplopods' midgut. Microsc. Res. Tech. 2012. © 2012 Wiley Periodicals, Inc.     

Morphological characterization of the nymphs Rhipicephalus sanguineus ticks (Latreille, 1806) (Acari: Ixodidae). Description of the testes, integument, Malpighian tubules, and midgut on the detachment day

This study presents the morpho-histological and histochemical characterization of the testes, integument, Malpighian tubules, and midgut of engorged Rhipicephalus sanguineus nymphs on the detachment day, showing the morphological and physiological characteristics to this phase in the life cycle of these individuals. The testis is constituted by germinative cells (only spermatogonia) with large, round-shaped and strongly stained nuclei which are organized into cysts by a thin layer of somatic cells. The integument consists of a cuticle subdivided into epicuticle (lipoprotein) and procuticle (glycoproteic), and a layer of epithelial cells which present glycolipoprotein elements. The procuticle presents two distinct regions: the exocuticle (next to the epicuticle) and the endocuticle (next to the epithelial layer). The Malpighian tubules present a simple epithelium with small flat and/or cubic cells, which form its wall and delimitates a lumen full of lipoprotein material. The midgut consists of an epithelial wall formed by two types of digestive cells, spent cells and empty digest cells, and by generative cells supported by a basal lamina and a thin layer of muscular tissue. This study described the main organs of engorged nymphs of R. sanguineus, to generate information that can help researchers to better understand the biology of these ectoparasites; which is fundamental for the development of compounds that are less aggressive to the environment. In addition, if the immature stages of the ticks are controlled, the number of adult ticks able to cause damages to the animals--and to the man as well--is also under control.     

Ultrastructure of male reproductive system of Eurydema ventrale Kolenati 1846 (Heteroptera: Pentatomidae)

The male reproductive system of Eurydema ventrale Kolenati 1846 is studied morphologically and histologically by using light, scanning and transmission electron microscopes. The reproductive system of the male E.ventrale consists of a pair of testis, a pair of vas deferens, a pair of seminal vesicles, accessory glands, a bulbus ejaculatorius, a pair of ectodermal sacs, and a ductus ejaculatorius. The testicular follicles have three different development zones (growth zone, maturation zone, and differentiation zone). The testes are connected to the seminal vesicles by the vas deferens that is a specialized in sperm storage. Sperm have an elongated head and a tail (flagellum) with an axonema and two mitochondrial derivatives. Vas deferens and seminal vesicles are fine, long, and cylindrical. The seminal vesicle is connected with bulbus ejaculatorius, which is balloon shaped and surrounded with accessory glands. The bulbus ejaculatorius is continuous with ductus ejaculatorius which is connected to the aedeagus. Microsc. Res. Tech. 78:643–653, 2015. © 2015 Wiley Periodicals, Inc.     

Diversity of the vomeronasal system in mammals: the singularities of the sheep model

The enormous morphological diversity and heterogeneity of the vomeronasal system (VNS) in mammals--as well as its complete absence in some cases--complicates the extrapolation of data from one species to another, making any physiological and functional conclusions valid for the whole Mammalian Class difficult and risky to draw. Some highly-evolved macrosmatic mammals, like sheep, have been previously used in interesting behavioral studies concerning the main and accessory olfactory systems. However, in this species, certain crucial morphological peculiarities have not until now been considered. Following histological, histochemical and immunohistochemical procedures, we have studied the vomeronasal organ (VNO) and the accessory olfactory bulb (AOB) of adult sheep. We have determined: (1) that all structures which classically define the VNO in mammals are present and well developed, providing the morphological basis for functional activity. (2) that, conversely, there is only a scant population of scattered mitral/tufted cells. One morphological consequence of both details is that the strata of the AOB in adult sheep are not as sharply defined as in other species; moreover, the small number of the mitral/tufted cells in the AOB may imply that the VNS of adult sheep is not capable of functioning in the way a well-developed VNS does in other species. (3) the zone to zone projection from the apical and basal sensory epithelium of the VNO to the anterior and posterior part of the AOB, respectively, typical in rodents, lagomorphs and marsupials, is not present in adult sheep.     

Descriptive studies on the tongue of two micro‐mammals inhabiting the Egyptian fauna; the Nile grass rat (Arvicanthis niloticus) and the Egyptian long‐eared hedgehog (Hemiechinus auritus)

The current study aimed to describe the anatomical features of the tongues of two micro‐mammals common in the Egyptian fauna; the Nile grass rat (Arvicathis niloticus), and the Egyptian long‐eared hedgehog (Hemiechinus auritus). The tongues of five adult individuals of each species were excised and processed histologically, histochemically, and morphometrically. Statistical analysis comparing the relative tongue length in both species showed that there was a significant difference, which may correlate with the difference in feeding preferences. Grossly, the Nile grass rat has a dorsal lingual prominence with bifurcated apex while, the long‐eared hedgehog has a median slight elevation with rounded apex. Numerous forms of mechanical and gustatory papillae are scattered along the lingual dorsal epithelium. The histochemical detection of keratin by Holland's trichrome stain showed an intense expression in the case of A. niloticus and mild expression in H. auritus. The framework of the tongue (entoglossum) is supported by either a core of cartilage in H. auritus or bone in A. niloticus which incorporated in the lingual root. The lingual glands also showed marked variation, the Nile grass rat exhibit dense populations of mucous‐secreting glands and lesser populations of serous‐secreting glands, the contrary is true in H. auritus. In conclusion, the micro and macro‐anatomical features of the tongues of both species showed adaptive changes to accommodate the feeding lifestyle. Such type of studies using mammals from different phylogenetic traits and almost have different feeding preferences provide answers to many research questions related to tongue evolution among mammalian vertebrates.     

Convergence in dental histology between the late Triassic semionotiform Sargodon tomicus (Neopterygii) and a late cretaceous (Turonian) pycnodontid (Neopterygii: Pycnodontiformes) species

Microstructural scanning electron microscope investigation was performed on sectioned and surface‐etched isolated, prehensile teeth of the Late Triassic semionotiform species Sargodon tomicus and Pycnodontidae incertae sedis from the Late Cretaceous. The teeth of both taxa display a system of vascular canals penetrating the dentine and the overlying hypermineralized acrodin cap; small tubules are radiating at an angle to the long axis of the canals, interpreted as residual spaces left by odontoblast cell processes. This is the first detailed account of vascular acrodin encountered in a pycnodont species. New information is revealed also about Sargodon dental histology in the shape of mineralized remnants of the basal lamina at the acrodin–dentine junction. This implies that deposition of the acrodin organic matrix proceeded centrifugally by the cells of the inner dental epithelium, probably with minor collagen contribution from odontoblasts. This is contrary to the more typical centripetal formation (beneath the basal lamina) of the acrodin layer implied for the studied pycnodontid teeth. The rare occurrences of vascular acrodin within Actinopterygii, and the demonstrated differences in its histogenesis, do not suggest the usefulness of the tissue as systematic character but rather point to its adaptive significance. The superficial increase in the order of acrodin bundle orientation, observed in both species, is similarly regarded as convergently acquired mechanical adaptation. The observed uneven shape of crystallite rows and lesser degree of mineralization of the inner collariform ganoin, compared to its outer portion, is indicative of epithelial‐ectomesenchymal interaction and qualifies the tissue as enameloid. Microsc. Res. Tech., 2011. © 2010 Wiley‐Liss, Inc.     

Postembryonic development of hard jaws (trophi) in a species belonging to the Brachionus plicatilis complex (Rotifera, Monogononta): a morphometric analysis

The presence of hard jaws (trophi), with species-specific shape and size, is a taxonomic feature of Rotifera, a group of microscopic metazoans. Since trophi are used to discriminate among species, it is important to know whether these structures change in taxonomically important ways during postembryonic development. Using both SEM and optical images, we analyzed more than 100 individuals of a single clonal lineage of a monogonont rotifer, Brachionus plicatilis, in order to describe body and trophi development after hatching. Body size, expressed as lorica width and length, was isometrically related to age of the animals only during preadult stages. Trophi size, expressed as length of the different parts, was unrelated to either age or body size. Therefore, trophi elements do not grow after hatching in B. plicatilis. Despite the dimensional invariance with age, some differences in trophi size among individuals of the same clone were recorded. No difference in left-right asymmetry of the trophi was shown; the asymmetric elements of the trophi named rami consistently had the right ramus longer than the left. This constancy is in contrast to the reported trophi asymmetries in bdelloid rotifers, in which left-right asymmetries are not constant within clonal lineages. In conclusion, we suggest that also trophi size, constant within the analyzed clone, may be used as an additional taxonomic feature to help in the discrimination of taxa within the B. plicatilis complex of cryptic species.     

Structural study of the salivary glands of Anocentor nitens (Acari: Ixodidae) during the feeding cycle

The salivary glands of Anocentor nitens (Neumann, 1897 ) occur in pairs and are located in the anterolateral region of the general cavity, with milky white color and approximately equal sizes. They consist of a secretory portion and an excretion duct. In some glandular acini, all the cells had a basophilic appearance they were stained by hematoxylin, whereas others presented cells with different staining affinities. In this work, we describe the variations observed in these glands during the feeding cycle of ticks [after feeding (0 h) and successively at 24, 48, 72, 96, 120, and 144 h]. The cells stained by hematoxylin were shown to be more reactive to Alcian blue, thus demonstrating the presence of acid glycosaminoglycans, whereas those stained using eosin presented weak or no reaction. A strong reaction was found by the use of the periodic acid‐Schiff (PAS) technique, thereby suggesting the presence of glycogen and/or glycoconjugates containing hexose, confirmed by using salivary amylase before PAS, with partial destaining of the slides. Continuing presence of residual staining in these cells suggests the presence of glycoconjugates containing hexose. Cells with nuclei of circular outline and few granules (of different sizes) were found in type II acini, 72 h after collection. Type I acini presented wide lumina and walls composed of larger numbers of cells of cubic to cylindrical shape. The pronounced degranulation shown in this study over the course of the feeding cycle was associated with the release of substances for oviposition. Microsc. Res. Tech., 2009. © 2009 Wiley‐Liss, Inc.     

Sensilla and secretory glands in the antennae of a primitive ant: Dinoponera lucida (Formicidae: Ponerinae)

Morphology of the antennae of the female workers of the ponerine ant Dinoponera lucida was examined by light and scanning electron microscopy. In several antennomers, we found secretory gland cells of class I and III. Class III gland cells release their secretion through single pores in the antennal surface, whereas class I secretory cells are seen as tall epidermal cells close to the cuticle. Both gland types have weak reaction for total proteins and neutral polysaccharides. Six distinct sensilla types were observed: trichodea, chaetica, campaniform, basiconica, placodea, and coeloconica. The possible sensory functions of these sensilla and the gland functions are discussed.     

Ultrastructure of oocytes of the Urostreptus atrobrunneus (Diplopoda,Spirostreptida, Spirostreptidae): A potential urban centers plague

The knowledge of the process of egg formation is indispensable for understanding the mechanisms involved in the reproduction of different species. In this context, the objective of this work was to describe the ultrastructure of the oocytes of Urostreptus atrobrunneus (Spirostreptida), a potential plague of urban centers in different locations of São Paulo State. The lack of knowledge about the morphology, physiology, and the reproductive behavior of the species have hindered an effective control of it. The oocytes of U. atrobrunneus presented three development stages: young oocyte or type I; intermediary oocyte or type II; and mature oocyte or type III. During the oocyte development, the cytoplasm become filled with several globules of protein, drops of lipids, and sphaerocrystals, and it was not observed in many organelles in the oocytes with exception of mitochondria, abundant, principally in young oocytes. The vitelline membrane is also deposited in a discontinuous form and the chorion does not present differentiation of layers. The follicular epithelium alters its shape according to the development phase of the oocyte. Part of the vitellus is from exogenous origin and part is endogenous. Before this, only two studies about the ultrastructural analysis of the female germ cells of diplopods were published. Microsc. Res. Tech. © 2012 Wiley Periodicals, Inc.     

Structural and ultrastructural features of the Malpighian tubules of Dolycoris baccarum (Linnaeus 1758), (Heteroptera: Pentatomidae)

The morphology and ultrastructure of the Malpighian tubules of Dolycoris baccarum were analyzed by scanning (SEM) and transmission (TEM) electron microscopy in order to determine their functional organization. The Malpighian tubules are compared with similar structures of other insects based on cell structure and functional organization. The Malpighian tubules of D. baccarum extend from the midgut–hindgut region of the digestive tract. The Malpighian tubules are divided into two regions: the proximal segment is short and flattened and the distal segment is long, stringy in shape and free in hemolymph. The tubules are generally long and narrow. There is a large number of trachea around the tubules. They consist of a single layer of epithelial cells. It is observed in the TEM observation that the epithelial cells have numerous microvilli at the apical side of the cells. At the basal side of the cells, there is a great number of membrane foldings and mitochondria among them. Besides some spherites, mitochondria, lysosome‐like bodies, and large or small granules can be distinguished in the cells. With this study, we aimed to demonstrate the ultrastructure of the Malpighian tubules of D. baccarum and differences or similarities with other species.     

Cytotoxic effects of thiamethoxam in the midgut and malpighian tubules of Africanized Apis mellifera (Hymenoptera: Apidae)

Due to its expansion, agriculture has become increasingly dependent on the use of pesticides. However, the indiscriminate use of insecticides has had additional effects on the environment. These products have a broad spectrum of action, and therefore the insecticide affects not only the pests but also non‐target insects such as bees, which are important pollinators of agricultural crops and natural environments. Among the most used pesticides, the neonicotinoids are particularly harmful. One of the neonicotinoids of specific concern is thiamethoxam, which is used on a wide variety of crops and is toxic to bees. Thus, this study aimed to analyze the effects of this insecticide in the midgut and Malpighian tubule cells of Africanized Apis mellifera. Newly emerged workers were exposed until 8 days to a diet containing a sublethal dose of thiamethoxam equal to 1/10 of LC₅₀ (0.0428 ng a.i./l L of diet). The bees were dissected and the organs were processed for transmission electron microscopy. The results showed that thiamethoxam is cytotoxic to midgut and Malpighian tubules. In the midgut, the damage was more evident in bees exposed to the insecticide on the first day. On the eighth day, the cells were ultrastructurally intact suggesting a recovery of this organ. The Malpighian tubules showed pronounced alterations on the eighth day of exposure of bees to the insecticide. This study demonstrates that the continuous exposure to a sublethal dose of thiamethoxam can impair organs that are used during the metabolism of the insecticide. Microsc. Res. Tech. 77:274–281, 2014. © 2014 Wiley Periodicals, Inc.     

Morphology and distribution of antennal sensilla in a parasitoid fly,Gymnosoma rotundatum (Diptera: Tachinidae)

Morphology of antennal sensilla and their distribution were investigated in male and female adults of the parasitoid fly Gymnosoma rotundatum (Diptera: Tachinidae) using scanning electron microscopy (SEM). The overall length and shape were not different between males and females from each other. Three basic types of sensilla (sensilla basiconica, s. chaetica, and s. coeloconica) were identified from both sexes, but with variations in numbers and distribution along the antennae. The s. basiconica and s. chaetica could be divided further into subtypes; s. basiconica into three subtypes and s. chaetica into two subtypes. All the basiconica subtypes 1, 2, and 3 were multiporous, indicating that their primary function was olfactory. The sensilla basiconica was most abundant on the antennae of both sexes. The abundance of s. basiconica subtype 1 was different, but other subtypes 2 and 3 were similar between males and females. There was no pore on the cuticular surface of the s. chaetica and s. coeloconica, suggesting that they are likely to be a mechanosensory or a thermohygroreceptory function. The abundance of the two sensillum types was similar between males and females. The morphological information obtained in our study provides a basis for future investigations into the sensory physiological function, and associated behaviors, of each type of sensilla in this parasitoid fly.     

Snout cutis of the domestic pig (Sus scrofa domesticus,Linnaeus, 1758): Using light and transmission electron microscopy

The current study was designed to give microscopic view on the snout skin of the domestic pig (Sus scrofa domesticus) to clear its adaptations with the function of exploring for the food and pushing the objects. This study carried out on the snout skin of apparently healthy 1 year five pigs (Sus Scrofa) and examined under the light and transmission electron microscopy. Our results clarify that the snout skin cutis composed of the epidermis and the dermis. The epidermis consisted of stratum corneum, stratum granulosum, stratum spinosum, and stratum basale. The stratum corneum and the stratum spinosum appeared thicker than other parts. The dermis consisted of a reticular and a papillary dermis. For tightness junction between the dermis and the epidermis, the hemidesmosomes were observed, while the desmosomes were presented in abundant numbers at the level of stratum basale to ensure the adhesion between the keratinocytes. The merocrine sweat glands were observed in abundant numbers to provide the wetness of the snout to avoid its injury from friction during food exploring or pushing of the objects. We concluded that the adaptation of the snout skin with the environmental condition surrounding the studied domestic pig.     

Structure and distribution of antennal sensilla in Pseudosymmachia flavescens (Brenske) (Coleoptera: Scarabaeidae: Melolonthinae)

Morphology, microstructure, and distribution of antennal sensilla were compared between female and male Pseudosymmachia flavescens (Brenske) (Coleoptera: Scarabaeidae: Melolonthinae). Lamellate antennae of P. flavescens were shown to have typical scape, pedicel, and flagellum segments. The flagellum consists of a four-segmented funicle and a three-segmented club. The lengths of their pedicel and funicle were found to be similar in females and males. Distinct sexual differences were observed in the length of lamellar segments. Nine types/subtypes of sensilla were identified on the antennae of both sexes, including Böhm sensilla, sensilla trichodea, sensilla basiconica (SB), and two subtypes for sensilla chaetica, sensilla coeloconica, and sensilla placodea each. Olfactory sensilla (e.g., SB and placodea) are mainly located on three lamellar segments of the antennal club. Variation was also seen in abundance of various types of antennal sensilla, with males possessing significantly more sensilla than females. Sensilla placodea were the most abundant, and their number in males was twice of that in females, showing a clear sexual dimorphism. The difference in the distribution of sensilla placodea might reflect their roles in sexual chemical communication.     

Histological and ultrastructural features of the rectum in Poecilimon cervus Karabağ, 1950 (Orthoptera: Tettigoniidae)

The morphology and ultrastructure of the rectum in Poecilimon cervus Karaba?, 1950 (Orthoptera, Tettigoniidae) were analyzed by light microscope, scanning (SEM) and transmission electron microscopes (TEM). The rectum is the final part of the digestive tract that plays an important role in water reabsorption in insects and so provides osmoregulation. In the transverse sections, six rectal pads and columnar epithelium can be distinguished. The cuticular intima lines the lumen at the apical side of the epithelium. In the cytoplasm, there are numerous mitochondria, some endocytic vesicles, secreting vesicles whose sizes differ according to the area in the cell, and a nucleus with globular in shape. With this study, we aimed to demonstrate the ultrastructure of the rectum of P. cervus and differences or similarities of with other species.     

Morphology of laryngeal cartilage of the nine‐banded armadillo (Dasypus novemcinctus) Linnaeus, 1758

Armadillos, Xenarthras representatives, known for adaptability to different ecosystems, own specific morphophysiological characteristics that are not known and deserve to be studied. The aim of this study was to describe the morphology of cartilage of the larynx of the nine‐banded armadillo (Dasypus novemcinctus). Five dead armadillos were donated by the Chico Mendes Institute of Biodiversity (ICMBio‐PI) to the Federal University of Piauí. The animals were fixed and dissected for removal of the larynx. The cartilages were identified and described, photodocumented, and schematized. Fragments with about 0.5 cm of each cartilage were collected and submitted to classical histology for Hematoxylin–Eosin coloring. The slides were assembled in enterlan and analyzed under a light microscope. The larynx of the armadillo (D. novemcinctus) is located in the mentonian region, ventral to the esophagus, and due to the total positioning of the tongue in the oral cavity, there is also a cranial cervical position in this species. The larynx has five cartilages, they are: a cricoid, a thyroid, an epiglottis, and two arytenoids. The corniculate process is present; however, the cuneiform process is absent. The epiglottis has a discrete bifurcation at its apex. In all cartilages epithelial variations are observed. The tissues are varied from squamoso stratified to cylindrical pseudostratified, with propria lamina rich in mucoserosas glands. With the exception of epiglottic cartilage, predominantly elastic, the rest are hyaline. The larynx of D. novemcinctus, although the same number of cartilages, differs morphologically and microscopically from the larynx of other species.     

Morphological description of the immature stages of Hermetia illucens (Linnaeus, 1758) (Diptera: Stratiomyidae)

In the dipteran genus Hermetia, only 6 of the 78 valid species have documented immature stages: H. albitarsis Fabricius, 1805, H. aurata Bellardi, 1859, H. concinna Williston, 1900, H. illucens (Linnaeus, 1758), H. panamensis Greene, 1940 and H. pulchra Weidemann, 1830. In particular, H. illucens stands out due to its reported applicability for forensic, medical and economic purposes. Here, we described the morphology of eggs and immature stages of this species, with a view to detecting differences between instars and in the pupal stage, which should eventually help properly identifying larval age. We utilized both optical and scanning electron microscopy tools. The eggs are elliptical and elongated, and color varies from cream white to yellowish. The larvae are apodal, hemichephalic and holopneustic, flattened dorso‐ventrally and may be recognized by the head elongated, dorsal and ventral chaetotaxy of the cephalic capsule, thoracic and abdominal segments, and the morphology of the anterior and posterior spiracles. The pupae are adecticous and coarctate, tegument dark brown and pruinescence varying from brown to golden. The overall morphology across instars is similar, but marked variations were observed in the shape of the antennal articuli and the shape of the setae (first instar compared to the others). Our results supplement the biological information on Hermetia illucens and should aid the proper identification and aging of juveniles in the field, as a way to minimize errors in the calculation of the post‐mortem interval.