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Trypanosoma brucei spp. development in the tsetse fly: characterization of the post-mesocyclic stages in the foregut and proboscis.

Post-mesocyclic development of Trypanosoma brucei in the tsetse fly in its migration from midgut to salivary glands, was revisited by sequential microdissection, morphometry and DNA-cytofluorometry. This development started by day 6 after the infective feed, with passage of mesocyclic midgut trypomastigotes through proventriculus and upward migration along foregut and proboscis to the salivary gland ducts. Kinetics of salivary gland infection showed that colonization of the salivary glands by epimastigotes occurred only during the time-limited presence of this developmental phase in the foregut and proboscis. Post-mesocyclic trypanosomes in the foregut and proboscis were pleomorphic, with 4 morphological stages in various constant proportions and present all through from proventriculus up to the salivary gland ducts: 67% long trypomastigotes, 27% long epimastigotes, 4% long epimastigotes undergoing asymmetric cell division and 2% short epimastigotes. Measurements of DNA content demonstrated a predominant tetraploidy for 67% of these trypanosomes, the remainder consisting of the homogeneous diploid short epimastigotes and some long epimastigotes. According to the experimental data, the following sequence of trypanosome differentiation in the foregut and proboscis is proposed as the most obvious hypothesis. Incoming mesocyclic trypomastigotes (2N) from the ectoperitrophic anterior midgut start to replicate DNA to a 4N level, are arrested at this point, and differentiate into the long epimastigote (4N) which give rise, by an asymmetric cell division, to 2 unequal, diploid daughter cells: a long, probably dead-end long epimastigote and a short epimastigote. The latter is responsible for the epimastigote colonization of the salivary glands if launched at the vicinity of the gland epithelium by the asymmetric dividing epimastigote.

Animals↗

Differential expression of fos and jun family members in the developing chicken gastrointestinal tract.

We have analysed the expression patterns of all the known fos/jun family genes, which encode the components of the transcription factor AP-1, in the chicken embryonic digestive tract that develops into the esophagus, proventriculus, gizzard, small intestine, ceca and large intestine. From soon after formation of the tubular structure, each gene transcript was localized in distinct domains of the epithelium and mesenchyme in all of these major gastrointestinal organs, independently of the anterior-posterior axis. fra-2 was expressed predominantly in epithelium, which also expressed junD, while low-level expression of junD was also detected in smooth muscle cell precursors in mesenchyme. Expression of c-jun and c-fos was detectable in both mesenchyme and epithelium through the whole tract. In the differentiated proventriculus, the developed glandular epithelium expressed c-jun and junD, but not fra-2, while luminal epithelium expressed fra-2 and junD, but not c-jun. These results suggest that distinct Fos/Jun protein heterodimers play important roles in maintaining the epithelial-mesenchymal interactions. Similar expression patterns to those of fra-2 and junD were established from earlier stages by Sonic hedgehog gene and the Indian hedgehog gene, respectively, both of which are important in forming the inductive network between epithelium and mesenchyme of the digestive tract.

Animals↗

Characterization of genes expressed in the salivary glands of the tsetse fly, Glossina morsitans morsitans.

Salivary gland products of haematophogous insects including tsetse flies (Diptera: Glossinidia) are involved in antihaemostasis to allow for efficient blood feeding. In addition, salivary products of tsetse are thought to indirectly support the metacyclogenesis and eventual transmission of the African trypanosome protozoan parasites to their mammalian hosts. We have previously characterized the major anticoagulant, Tsetse Thrombin Inhibitor (TTI), from salivary extracts, and described molecular aspects of its cDNA from a Glossina morsitans morsitans salivary gland cDNA library. In addition, a family of two related genes with growth factor and adenosine-deaminase motifs (TSGF-1 and TSGF-2) have also been described. Here, we report on the molecular aspects of three different cDNAs and their putative products expressed in salivary glands: cDNAs TAg5, Tsal1 and Tsal2. The full-length transcript encoded by Tsetse Antigen 5 (TAg5) cDNA is 926 bp excluding the poly(A) stretch, and has an open reading frame of 259 amino acids that can encode for a protein of 28 925 Da. The putative product of TAg5 shows extensive similarities to cDNAs characterized from Drosophila (Agr and Agr2) and sandfly Lutzomyia (LuLoAG5). The cDNAs Tsal1 and Tsal2 are predicted to encode for mature proteins of 45 612 Da (399 amino acids) and 43 930 Da (389 amino acids), respectively, and their putative products exhibit over 42% identity to one another. The N terminus of each putative protein contains a hydrophobic region with signal peptide characteristics indicating that they may be secretory in nature. Transcripts specific for TAg5 and Tsal2 genes can be detected in all developmental stages of tsetse while Tsal1 expression is limited to adult and larval stages. A reverse transcription polymerase chain reaction based amplification approach indicates that TAg5 transcipts can be detected from proventriculus and midgut tissues of the fly in addition to salivary glands, while Tsal1 and Tsal2 expression is restricted to salivary gland and proventriculus. The salivary glands of adult males are found to express higher levels of TAg5 and Tsal2 in comparison to females while no significant sex-based difference is observed for Tsal1 expression. The expression of these cDNAs in different tsetse species (G. m. morsitans, Glossina austeni and Glossina fuscipes) shows wide variations.

Amino Acid Sequence↗

The oral bioavailability of ibuprofen enantiomers in broiler chickens.

Doses of racemic ibuprofen ranging from 5 to 20 mg/kg body weight were administered intravenously (i.v.) and orally to broiler chickens and plasma concentration-time profiles for both ibuprofen enantiomers were determined. The absorption of ibuprofen was evaluated after a bolus administration of a commercially available suspension into the crop and proventriculus, respectively. An enterohepatic circulation as described for other nonsteroidal anti-inflammatory drugs (NSAIDs) in other species could be suggested for both enantiomers after i.v. and oral administration. Significantly higher area under the curve (AUC) values for S(+)-ibuprofen compared with R(-)-ibuprofen were collected after crop and proventriculus administration. Several factors could be responsible for the significant differences in AUC values between both enantiomers.

Absorption↗

In vitro analysis of mesenchymal influences on the differentiation of stomach epithelial cells of the chicken embryo.

It is well established that epithelial-mesenchymal interactions play important roles in the differentiation of stomach epithelial cells in the chicken embryo. To analyze mesenchymal influences on the differentiation of the epithelial cells, we developed a tissue culture system for stomach (proventriculus and gizzard) epithelia of chicken embryo, and examined their differentiation in the presence or absence of mesenchyme. Stomach epithelium from 6-day chicken embryo did not express embryonic chicken pepsinogen (ECPg), a marker molecule of glandular epithelial cells of proventriculus, while it expressed marker molecules of epithelial cells of the luminal surface of stomach, when cultured alone on the Millipore filter, covered with the gel consisting of extracellular matrix components. When the epithelium was recombined with mesenchyme separated by the filter, differentiation of the epithelium was affected by the recombined mesenchyme. Proventricular and lung mesenchymes induced the expression of ECPg in epithelial cells, and the expression was extensive when the gel contained basement membrane components. Proventricular and gizzard epithelia showed different responses to the mesenchymal action. We tested the effects of some growth factors on the differentiation of epithelial cells using this culture system. Furthermore we devised a "conditioned semi-solid medium experiment" for analysis of the inductive properties of proventricular and lung mesenchymes. The results of this experiment clearly demonstrated for the first time that diffusible factors from mesenchyme induce the differentiation of glandular epithelial cells in the absence of mesenchymal cells.

Animals↗

The fat-like gene of Drosophila is the true orthologue of vertebrate fat cadherins and is involved in the formation of tubular organs.

Fat cadherins constitute a subclass of the large cadherin family characterized by the presence of 34 cadherin motifs. To date, three mammalian Fat cadherins have been described; however, only limited information is known about the function of these molecules. In this paper, we describe the second fat cadherin in Drosophila, fat-like (ftl). We show that ftl is the true orthologue of vertebrate fat-like genes, whereas the previously characterized tumor suppressor cadherin, fat, is more distantly related as compared with ftl. Ftl is a large molecule of 4705 amino acids. It is expressed apically in luminal tissues such as trachea, salivary glands, proventriculus, and hindgut. Silencing of ftl results in the collapse of tracheal epithelia giving rise to breaks, deletions, and sac-like structures. Other tubular organs such as proventriculus, salivary glands, and hindgut are also malformed or missing. These data suggest that Ftl is required for morphogenesis and maintenance of tubular structures of ectodermal origin and underline its similarity in function to a reported lethal mouse knock-out of fat1 where glomerular epithelial processes collapse. Based on our results, we propose a model where Ftl acts as a spacer to keep tubular epithelia apart rather than the previously described adhesive properties of the cadherin superfamily.

Amino Acid Motifs↗

Performance of gross lesions at postmortem for the detection of outbreaks during the avian influenza A virus (H7N7) epidemic in The Netherlands in 2003.

A total of 123 submissions (on average, five birds per submission) from poultry flocks with a suspicion of an infection with highly pathogenic avian influenza virus were investigated at postmortem during the 2003 epidemic in The Netherlands. A total of 86 of these submissions were from infected flocks (positive submissions), and 37 submissions were from non-infected flocks (negative submissions). Peritonitis was the most frequently (62%) recorded pathological finding in positive submissions, followed by tracheitis (43%), oedema of the neck and/or wattles (12%) and (petechial) haemorrhages in the proventriculus (4%). A total of 17% of the positive submissions consisted of birds without any macroscopic lesion. The pathological lesions observed in our study do not fully comply with descriptions in literature, especially the lack of haemorrhagic changes in tissues and low prevalence of oedema of the neck, comb and wattles. It is recommended that if peritonitis, or tracheitis, or oedema of the neck and/or wattles or (petechial) haemorrhages in the proventriculus is observed at postmortem examination, especially if accompanied by an anamnesis describing acute and high mortality in a flock, this should consistently result in follow-up action to exclude highly pathogenic avian influenza in the differential diagnosis as cause of the disease problems by testing tissue samples with an avian influenza-specific laboratory test at the avian influenza reference laboratory.

Animals↗

Localization of myosuppressinlike peptides in the hypocerebral ganglion of two blood-feeding flies: horn fly and stable fly (Diptera:Muscidae).

The insect peptides leucomyosuppressin (pEDVDHVFLRFamide) and dromyosuppressin (TDVDHVFLRFamide) have identical chemical sequences with the exception of the N-terminal amino acid; both inhibit spontaneous contraction of insect visceral muscles. Neurons in the hypocerebral ganglion of horn fly, Hematobia irritans (L.), and stable fly, Stomoxys calcitrans (L.), were found to contain material immunoreactive to antiserum produced against the C-terminal of leucomyosuppressin, but not to the N-terminal of dromyossuppressin. Two large lateral clusters containing 8 cells, linked dorsally and ventrally by 2 chains of 6 cells, encircled the anterior surface of the proventriculus and were immunoreactive of leucomyosuppressin and FMRFamide antisera. Axons from these cells were traced to the wall of the aorta and over the surface of the proventriculus. Ultrastructural analysis revealed these cells contained a singular type of elementary secretory granule that contained material of relatively low electron density, both in the cell body and at the axon terminals.

Animals↗

Sites of organic acid production and pattern of digesta movement in the gastrointestinal tract of geese.

Sixteen geese were used to assess the movement of fluid and particulate digesta through their gastrointestinal tracts and to determine the diurnal variation in organic acid levels for the various segments of the tract. Fluid (polyethylene glycol and chronium-labeled ethylenediaminetetraacetic acid) and particulate markers (2 and 5 mm long) were administered with the meal. Animals were killed at given intervals after the administration of markers. The gastrointestinal tract was divided into nine segments for measurement of markers, pH, volatile fatty acids (VFA), and lactic acid contents. The data indicated a rapid evacuation of fluid marker from the foregut (crop, ventriculus, and proventriculus), while particulate markers were retained for extended periods of time. Retention of fluid marker was observed only within the cecum. Retrograde movement of particulate marker was demonstrated from the duodenum to ventriculus and proventriculus. Retrograde movement of fluid marker was observed from the cloaca to the colon, cecum, and distal third of the small intestine. However, particulate marker showed no retrograde movement in these segments of tract. Highest VFA levels were observed in the cecum. Retention of digesta and production of VFA within the colon were less than those noted for the dog, pig and pony. Lactic acid comprised less than 10% of the organic acids present in the gastrointestinal tract and were at their highest levels in the proximal and mid small intestine.

Animals↗

Toxicological evaluations of cyclopiazonic acid and ochratoxin A in broilers.

The individual and combined effects of ochratoxin A (OA) and cyclopiazonic acid (CPA) were evaluated in Petersen x Hubbard broiler chickens from 1 d to 3 wk of age. The experimental design was a 2 x 2 factorial with treatments of 0 and 2.5 mg OA/kg feed and 0 and 34 mg CPA/kg feed. Production performance, serum biochemistry, and gross pathological observations were evaluated. Body weight gain was reduced (P < 0.05) by OA, CPA, and OA-CPA in combination at the end of 3 wk. Ochratoxin A significantly increased the relative weight of the kidney and serum concentrations of uric acid and triglycerides and decreased total protein, albumin, and cholesterol. The toxicity of CPA was expressed primarily through increased relative weights of the proventriculus and increased activity of creatine kinase. Exposure to OA-CPA was characterized by increased relative weights of the liver, kidney, pancreas, and proventriculus; decreased concentrations of serum albumin, total protein, and cholesterol; increased activity of creatine kinase; and increased concentrations of triglycerides and uric acid. Postmortem examination revealed that the chickens fed CPA or OA-CPA had thickened mucosa and dilated proventricular lumen. Data from this study demonstrate that OA, CPA, and the OA-CPA combination can limit broiler performance and adversely affect broiler health. The interaction of the compounds was primarily additive or less than additive in the parameter in which the interaction occurred.

Animal Feed↗

Intestinal mesenchyme provokes differentiation of intestinal endocrine cells in gizzard endoderm.

The gizzard (muscular stomach) of chicks is deficient in endocrine cells at hatching. It has previously been shown that proventricular types and proportions of endocrine cells can be induced in gizzard endoderm under the influence of proventricular (glandular stomach) mesenchyme. In order to test its capacity to form nongastric endocrine cell types, gizzard endoderm of 3.75- to 5-day chick embryos was combined with mesenchyme from the small intestine of 3.5- to 4-day quail embryos. The combinations were grown as chorio-allantoic grafts until they attained an incubation age comparable to that of hatching chicks. Controls comprised reassociated endoderm and mesenchyme of chick gizzard and of quail intestine. In the experimental grafts, morphogenesis was predominantly intestinal but some grafts showed gizzard-like features, particularly if the endoderm had been provided by older donors. All intestinal endocrine cell types, including those also found in the normal proventriculus (serotonin-, glucagon-, pancreatic polypeptide-, neurotensin- and somatostatin-immunoreactive cells) differentiated in experimental grafts, some even where morphogenesis was gizzard-like. Hence progenitors of not only gastric, but also intestinal, endocrine cells are indeed present in gizzard endoderm. The possibility that gizzard mesenchyme is inhibitory to endocrine cell differentiation is mooted. Motilin- and secretin-immunoreactive cells, which are characteristic of the intestine but not of the proventriculus of chicks at hatching, were respectively sparse or absent when the endoderm was derived from older donors. Thus the ability of gizzard endoderm to differentiate into nongastric endocrine cell types declines before its capacity to form gastric types. The unexpected appearance of gastrin-releasing peptide (GRP)-immunoreactive cells, a proventricular type not found in normal chick intestine, suggests that the intestinal mesenchyme, at least in this instance, was exercising a permissive role.

Animals↗

Functional specification in the Drosophila endoderm.

The discovery of homeobox gene clusters led us to realize that the mechanisms for body patterning and other developmental programs are evolutionally-conserved in vertebrates and invertebrates. The endoderm contributes to the lining of the gut and associated organs such as the liver and pancreas, which are critical for physiological functions. Our knowledge of endoderm development is limited; however, recent studies suggest that cooperation between the HNF3/Fork head and GATA transcription factors is crucial for endoderm specification. It is necessary to further understand the mechanism through which cells become functionally organized. Molecular genetic analyses of the Drosophila endoderm would provide insights into this issue. During proventriculus morphogenesis, a simple epithelial tube is folded into a functional multilayered structure, while two functions of midgut copper cells (i.e. copper absorption and acid secretion) can be easily visualized. The homeobox gene defective proventriculus (dve) plays key roles in these functional specifications.

Animals↗

Left-right asymmetry in the alimentary canal of the Drosophila embryo.

In many animal groups, left-right (LR) asymmetry within the body is observed. The left and right sides of the body are generally defined with reference to the anterior-posterior (AP) and dorsal-ventral (DV) axes. In this study, we investigated whether LR asymmetry is solely dependent on the AP and DV polarities in Drosophila embryos. We focused on the proventriculus, a posterior part of the foregut, and the hindgut because LR asymmetry in these body parts is highly stable in normal embryos. In embryos with a fully reversed AP polarity, LR asymmetry in both the proventriculus and the hindgut was re-oriented in relation to the reversed AP polarity. This demonstrates that inversion of AP polarity does not affect LR asymmetry of these tissues, and implies that LR asymmetry is specified in relation to the AP and DV polarities. Our findings were not consistent with the alternative hypothesis that LR asymmetry is predetermined by maternal signals that localize asymmetrically along the LR axis in the oocyte and/or early embryo.

Animals↗

Occurrence and polymorphism of bombesin-like peptides in the gastrointestinal tract of birds and mammals.

The gastrointestinal tract of mammals and birds, especially stomach and upper small intestine, contains bombesin-like peptides. This has been unequivocally demonstrated by radioimmunoassay and bioassay. Concentrations of bombesin-like activity may range from a few ng to 500-600 ng per g fresh tissue. Last values refer to the chicken proventriculus, which has been the object of a more thorough investigation. The bombesin-like peptide of the chicken proventriculus showed a marked heterogeneity. All forms probably stem from a pro-bombesin, a large precursor molecule which is insoluble in methanol, acetone, and even boiling water, but may be cleaved by acid hydrolysis. Methanol extracts contain at least two forms of the bombesin-like peptide; HCl extracts at least three forms; HCl extracts of the residue of methanol extraction at least four forms. Whereas some forms--for example, the methanol extractable forms--probably pre-exist in the tissue, other forms may be artefacts arising from acid treatment. The various forms may be distinguished from each other not only by their elution profile, but also by bioassay. In fact, though all forms show the activity spectrum characteristic for the amphibian bombesin-like peptides, they present considerable quantitative differences in activity. Pro-bombesin(s) probably occur also in the rat and guinea-pig stomach; similarly, a clear-cut heterogeneity is appreciable for the bombesin-like peptide of the human gastric mucosa.

Animals↗

Effect of pH, temperature and incubation period on the study of intrinsic innervation of various organs by thiocholine technique in certain vertebrates.

The adrenergic fibres can be demonstrated if the optimal time and temperature are maintained. The cholinergic fibres were demonstrated at pH 5.2, incubation period l4h and temperature 37 degrees C. To ascertain the cholinesterase activity in tongue, heart, lung(birds, mammals and reptiles), gizzard and proventriculus (birds) and penis (rat and embryo of squirrel), pH 4.9, 16h incubation and temperature 37 degrees C, were quite suitable. It was possible to demonstrate the intrinsic innervation in tongue, lung, heart (mammals, birds and reptiles), gizzard and proventriculus (birds) and penis (rat and embryo of squirrel), at pH 5.2, 20h incubation and temperature 40 degrees C.

Animals↗

Pathological and immunohistochemical findings in American crows (Corvus brachyrhynchos) naturally infected with West Nile virus.

Twenty-one American crows were identified as being West Nile virus (WNV) infected by WNV-specific reverse transcriptase-polymerase chain reaction (RT-PCR) performed on fresh brain tissue (cerebrum and cerebellum of 16 crows) or by WNV-specific immunohistochemistry of various organs (21 crows). Consistent gross lesions attributable to WNV infection were not detected. Common histological lesions included necrosis of spleen and bone marrow. West Nile virus antigen was consistently detected in heart and kidney (100%). In addition, bone marrow (92%), duodenum (89%), proventriculus (87%), liver (86%), lung (85%), spleen (80%), pancreas (61%), and brain (45%) contained WNV antigen-positive cells. Infected cells included cardiomyocytes; neurons; endothelial cells and vascular smooth muscle cells; hematopoietic cells of bone marrow; and macrophages of spleen, liver (Kupffer cells), and lungs. Epithelial cells of renal tubules, duodenum, pancreas, and proventriculus were also infected. The diagnostic histopathologist should consider WNV infection in crows in the absence of any inflammatory lesions. Immunohistochemistry of heart and kidney is as reliable in detecting WNV infection in American crows as RT-PCR of fresh brain tissue.

Animals↗

Gastrointestinal pathogenicity of adenoviruses and reoviruses isolated from broiler chickens in Alabama.

Adenoviruses and reoviruses isolated from commercial broiler chickens were evaluated for gastrointestinal pathogenicity in specific-pathogen-free Leghorn chickens. The viruses were originally isolated from either the proventriculus or a gastrointestinal pool of tissues of broiler chickens with proventriculitis or enteritis. Isolates were cloned by terminal dilution. Day-old chickens were inoculated by oral and ocular routes with undiluted tissue culture fluids (titers of 10[2]-10[4] TCID50/ml) and then examined at necropsy on days 5, 10, and 15 postinoculation. Chickens in all virus groups (but not the control group) developed wet, unformed fecal droppings that persisted for the duration of the study. Mild lesions occurred in reovirus-inoculated chickens and included hyperplasia of lymphocyte aggregates in various organs and mild gizzard erosions. Chickens inoculated with adenovirus isolates developed marked gizzard erosions and necrotizing pancreatitis as well as mild proventriculitis. Intranuclear viral inclusion bodies occurred in gizzard epithelium and pancreatic acinar cells at the sites of lesions. Lymphocytic atrophy occurred in the bursa of Fabricius. Respective viruses were reisolated from proventriculus and duodenum collected from chickens of each group; no viruses were isolated from controls. Under the conditions of this study, adenovirus isolates were more pathogenic than the reovirus isolates in the digestive system.

Adenoviridae Infections↗

Target organs for avian pancreatic polypeptide.

The problem of the physiological function of pancreatic polypeptide (PP) has been approached by attempting to identify target organs. Avian PP (aPP) labeled with 125I at either the C-terminus (aPP-C) or the N-terminus (aPP-N) was injected into fasted chickens and allowed to circulate for 3-120 min. At the end of the equilibration period, the anesthetized bird was perfused first with saline, then with Buoin's solution. Samples of fixed tissue from various organs were collected, weighed, and counted. Control experiments consisted of coinjection of unlabeled aPP to compete for receptors. The rate of disappearance of aPP-N from plasma was greater than that of aPP-C. Binding of aPP-N by spleen, duodenum, ileum, pancreas, and bone marrow was markedly reduced by coinjection of unlabeled aPP. A similar but less marked reduction in binding was found in liver and proventriculus. aPP-C gave less conclusive results. The maximal competitive effect of unlabeled PP could be achieved in most cases with 30 microgram unlabeled aPP. It is concluded that pancreas, duodenum, ileum, spleen, and bone marrow, and probably liver and proventriculus, are target organs for aPP in the chicken and that the C-terminal region of aPP is involved in receptor binding.

Animals↗