Anatomical, histological, and scanning electron microscopic features of the esophagus and crop in young and adult domestic pigeons (Columba livia Domestica).

BACKGROUND: Pigeons (Columba livia) are mainly raised as a source of animal protein, racing, leisure and as an experimental animal. The present study investigated the morphology of the esophagus in the young and adult domestic pigeon, Columba livia domestica. METHODS: Ten young and ten adult, normal, and healthy pigeons were collected from the local breeders. Samples from different parts of esophagus and crop were examined grossly, by stereomicroscopy, scanning and light microscopy. RESULTS: The esophagus consisted of a long cervical part, a crop, and a short thoracic part. The crop was represented by a thin-walled outpouching with two lateral diverticula. The mucosa presented wavy fine folds in the cervical esophagus, irregular folds in the lateral diverticula giving it a corrugated appearance, and prominent longitudinal folds with several gland openings in the middle and lower parts of the crop, as well as in the thoracic esophagus. The density of gland openings was higher in adult pigeons than that in young pigeons. The mucosa of the esophagus was lined by non-keratinized stratified squamous epithelium. The shape, height, and branching of the mucosal folds differed between young and adult pigeons. Mucous-secreting alveoli were detected in the middle part of the crop as well as in the thoracic esophagus, but not in the cervical esophagus or lateral diverticula of the crop. CONCLUSION: The variations between the young and adult pigeons suggest a functional adaptation of adult pigeons to their diet compared to young pigeons.

Antigenic epitope targets of rhesus macaques self-curing from Schistosoma mansoni infection.

The self-cure of rhesus macaques from a schistosome infection and their subsequent strong immunity to a cercarial challenge should provide novel insights into the way these parasites can be eliminated by immunological attack. High-density arrays comprising overlapping 15-mer peptides from target proteins printed on glass slides can be used to screen sera from host species to determine antibody reactivity at the single epitope level. Careful selection of proteins, based on compositional studies, is crucial to encompass only those exposed on or secreted from the intra-mammalian stages and is intended to focus the analysis solely on targets mediating protection. We report the results of this approach using two pools of sera from hi- and lo-responder macaques undergoing self-cure, to screen arrays comprising tegument, esophageal gland, and gastrodermis proteins. We show that, overall, the target epitopes are the same in both groups, but the intensity of response is twice as strong in the high responders. In addition, apart from Sm25, tegument proteins elicit much weaker responses than those originating in the alimentary tract, as was apparent in IFNγR KO mice. We also highlight the most reactive epitopes in key proteins. Armed with this knowledge, we intend to use multi-epitope constructs in vaccination experiments, which seek to emulate the self-cure process in experimental animals and potentially in humans.

Immunoreactive Proteins in the Esophageal Gland Cells of Anisakis Simplex Sensu Stricto Detected by MALDI-TOF/TOF Analysis.

In plant and animal nematode parasites, proteins derived from esophageal gland cells have been shown to be important in the host-nematodes relationship but little is known about the allergenic potential of these proteins in the genus Anisakis. Taking into account the increase of anisakiasis and allergies related to these nematodes, immunoreactive properties of gland cell proteins were investigated. Two hundred ventricles were manually dissected from L3 stage larvae of Aniskakis simplex s.s. to allow direct protein analysis. Denaturing gel electrophoresis followed by monochromatic silver staining which revealed the presence of differential (enriched) proteins when compared to total nematode extracts. Such comparison was performed by means of 1D and 2D electrophoresis. Pooled antisera from Anisakis spp.-allergic patients were used in western blots revealing the presence of 13 immunoreactive bands in the ventricular extracts in 1D, with 82 spots revealed in 2D. The corresponding protein bands and spots were excised from the silver-stained gel and protein assignation was made by MALDI-TOF/TOF. A total of 13 (including proteoforms) were unambiguously identified. The majority of these proteins are known to be secreted by nematodes into the external environment, of which three are described as being major allergens in other organisms with different phylogenetic origin and one is an Anisakis simplex allergen.

Antigenic epitope targets of rhesus macaques self-curing from Schistosoma mansoni infection

The self-cure of rhesus macaques from a schistosome infection and their subsequent strong immunity to a cercarial challenge should provide novel insights into the way these parasites can be eliminated by immunological attack. High-density arrays comprising overlapping 15-mer peptides from target proteins printed on glass slides can be used to screen sera from host species to determine antibody reactivity at the single epitope level. Careful selection of proteins, based on compositional studies, is crucial to encompass only those exposed on or secreted from the intra-mammalian stages and is intended to focus the analysis solely on targets mediating protection. We report the results of this approach using two pools of sera from hi- and lo-responder macaques undergoing self-cure, to screen arrays comprising tegument, esophageal gland, and gastrodermis proteins. We show that, overall, the target epitopes are the same in both groups, but the intensity of response is twice as strong in the high responders. In addition, apart from Sm25, tegument proteins elicit much weaker responses than those originating in the alimentary tract, as was apparent in IFNγR KO mice. We also highlight the most reactive epitopes in key proteins. Armed with this knowledge, we intend to use multi-epitope constructs in vaccination experiments, which seek to emulate the self-cure process in experimental animals and potentially in humans.

In situ Hybridization of Plant-parasitic Nematode Globodera pallida Juveniles to Detect Gene Expression.

In this study, we describe a standard whole mount in situ hybridization method which is used to determine the spatial-temporal expression pattern of genes from Globodera spp. Unlike more invasive radioactive labeling approaches, this technique is based on a safe, highly specific enzyme-linked immunoassay where a Digoxigenin (DIG)-tagged anti-sense probe hybridized to a target transcript is detected by anti-DIG antibodies conjugated with alkaline phosphatase enzyme (AP) (anti-DIG-AP). The hybrid molecules are visualized through an AP-catalyzed color reaction using as the substrate 5-bromo-4-chloro-3-indolyl phosphate (BCIP) and nitro blue tetrazolium chloride (NBT). This method can be applied to both free-living pre-parasitic juveniles and early endoparasitic stages of cyst nematodes.

CDX2 protein expression compared to alcian blue staining in the evaluation of esophageal intestinal metaplasia.

AIM: To compare the sensitivity and specificity of CDX2 and alcian blue (AB) pH 2.5 staining in identifying esophageal intestinal metaplasia. METHODS: One hundred and ninty-nine biopsies from 186 patients were retrospectively reviewed and categorized as Barrett's esophagus (BE) (n = 108); non-Barrett's esophagus (NBE) (n = 48); columnar blue cells (CB) and esophageal glands (EG) (n = 43). The biopsies were stained with AB and immunostained for CDX2 using a mouse monoclonal antibody from Biogenex (clone CDX2-88) and the Ventana Discovery X automated immunostainer. The positive and negative predictive value of each group was used to determine the predictive power of CDX2 and AB in diagnosing intestinal metaplasia. RESULTS: All of the 108 BE biopsies (100%) were positive for AB and 102 of them (94.4%) were positive for CDX2. The six BE patients (5.6%) who failed to stain with CDX2 were found to have lost the focus of intestinal metaplasia upon deeper sectioning for immunostaining. Both AB and CDX2 were negative in 43 out of 48 (89.6%) NBE cases. Five NBE patients (10.4%) were falsely positive for AB due to the presence of EG and CB in these biopsies. These cases were all CDX2 negative. In addition, 5 AB negative NBE were found to be CDX2 positive. Based on these results the CDX2 immunostain had similar sensitivity but higher specificity (100% vs about 91%) than AB in detecting intestinal type metaplasia in these samples. Our data shows that CDX2 has a better PPV in detecting intestinal metaplasia as compared to AB (95.6% vs 71.5%, respectively). CONCLUSION: CDX2 has a better positive predictive value than AB in detecting intestinal metaplasia. CDX2 may be useful when challenged by gastro-esophageal biopsies containing mimikers of BE.

Electron Microscopy of the Stomatostylet and Esophagus of Criconemoides curvatum.

The stomatostylet of Criconemoides curvature consists of three parts: tooth cone, shaft, and knobs. The tooth cone constitutes the outer conical covering and inner lining of the anterior half of the stylet lumen. The tooth cone is easily separated from the shaft by treating an isolated styler with 0.5% sodium hypochlorite. The posterior half of the shaft is cylindrical, tapering anteriorly to form the shaft extension, wedged between the inner and outer tooth cone. The shaft extension extends to the stylet lumen orifice, which is subterminal and ventral. Six ducts enter the shaft through the junction between the shaft and knobs. They extend anteriad toward the tip of the shaft extension. Cytoplasmic connections between the ducts and the cells surrounding the stylet occur near tile junction between the shaft and the basal knobs. Ribosome and membranous structures are observed in these ducts. The esophagus of the adult female consists of a fused procorpus and metacorpus with a large valve possessing thickened cuticular walls at the anterior and posterior ends. The dorsal esophageal gland reservoir is composed of many honeycomb-like compartments made up of two types of differing electron density. The subventral esophageal glands, however, consist of only one type of granules. Both dorsal and subventral esophageal glands open into the esophageal lumen through trachea-like branched multiple canals.

Post-infection Development and Morphology of Meloidogyne cruciani.

The development and life stages of Meloidogyne cruciani on tomato was studied at 28 C. Roots of 2-wk-old 'Rutgers' tomato seedlings were exposed to inoculum for 24 h, rinsed, and the seedlings repotted. No major changes in juvenile development were observed prior to 8 days after inoculation. At 11 days the second-stage juvenile had enlarged considerably. The genital primordium had not yet asumed the V-shape characteristic of developing females, but the presence of rectal glands identified the juveniles as females. At this time (11 days), two additional, previously undescribed esophageal lobes were first observed; they were adjacent to the dorsal and subventral glands. After molting from second to third stage, the stylet cone, shaft, and the lumen of the stylet knobs are shed and remain attached to the second-stage cuticle. The excretory duct of the third-stage juveniles was directed anteriorly from the excretory pore of the second-stage cuticle and appear attached to the body wall of the third-stage juveniles opposite the procorpus. At 19 days after inoculation, the last molt took place. The adult female possessed a new stylet, a large five-gland esophagus, a prominent excretory system ending in a unicellular gland and a fully developed reproductive system.

Changes in Esophageal Gland Activity During the Life Cycle of Nacobbus aberrans (Nemata: Pratylenchidae).

Electron and light microscopy were used to study the dorsal gland (DG) and the two subventral glands (SvG) of seven developmental phases of Nacobbus aberrans: pre-parasitic second-stage juveniles (J2), parasitic J2, third- (J3) and fourth- (J4) stages, migratory females, young sedentary females, and mature sedentary females. In each developmental phase the level of esophageal gland activity, was estimated by the abundance of organelles associated with secretory pathways, including endoplasmic reticulum, ribosomes, Golgi, multivesicular bodies, and secretory granules. All esophageal glands were metabolically active in all J2 examined, although only in parasitic J2 were there numerous secretory granules in the esophageal gland extensions and ampullae. No evidence of secretory activity was observed in the esophageal glands of the coiled and relatively inactive J3 and J4, nor in migratory females; these stages apparently do not feed. Observations suggest that reserves stored by J2 sustain three ecdyses and the migratory female's search for a feeding site and induction of a syncytium. Feeding activity is resumed in young and mature sedentary females, in which the DG is highly active and enlarged. The SvG are metabolically active, but with little synthesis of secretory granules, suggesting that in sedentary females the SvG may have physiological roles other than digestion.

Pararotylenchus n. gen. (Pararotylenchinae n. subfam., Hoplolaimidae) with Six New Species and Two New Combinations.

A new subfamily, Pararotylenchinae, (Hoplolaimidae) is described. It includes a single genus, Pararotylenchus n. gen., six new species, attd two new combinations, Pararotylenchus (syn. Tylenchorhynchus) brevicaudatus (Hopper, 1959) n. comb. and Pararotylenchus (syn. Rotylenchus) pint (Mamiya, 1968) n. comb. Pararotylenchinae is similar to certain other Hoplolaimidae, such as Rotylenchinae, with respect to most characters including the short tail, the position of the phasmids near the anus, and the relatively great distance of the dorsal gland orifice from the base of the stylet knobs. The lip region, as observed with the scanning electron microscope, conforms to the basic pattern for Hoplolaimidae. The labial disc is round with slit-like amphidial openings at the lateral sides of the periphery of the disc. The distinctive anterior-most lip annule is segmented into six sectors. Unlike other Hoplolaimidae, however, the esophageal glands of Pararotylenchinae form a basal bulb similar to that of Tylenchorhynchidae; other characters do not resemble Tylencborhynchidae. Specimens of Pararotylenchus have been recovered only from cool regions at high elevations and Pacific coastal areas in the Western United States, Japan, and Korea.

Meloidogyne cruciani n. sp. a Root-knot Nematode from St. Croix (U.S. Virgin Islands) with Observations on Morphology of This and Two Other Species of the Genus.

Meloidogyne cruciani n. sp. infecting tomato (Lycopersicon esculentum Mill.) in the U.S. Virgin Islands is described and illustrated. M. cruciani is distinguished from other species of the genus by having punctations around the anus of the female and by the second-stage juveniles possessing tri-lobed esophageal glands which are longer than most other species, with their posterier end at about 46.4% of the body length. The esophageal glands of the immature and adult females are contained in five separate lobes.