Histochemical and immunohistochemical adaptations on the gastroesophageal tube of the blue-fronted parrot (Amazona aestiva Linnaeus, 1758)

Amazona aestiva Linnaeus, 1758 is known as the blue-fronted parrot. These animals are considered seed dispersion, contributing to ecological balance. Despite its ecological importance, A. aestiva is listed as an imminent danger of becoming extinct, as it is one of the most illegally traded native species in Brazil, besides losing its habitat due to deforestation and forest fires. The cranial oesophagus mucosa is lined with a non-keratinized stratified squamous epithelium, and the basal layer presents melanocyte accumulation. No mucous glands were observed in the lamina propria/submucosa. The ingluvium two different muscular layer patterns were observed, one internal and circular and one very thick, external and longitudinal. The proventriculum submucosa consists of polygonal proventricular glands and contents into a broad central duct lined with simple columnar epithelium Periodic acid-reactive Schiff positive (PAS+) and Alcian blue positive (AB+). The 5-HT cells are dispersed in the proventricular glands located at base and apex. The ventricle mucous layer contains numerous branched folds lined by a simple columnar epithelium. The epithelium was negative reaction for PAS and positive for AB only on the fold surfaces. The 5-HT cells were scarce and observed only at the base of the ventricular gland, as well as in the adjacent connective tissue.     

Ultrastructure and regeneration of midgut epithelial cells in Lithobius forficatus (Chilopoda,Lithobiidae)

Lithobius forficatus (Myriapoda, Chilopoda, Lithobiidae) is a widespread species of centipede that is common across Europe. Its midgut epithelial cells are an important line of defense against toxic substances that originate in food, such as pathogens and metals. Despite this important role, the biology of the midgut epithelium is not well known. Here we describe the ultrastructure of the midgut epithelium, as well as the replacement of degenerated midgut epithelial cells. The midgut epithelium of L. forficatus is composed of digestive, secretory, and regenerative cells. The cytoplasm of digestive cells shows regionalization in organelle distribution, which is consistent with the role of these cells in secretion of enzymes, absorption of nutrients, and accumulation of lipids and glycogen. Secretory cells, which do not reach the luminal surface of the midgut epithelium, possess numerous electron‐dense and electron‐lucent granules and may have an endocrine function. Hemidesmosomes anchor secretory cells to the basal lamina. Regenerative cells play the role of midgut stem cells, as they are able to proliferate and differentiate. Their proliferation occurs in a continuous manner, and their progeny differentiate only into digestive cells. The regeneration of secretory cells was not observed. Mitotic divisions of regenerative cells were confirmed using immunolabeling against BrdU and phosphohistone H3. Hemocytes associate with the midgut epithelium, accumulating between the visceral muscles and beneath the basal lamina of the midgut epithelium. Hemocytes also occur among the digestive cells of the midgut epithelium in animals infected with Rickettsia‐like microorganisms. These hemocytes presumably have an immunoprotective function in the midgut.     

Effect of gluten on the digestive tract and fat body of Telodeinopus aoutii (Diplopoda)

Adult specimens or larvae of invertebrates used as food for vertebrates are often maintained close to gluten so they might become vectors for cereal proteins. However, the tissues and internal organs can respond differently in animals with different feeding habits. The midgut epithelium might be a first and sufficient barrier preventing uptake and effects of gluten on the whole body, while the fat body is the main organ that accumulates different xenobiotics. Good models for such research are animals that do not feed on gluten-rich products in their natural environment. The project's goal was to investigate alterations in the midgut epithelium and fat body of the herbivorous millipede Telodeinopus aoutii (Diplopoda) and analyze cell death processes activated by gluten. It enabled us to determine whether changes were intensified or reversed by adaptive mechanisms. Adult specimens were divided into control and experimental animals fed with mushrooms supplemented with gluten and analyzed using transmission electron microscopy, flow cytometry, and confocal microscopy. Two organs were isolated for the qualitative and quantitative analysis: the midgut and the fat body. Our study of the herbivorous T. aoutii which does not naturally feed on gluten containing diet showed that continuous and prolonged gluten feeding activates repair processes that inhibit the processes of cell death (apoptosis and necrosis) and induce an increase in cell viability.     

Effects of citronella oil (Cymbopogon winterianus Jowitt ex Bor) on Spodoptera frugiperda (J. E. Smith) midgut and fat body

The armyworm, Spodoptera frugiperda, is the principal pest of corn in Brazil. Control is achieved primarily by synthetic insecticides, which cause problems for the agro-ecosystem. Alternative methods of control are under investigation and citronella (Cymbopogon winterianus) essential oil appears to be a promising agent. We investigated the effects of citronella oil using histological, histochemical and immunohistochemical methods. The midgut of larvae treated with citronella exhibited altered epithelium including cytoplasmic protrusions, columnar cell extrusion, pyknotic nuclei, and increased periodic acid-Schiff positive granules. Regenerative cells in the epithelium of the midgut increased in number, which facilitated subsequent regeneration of this tissue. After exposure to citronella, trophocytes, the principal cell type of the fat body, possessed enlarged vacuoles and mitotic bodies, and contained reduced amounts of glycogen, lipid, and protein. Citronella oil caused morphological changes of the midgut and reduction of stored resources in the fat body, which may adversely affect insect reproduction and survival.     

Ontogenetic divergence of the phytoparasitic nematodesMacroposthonia nainitalensis andM. tenuiannulata (Criconematidae, tylenchia)

Promorphogenesis, egg cleavage, and gastrulation proceed inMacroposthonia nainitalensis similarly to other phytonematodes of the subclass Tylenchia. Before the onset of gastrulation, the greatest similarity was observed in a descendant of the micromere 2b, which separates macromeres of the third division. Deviations from the general line of development started in this species during embryonal organogenesis with the formation of an external epithelium with a special rough surface layer, which was absent in the embryos of other phytonematodes. In addition, during this period of acceleration, the digestive system developed, while peristaltic changes in body shape appeared at the tadpole stage. Later, somatic muscle development was enhanced in the first instar larvae ofM. nainitalensis, while in the second instar larvae ofMacroposthonia spp., somatic muscle cells were located at an acute angle to the longitudinal body axis. In addition, denticles—microchaetae—appeared in the second to fifth instar larvae of these nematodes. All of these features, which have not been observed in other phytonematodes of the subclass Tylenchia, help the peristaltic crawling ofMacroposthonia spp. In contrast to other Tylenchia, the musculocutaneous sac did not develop inMacroposthonia spp. However, there are similarities in the development of the digestive and genital systems.     

Structure of the Digestive Tube in the Holothurian Eupentacta fraudatrix (Holothuroidea: Dendrochirota)

In the holothurian Eupentacta fraudatrix,the gut wall exhibits trilaminar organization. It consists of an inner digestive epithelium, a middle layer of connective tissue, and an outer mesothelium (coelomic epithelium). The pharynx, esophagus, and stomach are lined with a cuticular epithelium composed of T-shaped cells. The lining epithelium of the intestine and cloaca lacks a cuticle and consists of columnar vesicular enterocytes. Mucocytes are also encountered in the digestive epithelium. The connective tissue layer is composed of a ground substance, which houses collagen fibers, amoebocytes, morula cells, and fibroblasts. The gut mesothelium is a pseudostratified epithelium, which is dominated by peritoneal and myoepithelial cells and also includes the perikarya and processes of the neurons of the hyponeural plexus and vacuolated cells.     

An ultrastructural study of alimentary tract development in the cercariae of Prosorhynchoides borealis (Digenea,Bucephalidae)

Podvyaznaya I. 2011. An ultrastructural study of alimentary tract development in the cercariae of Prosorhynchoides borealis (Digenea, Bucephalidae). —Acta Zoologica (Stockholm) 92 : 170–178. The development of digestive system in Prosorhynchoides borealis cercariae was studied using transmission electron microscopy. The foregut and caecum primordia arise in early cercarial embryos as two adjoining cellular cords. The primordial pharynx appears as a cluster of myoblasts in the mid‐part of the foregut primordium whose proximal end abuts onto the ventral embryonic tegument. Later, a lumen develops within the gut primordia and their component cells form the embryonic cellular epithelium with an essentially similar structure in the foregut and caecal regions. Subsequently, the foregut epithelial cells merge to form a syncytium. This process proceeds asynchronously and the most proximal foregut area remains cellular for the longest time. The syncytial lining of the foregut establishes syncytial connections with secretory cytons differentiating in the surrounding parenchyma. These cytons produce secretory granules, which are transported through cytoplasmic connections to the foregut syncytium. Before cercariae reach maturity, their foregut epithelium becomes anucleate and continuous with the external tegument. By the end of cercarial development, numerous short lamellae appear on the luminal surface of the caecal epithelium. The caecal cells become involved in secretory activity as indicated by the presence of Golgi‐derived secretory bodies in their cytoplasm.     

Light and electron microscopic study of the anterior oesophagus of Bulla striata (Mollusca,Opisthobranchia)

Lobo‐da‐Cunha, A., Oliveira, E., Alves, Â., Coelho, R. and Calado, G. 2010. Light and electron microscopic study of the anterior oesophagus of Bulla striata (Mollusca, Opisthobranchia). —Acta Zoologica (Stockholm) 91 : 125–138. The anterior oesophagus of Bulla striata was investigated with light and electron microscopy. In the most anterior region, the ridges of the oesophageal wall are covered by a ciliated columnar epithelium forming large apical blebs which are released into the lumen, an activity that is particularly intense in the oesophageal pouch. In the last two‐thirds of the anterior oesophagus, the epithelium is covered with microvilli embedded in a cuticle, but apocrine secretion and cilia are absent. Subepithelial secretory cells are very abundant in the oesophageal wall, except in the roof of the pouch. They have a long neck that crosses the epithelium, whereas the cell body containing the nucleus is embedded in the connective tissue. Large electron‐lucent secretory vesicles and many Golgi stacks fill most of their cytoplasm. The histochemical and cytochemical assays show that these cells secrete acid mucopolysaccharides. With the current and future studies we aim to obtain data for the establishment of relationships between morphofunctional features of the digestive system and food types in cephalaspideans. Additionally, the new data about the oesophageal pouch of B. striata may be useful for the establishment of eventual homologies with the oesophageal diverticula of other opisthobranchs.     

Apoptosis in the digestive tract of herbivorous Rana pipiens larvae and carnivorous Ceratophrys ornata larvae: An immunohistochemical study

The lifespan of herbivorous Rana pipiens larvae is ～3 months, while that of carnivorous Ceratophrys ornata larvae is only about 2 weeks. During metamorphic climax, the larval gut shortens dramatically, especially in R. pipiens, and its luminal epithelium is replaced by adult‐type epithelium. To determine when programmed cell death occurs during the metamorphic restructuring of the gut, we prepared cross‐sections of the stomach, small intestine, and large intestine from representative larval stages and from juvenile frogs of both species. The sections were incubated with monoclonal antibody against active caspase‐3, one of the key enzymes in the apoptotic cascade. We observed apoptosis in some luminal epithelial cells in each of the three regions of the larval gastrointestinal tract of both species. However, apoptotic cells appeared earlier in larval stages of R. pipiens than C. ornata and few were seen in juvenile frogs of either species. The results demonstrate the occurrence of apoptosis in the metamorphic remodeling of the gut of both R. pipiens larvae and C. ornata larvae. J. Morphol., 2011. © 2011 Wiley Periodicals, Inc.     

Morphological characteristics of the Pterodoras granulosus digestive tube (Valenciennes, 1821) (Osteichthyes,Doradidae)

Little is known about the digestive tube (DT) morphology of the fish Pterodoras granulosus. Therefore, macro‐, meso‐ and microscopic aspects of 15 P. granulosus DTs were analysed. The muscular layer was composed of striated skeletal muscle in the oesophagus and smooth muscle in the other segments. The epithelium progressed from a stratified pavement in the oesophagus to a simple column in the other segments, with a flat striated border in the intestine. A large number of mucus‐secreting periodic acid‐Schiff (PAS)‐positive cells were observed in the oesophagus. In the stomach, the number of glands in the region decreased towards the cardiac–fundic region, and none were found in the pylorus. The intestine showed an epithelium with absorption cells and an increasing number of PAS‐positive caliciform cells towards the distal region. Tests showed that the oesophagus is adapted for passing and preparing food for the chemical digestion that occurs in the stomach, which also has storage functions without grinding action. The proximal intestinal region was consistent with fat absorption, and the medium region, with the absorption of other nutrients. The distal region was short and consistent with a role in absorption for osmoregulation as well as in the formation, storage and disposal of faeces.     

生物发酵稻秸对湖羊消化道上皮形态及微生物区系的影响

【目的】本研究旨在通过显微观察和16SrRNA基因高通量测序技术来探究饲喂生物发酵稻秸对湖羊肠道上皮形态及微生物区系的影响。【方法】试验选择70日龄、体重相近(25.15±0.47)kg的湖羊公羔21只,根据饲粮中粗饲料的组成随机分为3组：稻秸组、生物发酵稻秸组和苜蓿干草组,饲粮精粗比为6:4,试验持续7周,其中适应期3周,正试期4周,结束后屠宰取样,采集瘤胃、空肠和结肠上皮组织进行观察测量,收集对应肠段内容物用于微生物区系和代谢产物测定。【结果】与稻秸组相比,饲喂生物发酵稻秸显著提高湖羊瘤胃中纤维杆菌(Fibrobacteres)的相对丰度和挥发性脂肪酸(volatile fatty acids, VFA)的含量,促进瘤胃上皮的发育;显著提高湖羊空肠中厚壁菌门(Firmicutes)和疣微菌门(Verrucomicrobia)的相对丰度,改变空肠微生物菌群结构,促进空肠上皮组织的发育;改变湖羊结肠微生物菌群结构。【结论】饲喂生物发酵稻秸有利于湖羊肠道上皮的发育,并增加消化道内的微生物多样性。     

香鱼消化道及肝脏的形态结构特征

采用解剖及石蜡切片显微技术观察了香鱼消化道及肝脏的组织学结构。香鱼消化道由口咽腔、食道、胃及肠构成。口咽腔大且狭长,其底壁前部有一对粘膜褶,两颌边缘着生宽扁梳状齿,腭骨及舌骨具齿,犁骨无齿;舌由基舌骨突出部分覆盖粘膜构成,舌粘膜上皮为复层扁平上皮,含有较多的杯状细胞和味蕾。食道、胃及肠均由粘膜层、粘膜下层、肌层及外膜构成。食道粘膜层上皮为复层扁平上皮,杯状细胞发达。胃呈V形,由贲门部、胃体部及幽门部组成,胃壁粘膜上皮为单层柱状上皮,贲门部与胃体部的固有层中有胃腺。肠较短,由前、中、后肠构成,肠壁粘膜上皮为单层柱状上皮,其游离面具微绒毛;上皮细胞间有杯状细胞。幽门盲囊有350～400条,其组织学结构与肠相同。肝脏单叶,外被浆膜;肝细胞形态不规则,肝小叶界限不明显。     

Regeneration of the Digestive Tube in the Holothurian Apostichopus japonicusafter Evisceration

We performed a TEM study of regeneration of the intestine in the Far Eastern trepang, the holothurian Apostichopus japonicus, after evisceration. The following stages were distinguished in the restoration process of the digestive tube: the growth of connective tissue along the margin of the mesenterium, in the place of rupture; dedifferentiation of cells and their migration and proliferation; the rooting of the esophagus lining into the connective tissue anlage; and the transformation of esophagus cells into cells of the middle part of the intestine. The migration of epithelium into the area of regeneration takes place through a solid cellular layer, without breaking of the cell contacts. The mitotic activity was registered in all stages of restoration; the dividing cells were located chaotically, without the development of a blastema.     

Imidacloprid impairs the post‐embryonic development of the midgut in the yellow fever mosquito Stegomyia aegypti (=Aedes aegypti)

The mosquito Stegomyia aegypti (=Aedes aegypti) (Diptera: Culicidae) is a vector for the dengue and yellow fever viruses. As blood digestion occurs in the midgut, this organ constitutes the route of entry of many pathogens. The effects of the insecticide imidacloprid on the survival of St. aegypti were investigated and the sub‐lethal effects of the insecticide on midgut development were determined. Third instar larvae were exposed to different concentrations of imidacloprid (0.15, 1.5, 3.0, 6.0 and 15.0 p.p.m.) and survival was monitored every 24 h for 10 days. Midguts from imidacloprid‐treated insects at different stages of development were dissected and processed for analyses by transmission electron microscopy, immunofluorescence microscopy and terminal deoxynucleotidyl transferase dUTP nick‐end labelling (TUNEL) assays. Imidacloprid concentrations of 3.0 and 15.0 p.p.m. were found to affect midgut development similarly. Digestive cells of the fourth instar larvae (L4) midgut exposed to imidacloprid had more multilamellar bodies, abundantly found in the cell apex, and more electron‐lucent vacuoles in the basal region compared with those from untreated insects. Moreover, imidacloprid interfered with the differentiation of regenerative cells, dramatically reducing the number of digestive and endocrine cells and leading to malformation of the midgut epithelium in adults. The data demonstrate that imidacloprid can reduce the survival of mosquitoes and thus indicate its potentially high efficacy in the control of St. aegypti populations.     

Ultrastructural, histochemical and cytochemical characterization of intestinal epithelial cells in Aplysia depilans (Gastropoda, Opisthobranchia)

To improve the current knowledge about the digestive system in opisthobranchs, light and electron microscopy methods were used to characterize the epithelial cells in the mid‐intestine of Aplysia depilans. This epithelium is mainly formed by columnar cells intermingled with two types of secretory cells, named mucous cells and granular cells. Columnar cells bear microvilli on their apical surface and most of them are ciliated. Mitochondria, multivesicular bodies, lysosomes and lipid droplets are the main components of the cytoplasm in the region above the nucleus of these cells. Peroxisomes are mainly found in middle and basal regions, usually close to mitochondria. Mucous cells are filled with large secretory vesicles containing thin electron‐dense filaments surrounded by electron‐lucent material in which acidic mucopolysaccharides were detected. The basal region includes the nucleus, several Golgi stacks and many dilated rough endoplasmic reticulum cisternae containing tubular structures. The granular cells are characterized by very high amounts of flat rough endoplasmic reticulum cisternae and electron‐dense spherical secretory granules containing glycoproteins. Enteroendocrine cells containing small electron‐dense granules are occasionally present in the basal region of the epithelium. Intraepithelial nerve fibres are abundant and seem to establish contacts with secretory and enteroendocrine cells.     

Quantitative comparative analysis of the nasal chemosensory organs of anurans during larval development and metamorphosis highlights the relative importance of chemosensory subsystems in the group

The anuran peripheral olfactory system is composed of a number of subsystems, represented by distinct neuroepithelia. These include the main olfactory epithelium and vomeronasal organ (found in most tetrapods) and three specialized epithelia of anurans: the buccal‐exposed olfactory epithelium of larvae, and the olfactory recess and middle chamber epithelium of postmetamorphic animals. To better characterize the developmental changes in these subsystems across the life cycle, morphometric changes of the nasal chemosensory organs during larval development and metamorphosis were analyzed in three different anuran species (Rhinella arenarum , Hypsiboas pulchellus , and Xenopus laevis ). We calculated the volume of the nasal chemosensory organs by measuring the neuroepithelial area from serial histological sections at four different stages. In larvae, the vomeronasal organ was relatively reduced in R. arenarum compared with the other two species; the buccal‐exposed olfactory epithelium was absent in X. laevis , and best developed in H. pulchellus . In postmetamorphic animals, the olfactory epithelium (air‐sensitive organ) was relatively bigger in terrestrial species (R. arenarum and H. pulchellus ), whereas the vomeronasal and the middle chamber epithelia (water‐sensitive organs) was best developed in X. laevis . A small olfactory recess (likely homologous with the middle chamber epithelium) was found in R. arenarum juveniles, but not in H. pulchellus . These results support the association of the vomeronasal and middle chamber epithelia with aquatic olfaction, as seen by their enhanced development in the secondarily aquatic juveniles of X. laevis . They also support a role for the larval buccal‐exposed olfactory epithelium in assessment of oral contents: it was absent in X. laevis , an obligate suspension feeder, while present in the two grazing species. These initial quantitative results give, for the first time, insight into the functional importance of the peripheral olfactory subsystems across the anuran life cycle.     

Effects of vitamin A-deficiency and inflammation on the conducting airway epithelium of Syrian golden hamsters

The effects of vitamin A-deficiency and inflammation were studied in the conducting airways of Syrian golden hamsters. An important goal of the study was to characterize epithelial changes that occur early in vitamin A-deficiency, that might precede yet predispose to infection, and precipitate inflammatory changes in the lungs. Age-matched vitamin A-replete control and vitamin A-deprived hamsters were killed at 33 days of age (preweight-plateau); at 41 days of age (weight plateau-early weight loss); and at 48–55 days of age (prolonged weight plateau followed by weight loss). A tablet containing bromodeoxyuridine (BrdU) was implanted subcutaneously into each hamster 7 h before it was killed. No changes were seen in the conducting airway epithelium of vitamin A-deprived hamsters in the preweight plateau. However, labelling of secretory cells for BrdU was reduced 6–7 fold in the epithelium lining the lobar bronchus (p< 0.0002) and the bronchioles (p< 0.0001), and the proportions of ciliated cells were decreased (p<0.0001) at both airway levels in vitamin A-deficient hamsters in the weight plateau-early weight loss stage. Changes in cellular morphology were minimal in the intrapulmonary airway epithelium at this time but a few small focal patches of epidermoid metaplasia were seen in the tracheal epithelium. Small foci of inflammation were closely associated with the airways in the weight plateau, and the inflammation became more widespread when the deficiency was prolonged. The results suggest that the defense of the lungs to infection was impaired initially in the vitamin A-deficient hamsters by a widespread reduction in the numbers of ciliated cells throughout the epithelium of the conducting airways (trachea, bronchi, bronchioles). At the foci of inflammation, labelling of epithelial secretory cells for BrdU was greatly increased at all airway levels. A highly stratified cornifying epidermoid metaplasia developed in the tracheal epithelium, and goblet cell metaplasia developed in the cranial portion of the lobar bronchus, in association with submucosal inflammation. Goblet cell metaplasia appeared to be the only abnormality that wasnot reversed when vitamin A was restored to the diet. This is contribution no. 2911 from the Pathobiology Laboratory     

Validation of a planimetric procedure to quantify stress in Littorina littorea (Gastropoda: Mollusca): is it independent of the reproductive cycle?

This study forms part of a larger project in which five planimetric parameters have been used to study changes in the digestive epithelium of Littorina littorea under different environmental and physiological conditions. Our aim was to examine the effect of the reproductive cycle on these parameters in order to assess their usefulness as indicators of stress. A one way anova shows that the absolute parameters of mean epithelial thickness (MET), mean diverticulum radius (MDR) and mean luminal radius (MLR) vary significantly depending on the time of the year. This variation is highly correlated to the amount of digestive tissue present and hence negatively correlated to the reproductive state because gonad and digestive tissue volume are inversely related. Consequently, these parameters are not good indicators of stress. However, whereas the absolute size of the digestive acinus varies with the reproductive state of the animal, the MET, MDR and MLR retain the same proportions and therefore the ratios MET/MDR and MLR/MET remain constant. This makes them useful indicators of stress because they are independent of intrinsic variables such as the reproductive cycle.