PROTEOME ANALYSIS OF GUT AND SALIVARY GLAND PROTEINS OF FIFTH-INSTAR NYMPH AND ADULTS OF THE SUNN PEST, Eurygaster integriceps

In the digestive system of the sunn pest, Eurygaster integriceps Puton (Hemiptera: Scutelleridae), the salivary gland has a key role in extra oral digestion and the gut is the main site for digestion of food. In this study, proteomics was used to study the role of proteins involved in digestion. The amount of feeding on wheat grain by adult insects increased by comparison to fifth-instar nymphs. Proteins of the gut and salivary gland in adults and fifth-instar nymphs were analyzed 1 day after feeding. The proteins related to digestion, metabolism, and defense against toxins were accumulated in the gut of adult insects. Three plant proteins including serpin, dehydroascorbate reductase, and β-amylase were accumulated in guts of adults. In the salivary gland, phospholipase A2 and arginine kinase were increased in adults. Heat shock protein 70 increased in the gut of fifth-instar nymphs. Proteomic analysis revealed that most of changed proteins in digestive system of sunn pest were increased in adults. This study provided more targets derived from gut and salivary gland for pest management.     

Feeding and gut physiology in Acanthopleura spinigera (Mollusca)

The morphology and histology of the alimentary canal of the rock chiton Acanthopleura spinigera are described and the ability of regions of the gut to digest specific substrates investigated. The oesophagus is produced into a pair of thin-walled lateral pouches, the salivary glands or "sugar glands" which empty into the stomach. Folds of the capacious stomach are almost obscured by the large digestive gland over which is coiled the intestine. Histologically the gut consists of an outer layer of connective tissue, an inner muscular layer and a ciliated epithelium which varies in thickness from one region to the next. Proteases are most active in the stomach, digestive gland and anterior intestine at pH 6·5 and in the posterior intestine at pH 7·5-8·5. The digestion of lipoidal substance was greatest in the stomach and digestive gland and least in anterior intestine. There was little increase in the amount of digestion product obtained after 20 hours incubation. All regions of the alimentary canal and salivary gland were capable of digesting carbohydrates except that many low molecular weight carbohydrates were digested by salivary gland extracts only. The amylases were most active at pH 6–6·5. It is concluded that digestive enzymes are distributed throughout the intestinal tract but the amount of enzyme present varies from region to region, and is greatest just after feeding.     

The digestive gland of Viviparus ater (Mollusca, Gastropoda, Prosobranchia): an ultrastructural and histochemical study

The digestive gland of Viviparus ater was studied using histochemical and ultrastructural methods. Only one cell type was observed in the tubule epithelium of the gland. The cells are involved in an endocytotic process mediated by clathrin-coated vesicles and in the intracellular digestion of food materials (thus they can be regarded as digestive cells). The different stages of digestion and exocytotic extrusion of residual bodies into the tubule lumen were shown by electron microscopy. Very few, small mucocytes are scattered among the digestive cells. Calcium concretions, glycogen-containing cells and endocrine cells are scattered in the area of connective tissue present among the digestive tubules.     

THE FUNCTIONAL ANATOMY OF THE GUT OF THE PROSOBRANCH GASTROPOD POMACEA CANALICULATA AND OF SOME OTHER PILIDS

The structure and functioning of the gut of Pomacea canaliculata (D'Orb.) has been investigated using living and preserved material. Anatomical studies were also carried out on preserved specimens of Pila globosa, Turbinicola saxea and Lanistes ovum bangweolicus .
The gut of pilids is specialised for a macrophagous diet, usually of aquatic angiosperms. The mid-oesophagus is a crop for storage, and the stomach has a large triturating gizzard developed from the gastric shield area. This is the site of extra-cellular digestion; there is no intra-cellular digestion in any part of the gut. The ducts of the digestive gland open into a special region of the stomach, the vestibule, which is histologically similar to them. The style sac begins the compacting of the faeces, which is completed in tho intestine. There is no evidence that absorption ocrurs in the epithelium of tho stomach or intestine; soluble products of digestion are carried into the digestive gland, which is the main site of absorption. Its activity is supplemented by amoebocytes entering the lumen of the style sac and intestine. Two types of cell occur in the gland, one type producing digestive enzymes and absorbing soluble products of digestion, the other type being excretory in function. There is no sign of phagocytosis in either. The excretory activity of the kidney is further supplemented by an anal gland.     

Changes in the digestive gland of the loliginid squid Sepioteuthis lessoniana (Lesson 1830) associated with feeding

Changes associated with feeding in the histological and cytological structure of the digestive gland of the loliginid squid Sepioteuthis lessoniana were examined, along with the nature of both the intracellular and extracellular enzymes produced by the gland. The timing of the release of the extracellular enzymes during the digestive cycle was also determined using a quantitative experimental program. Like that of all coleoid cephalopods, the digestive gland was characterised by one type of cell with several functional stages. As is the case for other loliginid squids, however, the digestive cells did not contain the large enzyme-carrying boules that characterise the digestive glands of most cephalopods. Instead, smaller secretory granules were found in the digestive cells and these may be the enzyme carriers. The prominent rough endoplasmic reticulum, large mitochondria and active Golgi complexes present in the digestive cells are characteristic of cephalopods and indicate a high metabolic activity. Like that of other cephalopods, endocytotic absorption of nutrients and intracellular digestion occurs in the digestive gland of this squid. From quantitative and qualitative examinations of structural changes in the digestive gland of S. lessoniana after feeding, a schedule of its function during the course of digestion was proposed. This indicated that digestion was very rapid, being completed in as little as 4 h in S. lessoniana. Extracellular digestive enzymes were only released after the first hour following feeding, which implies that they are stored in the stomach between meals to increase digestive efficiency.     

The lysozyme locus in Drosophila melanogaster: an expanded gene family adapted for expression in the digestive tract

Lysozyme has been studied in insects as part of the system of inducible antibacterial defence in the haemolymph. We recently found two Drosophila lysozyme genes that are constitutively expressed in the digestive tract, and are probably involved in the digestion of bacteria in the food. To obtain an overview of the lysozyme genes in this species and their possible roles in immunity and digestion, we have now characterized all six lysozyme genes in the cloned part of the lysozyme locus at 61F, and a seventh gene that maps to the same chromosomal location. The expression of the genes follows four different patterns: firstly, four closely related genes, LysB, C, D and E, are all strongly expressed in the midgut of larvae and adults; secondly, LysP is expressed in the adult salivary gland; thirdly, LysS is expressed mainly in the gastric caecae of larvae; and finally, LysX is primarily expressed in the metamorphosing midgut of late larvae and early pupae. The LysD-like genes and LysS are strongly repressed in artificially infected animals, possibly reflecting a malaise reaction in the digestive tract. None of the genes is expressed in the fat body or haemocytes. Thus rather than being a component of the haemolymph, the Drosophila lysozymes are found mainly in the digestive tract where they are expressed at a high level. Furthermore all genes, except LysP, encode acidic proteins, in contrast to the strongly basic typical lysozymes. This is highly reminiscent of the situation in ruminants, where the lysozymes have been recruited for the digestion of symbiotic bacteria in the stomach.     

The Ultrastructure of the Gastrodermal Gland Cells in the Freshwater Planarian Dugesia gonocephala s.l.

Light and transmission electron microscopy have been used to study the gastrodermal gland cells of the triclad Dugesia gonocephala s.l. The events involved in the ultrastructural transformation and the secretion process in these cells were followed at four different stages in both fasted and fed animals. During the feeding stage their secretory granules are directly discharged into the intestinal lumen by means of a secretion process of the holocrine type that is described in this paper. It is suggested that such secretions contribute to extracellular digestion and that disintegration of the gland cells is accompanied by a differentiation of neoblasts into new gland cells, reflecting a turnover of gland cells during the triclad digestive stages.     

Location of immunoreactive leucine enkephalin in the digestive gland of the Mollusca Mizuhopecten yessoensis (Jay)

Distribution of leu-enkephalin-like peptides in the digestive gland of the Japanese scallop has been investigated by means of the indirect immunohistochemical method. The immunoreactive leu-enkephalin is revealed in supranuclear vacuoles of separate large secretory cells. These cells are situated in epithelium of the digestive tubules along the gland periphery. Their number is from 1-3 up to 5-6 per tubue section. The basal part of the cell and small apical vacuoles do not contain any products of the reaction. It is possible that in the bivalve molluscs, as in vertebrates, the leu-enkephalin-immunoreactive elements participate in regulation of digestion.     

Ultrastructural localization of esterase and acid phosphatase in digestive gland cells of fed and starvedCepaea nemoralis (L.) (Mollusca: Helicidae)

Summary The ultrastructural localizations of thiolacetic acid esterase, indoxyl acetate esterase and acid -glycerophosphatase have been studied in the digestive gland cells of fed and starvedCepaea nemoralis. In fed snails the major localization of all three enzymes was in the green granule vacuoles of digestive cells. In addition, the cytoplasm of calcium cells and the Golgi apparatus and GERL (?) of all cell types were acid phosphatase positive. Many digestive cells of starved snails showed a similar enzyme distribution to that found in fed snails but other digestive cells showed a very high cytoplasmic activity of all three enzymes. It is suggested that these cells are in the process of autolysis. New light is also thrown on the process by which food is transported from the digestive gland lumen to the phagosomes of digestive cells.     

Fine structure of the midgut gland of Phalangium opilio (Chelicerata,Phalangida)

Summary The fine structure of the midgut gland and the changes in composition associated with the digestive activity were examined in Phalangium opilio. In the epithelium four different types of cells are present: ferment cells, resorption cells, and digestion cells which probably turn into excretion cells, as can be seen by many intermediate stages. Ferment cells are found only in the midgut gland and in no other epithelia; therefore they should be regarded as a cell type. The relationship between digestion and resorption cells is not yet clear. No regeneration zone or single regeneration cells could be identified.The ultrastructural changes in these different cells during digestion are described, and their functional aspects are discussed. A hypothetical digestive cycle is constructed from these data. The results are compared with those on other chelicerate midgut glands.     

Phosphatase activity in the hepatopancreas of Helix aspersa

• 1.1. Phosphatase acid (PhA) activity in the digestive gland (hepatopancreas) of the common garden snail Helix aspersa has been investigated using cytochemical methods.
• 2.2. All the cells composing this gland show PhA activity, the distribution pattern differing according to the cell type.
• 3.3. The digestive cells show the most widely distributed reaction product (brush border, phagolysosomes, multivesicular bodies and autophagic vacuoles).
• 4.4. In the excretory cells this activity appears in large sacs, while in the calcium cells the reaction product is abundant in the calcium granules.
• 5.5. Cellular digestion processes performed by each of these cell types is discussed together with their role in the detoxification of heavy elements derived from the environment.

     

The lysozyme locus in Drosophila melanogaster: an expanded gene family adapted for expression in the digestive tract

Lysozyme has been studied in insects as part of the system of inducible antibacterial defence in the haemolymph. We recently found two Drosophila lysozyme genes that are constitutively expressed in the digestive tract, and are probably involved in the digestion of bacteria in the food. To obtain an overview of the lysozyme genes in this species and their possible roles in immunity and digestion, we have now characterized all six lysozyme genes in the cloned part of the lysozyme locus at 61F, and a seventh gene that maps to the same chromosomal location. The expression of the genes follows four different patterns: firstly, four closely related genes, LysB, C, D and E, are all strongly expressed in the midgut of larvae and adults; secondly, LysP is expressed in the adult salivary gland; thirdly, LysS is expressed mainly in the gastric caecae of larvae; and finally, LysX is primarily expressed in the metamorphosing midgut of late larvae and early pupae. The LysD-like genes and LysS are strongly repressed in artificially infected animals, possibly reflecting a malaise reaction in the digestive tract. None of the genes is expressed in the fat body or haemocytes. Thus rather than being a component of the haemolymph, the Drosophila lysozymes are found mainly in the digestive tract where they are expressed at a high level. Furthermore all genes, except LysP, encode acidic proteins, in contrast to the strongly basic “typical” lysozymes. This is highly reminiscent of the situation in ruminants, where the lysozymes have been recruited for the digestion of symbiotic bacteria in the stomach.     

A contribution to the functional morphology of the midgut gland in phalangiid harvestmen Gyas annulatus and Gyas titanus during their life cycle

The structure of the midgut gland and its changes in different seasons have been examined in the harvestmen Gyas annulatus and Gyas titanus (Arachnida: Opiliones: Phalangiidae). In both species, in the epithelium of the midgut gland two different types of cells are present: secretory and digestive ones. The secretory cells are characterized by plentiful rER and secretory granula. The digestive cells are characterized by an apical system of tubules. Both cells are connected by prominent specialized junctions. If a secretory cell is in contact with a digestive cell, rER cisterna are in close vicinity and parallel to these junctions. As found light- and electron microscopically and also histochemically, glycogen and lipids are stored in both cells. In both species, glycogen was seen to be used as energy compound during overwintering. At the end of their life, the digestive cells develop into excretory ones, containing metabolic wastes.     

Immunochemical localisation of proliferating cells in mussel digestive gland tissue

The distribution of proliferating cells in the digestive gland of the common marine mussel, Mytilus galloprovincialis Lmk, was investigated by means of immunochemical techniques employing PC10, a commercial monoclonal antibody to the proliferating cell nuclear antigen (PCNA). Immunoblot analysis of digestive gland whole homogenates revealed a single crossreactive band of 36–37kDa, identical to the corresponding protein of rat liver and murine melanoma cells. A band of slightly higher electrophoretic mobility (34–35kDa) was found in fish liver. In mussel digestive gland, the samples obtained from young specimens presented a more intense signal for PCNA than in those obtained from old mussels, suggesting that the digestive gland cells of young mussels exhibit a higher proliferative activity. In paraffin sections, PC10 specifically labelled nuclei of all cell types, but only a smaller number of cells lining the different digestive epithelia. PCNA expression was more intense in digestive cells than in basophilic cells. Hemocytes circulating along the interdiverticular spaces also presented immunoreactive nuclei. Electron microscopy revealed a specific and moderate PC10 labelling in nuclei. Thus, single gold particles appeared disseminated throughout the nuclei with accumulations of particles in the sites of DNA replication. Taken together, these data reveal that the capacity to proliferate resides within all cell types in the digestive diverticula and do not support the hypothesis of the existence of one stem cell in this epithelium. As opposed to the hepatopancreas of the crab, Carcinus maenas, where mitotic figures and PCNA immunoreactivity are only observed in the embryonic cells within the distal portions of the digestive diverticula, apparently there are not discrete regions of cell proliferation in the digestive gland of mussels.     

Pharyngeal and Gastrodermal Ultrastructure of Prorhynchus stagnalis Schultze, 1851 (Turbellaria,Lecithoepitheliata)

The digestive tract of a freshwater Lecithoepitheliata turbellarian, Prorhynchus stagnalis, has been studied at the ultrastructural level. The buccal tube connecting the mouth opening to the pharynx is lined by an insunk epithelium, and is provided with two kinds of secretory cells. The bulbous pharynx itself, typical of the genus, is a highly muscular organ, also lined internally by an insunk epithelium; it also bears at least two different types of gland cells. Externally the pharynx is enveloped by a thin, flat epithelium. The gastrodermis consists of phagocytes and Minotian gland cells, as typical for most Tricladida. No ciliary covering was observed along the whole digestive system; instead, phagocytic cells have long microvillar projections. Preliminary attempts to follow the digestive process using external markers and cytochemical reaction for acid phosphatase were successful in demonstrating endocytic activity in the phagocytes. These data, besides being a contribution to morphology and systematics, also speak in favour of the general digestive theory of Turbellaria proposed by Jennings and serve as a starting point for experimental studies on intracellular digestion within the Lecithoepitheliata.     

皱纹盘鲍消化腺细胞类型和分泌产物

对皱纹盘鲍消化腺进行了组织学、组织化学、超微结构及酶活性测定等研究。消化腺由消化细胞和嗜碱性细胞组成。消化细胞能内吞腺管腔内的外源性物质,细胞内充满大量与异噬功能有关的囊泡。消化细胞具有内吞和细胞内消化、分泌、贮存等功能。嗜碱性细胞含有发达的粗面内质网以及许多含铁的折光小体,具有分泌和贮存金属离子的功能。消化腺还呈现多种水解酶活性。     

Multiple sources of esterase enzymes in the crop juice ofCepaea (Mollusca: Helicidae)

Summary Previous workers have (a) compared pulmonate crop juice and digestive gland extracts and found a close similarity in the enzymic complements from these two sources, and (b) located specific enzymes within the various cell types of the digestive gland. The digestive gland seems to be the major source of extracellular enzymes but what is not clear is which of the enzymes associated with particular intracellular structures are actively secreted into the crop juice. The present study has used polyacrylamide disc gel electrophoresis to investigate the digestive gland and crop juice esterases ofCepaea nemoralis andC. hortensis. It appears that only some of the digestive gland esterases are specifically secreted. The variation shown in crop juice esterases suggests three independent sources in the digestive gland. Less detailed studies ofHelix aspersa andArianta arbustorum also indicate multiple sources of extracellular esterases.     

Microstructural and histochemical advances on the digestive gland of the common cuttlefish,Sepia officinalis L.

The cephalopod digestive gland is a complex organ that, although analogous to the vertebrate liver, has additional functions, with special (albeit not exclusive) note on its active role in digestion. Although the structure of the digestive cell and its main constituents are well known (among which “boules” and brown bodies are distinctive features), histological details of other cell types and the general structure of the digestive gland need still further research. By a thorough combination of histological and histochemical techniques, it is shown that the digestive gland diverticula of the common cuttlefish (Sepia officinalis L.) are comprised of three essential cell types: digestive, basal and excretory. Basal (“pyramidal”) cells are multi-functional, being responsible for cell replacement and detoxification, mainly through the production of calcic spherulae containing metals like copper and lead in a complex organic matrix of proteins and ribonucleins. Since copper- and lead-positive spherulae were almost absent from other cell types and lumen of the tubules, it appears that controlled bioaccumulation of these metals, rather than excretion, is the main detoxification mechanism. The results show that the organ is crossed by an intricate network of blood vessels, especially arteries and arterioles, whose contents share histochemical properties with a particular set of “boules” that are shed into the lumen of diverticula for elimination, suggesting that the organ actively removes unwanted metabolites from the haemolymph. Conversely, the rarer excretory cells appear to be specialized in the elimination of salts. Although the exact nature of many excretory and secretory products, as the metabolic pathways that originate them, remain elusive, the findings suggest an intricate interaction between the different cell types and between these and the surrounding media: haemolymph and digestive tract.     

Evolution of digestion of carbohydrates in the separate parts of the digestive tract of the edible snail Helix lucorum (Gastropoda: Pulmonata: Stylommatophora) during a complete 24-hour cycle and the first days of starvation

In the present study we examined carbohydrase activities during a complete 24-h cycle and during the first days of starvation in both adult and juvenile snails. The results indicated the predominant role of the digestive gland in the secretions of the enzymes responsible for degradation of most of the carbohydrates tested. Salivary glands secreted some digestive enzymes but in amounts lower than secreted by the digestive gland. Enzymatic activities fluctuated during the first hours of digestion and also after the digestive tract was empty. The relatively high enzymatic activities recorded 24 h after the intake of food and during starvation could be due to the circadian rhythm of this species and/or to the participation of an existing microflora in the digestive tract of Helix lucorum. The double origin (exogenous and endogenous) of some digestive enzymes such as cellulases is discussed.Abbreviations CMC Carboxymethyl cellulose - LSD-test least significant difference test - PNP p-nitrophenyl - SA specific activity - U units