• View in gallery

    Illustration of the position of a pigeon (Columba livia) undergoing gastroscopy (A) and close-up illustration of biopsy forceps being used to obtain a mucosal specimen from the proventriculus (B). A—The endoscope has been inserted through the oral cavity into the proventriculus, and the approximate position of the gastrointestinal tract compartments is shown. B—Notice that the forceps are positioned nearly parallel to the long axis of the proventriculus. (Medical illustration by Kip Carter, University of Georgia, 2016. Reprinted with permission.)

  • View in gallery

    Representative endoscopic images of the upper gastrointestinal tract of pigeons. A—Cervical portion of the esophagus. B—Crop infused with saline (0.9% NaCl) solution. C—Crop, with opening to the thoracic portion of the esophagus in the center. D—Lumen of the proventriculus. E—Close-up view of the proventricular mucosa. Notice the gastric glands. F—Biopsy forceps positioned parallel against the proventricular mucosa during biopsy specimen collection. G—Hemorrhage that occurred after proventricular biopsy. A clot developed rapidly. H—Lumen of the proventriculus. Notice the mucous covering. This was a normal finding, with no associated histologic lesions. I—Contracted isthmus separating the ventriculus. J—Entry into the ventriculus. Notice the grit and green coloration of the koilin, which were normal findings. K—Biopsy forceps in the process of obtaining a sample perpendicular to the ventriculus. L—Ventriculus following collection of 2 biopsy specimens. Notice the koilin defect and minimal mucosal hemorrhage.

  • View in gallery

    Representative photomicrographs of biopsy specimens collected during gastroscopy in pigeons. A—Mucosal specimen obtained from the proventriculus. This specimen is considered of adequate thickness and includes both plicated surface epithelium and underlying glands. B—Mucosal specimen obtained from the proventriculus. This sample is considered of inadequate thickness and includes only plicated surface epithelium. C—Mucosal specimen obtained from the ventriculus. This sample is considered of full thickness and includes koilin, mucosa, and tunica muscularis. D—Mucosal specimen obtained from the ventriculus. This specimen is considered inadequate and includes only koilin. H&E stain; bar = 200 μm.

  • 1. Sum S, Ward CR. Flexible endoscopy in small animals. Vet Clin North Am Small Anim Pract 2009;39: 881902.

  • 2. Langlois I. The anatomy, physiology, and diseases of the avian proventriculus and ventriculus. Vet Clin North Am Exot Anim Pract 2003;6: 85111.

    • Search Google Scholar
    • Export Citation
  • 3. Anjos BL, Diefenbach A, Rissi DR, et al. Anaplastic ventricular adenocarcinoma in a blue and gold macaw (Ara ararauna). Braz J Vet Pathol 2012;5: 7880.

    • Search Google Scholar
    • Export Citation
  • 4. De Voe R, Degernes L, Karli K. Dysplastic koilin causing proventricular obstruction in an eclectus parrot (Eclectus roratus). J Avian Med Surg 2003;17: 2732.

    • Search Google Scholar
    • Export Citation
  • 5. Gibbons PM, Busch MD, Tell LA, et al. Internal papillomatosis with intrahepatic cholangiocarcinoma and gastrointestinal adenocarcinoma in a peach-fronted conure (Aratinga aurea). Avian Dis 2002;46: 10621069.

    • Search Google Scholar
    • Export Citation
  • 6. Leach MW, Paul-Murphy J, Lowenstine LJ. Pet bird medicine. Three cases of gastric neoplasia in psittacines. Avian Dis 1989;33: 204210.

    • Search Google Scholar
    • Export Citation
  • 7. Yonemaru K, Sakai H, Asaoka Y, et al. Proventricular adenocarcinoma in a Humboldt penguin (Spheniscus humboldti) and a great horned owl (Bubo virginianus); identification of origin by mucin histochemistry. Avian Pathol 2004;33: 7781.

    • Search Google Scholar
    • Export Citation
  • 8. Campbell TW, Turner O. Carcinoma of the ventriculus with metastasis to the lungs in a sulphur-crested cockatoo (Cacatua galerita). J Avian Med Surg 1999;13: 265268.

    • Search Google Scholar
    • Export Citation
  • 9. Divers SJ. Avian diagnostic endoscopy. Vet Clin North Am Exot Anim Pract 2010;13: 187202.

  • 10. Mejia-Fava J, Divers SJ, Jiménez D, et al. Diagnosis and treatment of proventricular nematodiasis in an umbrella cockatoo (Cacatua alba). J Am Vet Med Assoc 2013;242: 11221126.

    • Search Google Scholar
    • Export Citation
  • 11. Stahl SJ, Hernandez-Divers SJ, Cooper TL. Evaluation of trans-cutaneous pulmonoscopy for examination and biopsy of the lungs of ball pythons and determination of preferred biopsy specimen handling and fixation procedures. J Am Vet Med Assoc 2008;233: 440445.

    • Search Google Scholar
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Evaluation of gastroscopy and biopsy of the proventriculus and ventriculus in pigeons (Columba livia)

Izidora Sladakovic BVSc, MVS1, Angela E. Ellis DVM, PhD2, and Stephen J. Divers BVetMed, DZooMed3
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  • 1 Department of Small Animal Medicine and Surgery, University of Georgia, Athens, GA 30602.
  • | 2 Department of Pathology, University of Georgia, Athens, GA 30602.
  • | 3 Department of Small Animal Medicine and Surgery, University of Georgia, Athens, GA 30602.

Abstract

OBJECTIVE To evaluate the efficacy and safety of gastroscopy and biopsy of the proventriculus and ventriculus in pigeons (Columba livia).

ANIMALS 15 adult pigeons.

PROCEDURES Each pigeon was anesthetized, and the upper gastrointestinal tract (from the cervical portion of the esophagus to the ventriculus) was endoscopically evaluated by use of a rigid endoscope inserted orally. Saline (0.9% NaCl) solution was orally infused to achieve lumen dilation and visibility. Two mucosal biopsy specimens were collected from each of the proventriculus and ventriculus, histologically evaluated, and graded for crush artifacts and depth. Pigeons were monitored for adverse effects for 3 to 6 days after the procedure, after which they were euthanized for necropsy.

RESULTS Gastroscopy via the oral approach provided excellent visibility of the lumen and mucosal surfaces of the proventriculus and cranial portion of the ventriculus and was safe provided that appropriate precautions were taken. Two intraoperative deaths occurred at the beginning of the study; following procedure refinement, no additional deaths occurred. No major adverse effects of the procedure were detected in the remaining 13 pigeons during the postoperative monitoring period or at necropsy. Diagnostic quality of proventriculus specimens was adequate for 10 of 13 pigeons. Eight of 13 ventriculus specimens were of inadequate quality, and only 3 were of adequate quality.

CONCLUSIONS AND CLINICAL RELEVANCE Gastroscopy was useful for evaluating the lumen and mucosal surface of the proventriculus and ventriculus in pigeons, and biopsy of those organs was safely performed with the appropriate technique. Further evaluation of these techniques is needed in birds with clinical disease and birds of other species.

Abstract

OBJECTIVE To evaluate the efficacy and safety of gastroscopy and biopsy of the proventriculus and ventriculus in pigeons (Columba livia).

ANIMALS 15 adult pigeons.

PROCEDURES Each pigeon was anesthetized, and the upper gastrointestinal tract (from the cervical portion of the esophagus to the ventriculus) was endoscopically evaluated by use of a rigid endoscope inserted orally. Saline (0.9% NaCl) solution was orally infused to achieve lumen dilation and visibility. Two mucosal biopsy specimens were collected from each of the proventriculus and ventriculus, histologically evaluated, and graded for crush artifacts and depth. Pigeons were monitored for adverse effects for 3 to 6 days after the procedure, after which they were euthanized for necropsy.

RESULTS Gastroscopy via the oral approach provided excellent visibility of the lumen and mucosal surfaces of the proventriculus and cranial portion of the ventriculus and was safe provided that appropriate precautions were taken. Two intraoperative deaths occurred at the beginning of the study; following procedure refinement, no additional deaths occurred. No major adverse effects of the procedure were detected in the remaining 13 pigeons during the postoperative monitoring period or at necropsy. Diagnostic quality of proventriculus specimens was adequate for 10 of 13 pigeons. Eight of 13 ventriculus specimens were of inadequate quality, and only 3 were of adequate quality.

CONCLUSIONS AND CLINICAL RELEVANCE Gastroscopy was useful for evaluating the lumen and mucosal surface of the proventriculus and ventriculus in pigeons, and biopsy of those organs was safely performed with the appropriate technique. Further evaluation of these techniques is needed in birds with clinical disease and birds of other species.

Gastroscopy with collection of gastric biopsy specimens is routinely performed in small animal medicine in the evaluation of gastric disorders, including identification of foreign bodies, ulceration, and neoplastic and infectious diseases.1 In birds, the proventriculus and ventriculus can be affected by various diseases, including those listed for small animals.2 These diseases can be difficult to distinguish and diagnose without performing a necropsy.2 Several case reports have been published regarding the clinical progression of a proventricular or ventricular disorder in birds, with a definitive diagnosis established only at necropsy3–7 or via coeliotomy,8 particularly in birds with neoplastic disease.

Endoscopy is a minimally invasive diagnostic tool that enables visual examination of various organs in birds and collection of biopsy specimens to obtain a diagnosis.9 Endoscopy-guided biopsy of the proventriculus reportedly facilitated antemortem diagnosis and successful treatment of proventricular nematodiasis in an umbrella cockatoo (Cacatua alba).10 However, to the authors’ knowledge, no studies have been conducted to evaluate the usefulness and safety of gastroscopy and endoscopically assisted collection of biopsy specimens from the proventriculus and ventriculus in birds. The objective of the study reported here was to evaluate the usefulness and safety of gastroscopy and biopsy of the proventriculus and ventriculus via an oral approach in pigeons (Columba livia).

Materials and Methods

Animals

Fifteen pigeons of various ages and both sexes were used in the study. The pigeons had been selected for culling from a breeding flock and were scheduled for euthanasia following an educational workshop. Each had been allowed to acclimatize to the research facility for 7 days prior to the start of the study. Pigeons were housed in indoor cages in groups of 3, with the environmental temperature maintained at 22° to 24°C and a 12-hour light, 12-hour dark cycle provided. They were given free access to a pelleted dieta and water. Food but not water was withheld overnight for 10 to 12 hours before the study procedures began. Also before the procedures began, pigeons were examined and weighed. The study protocol was approved by the University of Georgia Institutional Animal Care and Use Committee (protocol No. A2014 09-003-Y1-A0).

Anesthesia

Prior to gastroscopy, each pigeon was premedicated with butorphanol tartrateb (2 mg/kg, IM) and anesthesia was induced with 5% isofluranec in oxygen delivered via a facemask connected to a nonrebreathing circuit. Endotracheal intubation was performed with a Cole endotracheal tube (internal diameter, 2.5 mm). Anesthesia was maintained with isoflurane in oxygen, delivered by use of an intermittent positive pressure ventilator.d Isoflurane concentration was adjusted as necessary to maintain a surgical depth of anesthesia.

During anesthesia, pigeons were monitored for heart rate, respiration rate, end-tidal carbon dioxide concentration, and reflexes. Heart rate was measured by placing a Doppler ultrasonographic flow detectore over the brachial artery or by direct cardiac auscultation. End-tidal carbon dioxide concentration and respiration rate were monitored with a capnographf and by direct observation, respectively. Thermal support was provided by use of a recirculating warm water pad and pump.g Prior to starting gastroscopy, a blood sample was collected from a basilic vein for measurement of PCV and plasma total solids concentration.

Gastroscopy and biopsy specimen collection

Anesthetized pigeons were positioned in dorsal recumbency, with the head facing the person performing gastroscopy, tilted approximately 30° to 45°, and elevated (reverse Trendelenburg position; Figure 1). A small strip of self-adhesive bandage was placed around the neck to reduce backflow of infused saline (0.9% NaCl) solution into the cervical portion of the esophagus and oral cavity, and a strip of adhesive tape was placed across the neck to aid positioning. A 12 or 14F red rubber tube, attached to a 60-mL catheter-tip syringe, was passed through the oral cavity into the crop. Then, 30 mL of warm (40°C) sterile saline solution was infused to flush the lumen, and the infused saline solution was removed by aspiration. This process was repeated until the aspirated fluid appeared clear of debris. The red rubber tube was then advanced caudally into the proventriculus and, subsequently, the ventriculus, and the process repeated as above.

Figure 1—
Figure 1—

Illustration of the position of a pigeon (Columba livia) undergoing gastroscopy (A) and close-up illustration of biopsy forceps being used to obtain a mucosal specimen from the proventriculus (B). A—The endoscope has been inserted through the oral cavity into the proventriculus, and the approximate position of the gastrointestinal tract compartments is shown. B—Notice that the forceps are positioned nearly parallel to the long axis of the proventriculus. (Medical illustration by Kip Carter, University of Georgia, 2016. Reprinted with permission.)

Citation: American Journal of Veterinary Research 78, 1; 10.2460/ajvr.78.1.42

Irrigation for the gastroscopy procedure was provided by suspending a 1-L bag of warm sterile saline solution above the operating table and connecting the IV administration set to the ingress port of the sheath. A second IV administration set was connected to the egress port, and the end was placed under the table into a bucket. Both administration sets were primed with sterile saline solution before starting the procedure. A 2.7-mm × 18-cm, 30°C telescopeh housed within a 4.8-mm operating sheathi was connected to a xenon light source,j endovideo camera,k monitor, and digital capture device.l The telescope-sheath unit was then introduced through the oral cavity and advanced into the crop.

Irrigation with warm sterile saline solution was performed to reduce the risk of iatrogenic hypothermia and to dilate the lumen and allow visual examination of the mucosal surfaces. Inflow and outflow of saline solution was adjusted by turning the ingress and egress ports on and off as needed to provide good visibility while avoiding administration of an excess of solution. The crop was examined, and then the endoscope was advanced through the thoracic portion of the esophagus and into the proventriculus. The lumen and mucosa of the proventriculus were examined (Figure 1).

For the proventriculus biopsy procedure, a representative area of the proventriculus was identified and 5F biopsy forcepsm were introduced through the instrument channel of the operating sheath until they appeared in the center of the endoscopic field of view. One mucosal biopsy specimen was collected, and the biopsy forceps were withdrawn. The procedure was repeated to obtain a second specimen. The first time this procedure was performed, the proventriculus was perforated when specimen collection was attempted with the proventricular lumen slightly collapsed and the jaws of the forceps grasping a mucosal fold perpendicular to the proventricular wall. The perforation was appreciated by rapid movement of saline solution through the biopsy site and collapse of the proventricular lumen. The affected pigeon was immediately euthanized and excluded from the study. The biopsy technique was subsequently changed, and mucosal biopsy specimens were obtained by distending the proventricular lumen and performing the biopsy with the forceps nearly parallel to the long axis of the proventriculus (Figure 1).

For the ventricular biopsy procedure, the telescope-sheath unit was then advanced caudally into the ventriculus. A standard technique, with the biopsy forceps placed perpendicular to the wall, was used for the ventriculus. After the ventriculus was examined, 5F biopsy forceps were introduced through the instrument channel of the operating sheath until they appeared in the center of the endoscopic field of view. One biopsy specimen was collected of the koilin layer, and the biopsy forceps were withdrawn. Biopsy forceps were introduced again and inserted through the koilin defect, and a second specimen was collected of the mucosal layer. The procedure was repeated to obtain 2 more specimens.

Each collected tissue specimen was immediately transferred to a sterile red-top tube containing 2 mL of sterile saline solution by opening the jaws of the biopsy forceps and gently shaking the forceps to allow the specimen to fall into the solution. Specimens were then gently transferred to a biopsy cassette and placed in jars containing neutral-buffered 10% formalin.

Representative images and video recordings were obtained. All procedures, other than the first procedure involving the pigeon that was euthanized, were performed by the same author (IS), who was a first-year zoological medicine resident at the time the study was conducted. The ability to visually inspect the gastric compartments was graded as excellent, satisfactory, or impossible. Excellent was defined as clear visibility of the lumen and mucosal surfaces. Satisfactory was defined as some, but incomplete, visibility. Unsatisfactory was defined as inability to visually examine the lumen and the mucosal surfaces.

Postprocedure monitoring

After completion of the procedure, the infused saline solution was allowed to flow out of the egress port. The monitor was watched during the process, and as the gastrointestinal tract collapsed, the telescope was slowly withdrawn until as much as possible of the infused saline solution flowed out. Once the saline solution had been removed, isoflurane administration was discontinued. Endotracheal tubes were removed once pigeons recovered from anesthesia. Pigeons were then positioned in ventral recumbency, supported by towels, in a heated cage, where they were visually examined continuously until they were standing. Afterward, they were returned to their aviary.

Pigeons were monitored every 2 hours during the day of the procedure, 3 times/d on the day following the procedure, and 2 times/d thereafter for 3 to 6 days. Monitoring included assessment of general demeanor, appetite, and quality of droppings as well as once-daily measurement of body weight. A second blood sample was collected for measurement of PCV and plasma total solids concentration 24 hours after the procedure.

Necropsy

All pigeons were euthanized by administration of an overdose of a solutionn containing pentobarbital sodium and phenytoin sodium via a basilic vein 3 (for 4 pigeons), 4 (for 4 pigeons), 5 (for 4 pigeons), or 6 (for 2 pigeons) days following gastroscopy. A complete necropsy was performed on each pigeon; special attention was paid to the upper gastrointestinal tract, with inspection for evidence of perforation or ulceration. The entire upper gastrointestinal tract, including the crop, proventriculus, and ventriculus, was removed en bloc and placed in a jar containing neutral-buffered 10% formalin. After fixation of tissues was achieved, at least 2 representative samples of proventriculus and ventriculus specimens were prepared for histologic evaluation. Any grossly evident lesions in these organs, including biopsy sites, were also prepared for histologic evaluation. Samples were prepared as 2- to 3-mm, full-thickness cubes to resemble biopsy specimens. Although this size was considerably larger than an endoscopic biopsy specimen would be, it was small enough to duplicate issues with sample orientation that occurred during histologic processing of endoscopic biopsy specimens.

Histologic evaluation

Fixed biopsy and necropsy specimens were processed routinely, embedded in paraffin, sectioned at 5 μm, stained with H&E, and examined microscopically by a board-certified pathologist (AEE). Biopsy specimens were evaluated for crush artifact and the depth obtained. The degree of crush artifact was graded as minimal when only the periphery of the specimen was crushed and that artifact did not interfere with specimen evaluation. A more extensive degree of crush artifact was graded as mild, moderate, or severe on the basis of the resulting inability to evaluate tissue architecture or recognize cellular components. Depth of the specimen obtained was graded as inadequate, marginal, or adequate. Adequate was defined as a full-thickness mucosal sample, extending to and including the submucosa for the ventriculus or extending to the base of a gland for the proventriculus. Marginal was defined as containing both epithelium and lamina propria but not clearly extending to the submucosa for the ventriculus or to the base of a gland for the proventriculus. Inadequate was defined as containing koilin only, koilin and epithelium without lamina propria for the ventriculus, and plicated proventricular epithelium without underlying glands for the proventriculus.

Histologic changes identified in biopsy specimens were also described. In situations in which biopsy sites were identifiable at necropsy, any histologic changes in necropsy specimens obtained from those sites were described. Necropsy specimens were also histologically evaluated for abnormalities related to the biopsy procedure.

Statistical analysis

Values for body weight and hematologic variables are reported as mean ± SD. Values for PCV and plasma total solids concentration were compared between measurement points (baseline and 24 hours after the procedure) by use of the paired t test and statistical software.o Values of P < 0.05 were considered significant. Body weights of pigeons were compared among measurement points by use of repeated-measures ANOVA. The full model included a fixed factor representing measurement point and a random intercept representing individual pigeons. Results of multiple comparisons were corrected for those comparisons by use of the Tukey test. Results of histologic scoring are reported as descriptive statistics.

Results

Animals

Prior to gastroscopy and collection of biopsy specimens, physical examination findings were unremarkable for the 14 included pigeons (excluding the pigeon in which the proventriculus had been perforated at the beginning of the study). Mean ± SD body weight at that point was 456 ± 5 g. Mean PCV was 53 ± 3%, and mean plasma total solids concentration was 3.96 ± 0.38 g/dL.

Procedure

Preparation of the upper gastrointestinal tract was readily achieved by flushing with warm sterile saline solution prior to gastroscopy. On entering the oral cavity, the endoscope was easily passed into the crop, proventriculus, and cranial portion of the ventriculus (Figure 2). Minimal irrigation with saline solution was needed to dilate the lumen and allow visual examination as the endoscope was advanced caudally through the thoracic portion of the esophagus and into the proventriculus. In some pigeons, residual debris impeded visual examination of the mucosa, so the lumen was flushed by alternating between ingress and egress of saline solution to aid flushing. Conservative irrigation was important given that overzealous irrigation resulted in rapid crop distension and backflow of saline solution into the oral cavity. As the operator became more experienced, the incidence of these unwanted effects became less frequent and less saline solution was needed.

Figure 2—
Figure 2—

Representative endoscopic images of the upper gastrointestinal tract of pigeons. A—Cervical portion of the esophagus. B—Crop infused with saline (0.9% NaCl) solution. C—Crop, with opening to the thoracic portion of the esophagus in the center. D—Lumen of the proventriculus. E—Close-up view of the proventricular mucosa. Notice the gastric glands. F—Biopsy forceps positioned parallel against the proventricular mucosa during biopsy specimen collection. G—Hemorrhage that occurred after proventricular biopsy. A clot developed rapidly. H—Lumen of the proventriculus. Notice the mucous covering. This was a normal finding, with no associated histologic lesions. I—Contracted isthmus separating the ventriculus. J—Entry into the ventriculus. Notice the grit and green coloration of the koilin, which were normal findings. K—Biopsy forceps in the process of obtaining a sample perpendicular to the ventriculus. L—Ventriculus following collection of 2 biopsy specimens. Notice the koilin defect and minimal mucosal hemorrhage.

Citation: American Journal of Veterinary Research 78, 1; 10.2460/ajvr.78.1.42

Visibility of the proventriculus was excellent in all pigeons, with clear detail of the mucosal surface and the gastric glands appreciated (Figure 2). Mucosal biopsy specimens were easily and safely collected by distending the proventriculus and collecting the specimen with the biopsy forceps nearly parallel to the long axis of the proventriculus. After collection, mild hemorrhage occurred; however, a clot rapidly formed. No perforations occurred with this technique.

Visibility of the cranial sac of the ventriculus was excellent, with clear detail of the koilin layer (Figure 2). Biopsy of the koilin layer was easily performed with jaws of the biopsy forceps pressed perpendicular against the wall. The mucosal layer beneath the koilin was difficult to grasp with the forceps, regardless of whether the lumen was distended or deflated. Minimal hemorrhage was noticed with the ventricular biopsy procedure. Because of the length of the endoscope, it was not possible to advance caudally to visually examine the middle and caudal portions of the ventriculus. Therefore, biopsy of these regions was not attempted and visibility of these compartments could not be graded.

Conservative irrigation with saline solution was important to reduce the risk of aspiration. During the first half of the study, 1 pigeon went into cardiac arrest and died during the ventricular biopsy portion of the procedure. Only proventricular biopsy specimens were collected and evaluated for this pigeon. Pulmonary edema was detected at necropsy and was attributed to aspiration of saline solution, given that the lungs sank in the formalin solution used for tissue fixation. The remaining 13 pigeons recovered rapidly and uneventfully from the procedure. The anesthetic protocol was effective, and additional analgesic or antimicrobial treatment was not provided or deemed necessary.

Effects of the procedure on pigeons

Demeanor and appetite of the 13 surviving pigeons appeared to be quickly restored the day of the procedure. The excrement of some pigeons appeared dark in color (suggestive of melena) for 1 to 2 days after the procedure, but this resolved thereafter. One pigeon was suspected to have regurgitated, with dried food observed on its neck the morning after the procedure. Results of physical examination of that pigeon were otherwise unremarkable, and regurgitation resolved the following day.

No significant (P = 0.66) difference was identified in mean body weights of the 13 surviving pigeons over the study period. Compared with baseline values, no significant decrease was identified in postoperative PCV or plasma total solids concentration. The PCV in 1 pigeon decreased from 55% at baseline to 39% 24 hours later and then to 30% 48 hours after the procedure. Physical examination findings for this pigeon were unremarkable, and passing of dark-colored excrement ceased by 24 hours after the procedure. Necropsy findings for the bird were also unremarkable; however, histologic examination of the distal portion of the femur revealed hematopoietic neoplasia, most likely of myeloid origin, which was considered the cause of the low PCV.

At necropsy, no evidence was detected of trauma, perforation, or other changes associated with the gastroscopy or biopsy sites in the 13 pigeons that had survived the procedure. Biopsy sites were definitively identified grossly in only 2 pigeons. Of these, one had no histologic lesions associated with the site, whereas the other had only mild multifocal lymphoplasmacytic and heterophilic inflammation in the adjacent adipose tissue.

Histologic findings for biopsy and necropsy specimens

The degree of crush artifact was minimal to absent in all examined proventriculus and ventriculus biopsy specimens. For 1 pigeon, the 2 proventriculus specimens were lost during processing. Diagnostic quality of proventricular specimens was graded as adequate for 10 of 13 pigeons, marginal for 2 pigeons, and inadequate for 1 pigeon (Figure 3). Diagnostic quality of ventricular specimens was graded as inadequate in most (8/13) pigeons, consisting primarily of koilin with fragments of interdigitating surface epithelium. For 3 pigeons, ventriculus specimens were of adequate quality, and 2 were of marginal quality.

Figure 3—
Figure 3—

Representative photomicrographs of biopsy specimens collected during gastroscopy in pigeons. A—Mucosal specimen obtained from the proventriculus. This specimen is considered of adequate thickness and includes both plicated surface epithelium and underlying glands. B—Mucosal specimen obtained from the proventriculus. This sample is considered of inadequate thickness and includes only plicated surface epithelium. C—Mucosal specimen obtained from the ventriculus. This sample is considered of full thickness and includes koilin, mucosa, and tunica muscularis. D—Mucosal specimen obtained from the ventriculus. This specimen is considered inadequate and includes only koilin. H&E stain; bar = 200 μm.

Citation: American Journal of Veterinary Research 78, 1; 10.2460/ajvr.78.1.42

Although attempts had been made to obtain full-thickness sections at necropsy, histologic evaluation revealed that these specimens were often not full thickness. The small size (2 to 3 mm) of the necropsy samples obtained from the proventriculus and ventriculus for comparison with similarly sized biopsy specimens made orientation during embedding more difficult than if larger samples had been collected. Necropsy samples (38 each from the proventriculus and ventriculus, representing 2 or 3/pigeon), which had been consistently obtained in full thickness, were confirmed histologically to be full thickness for 26 proventriculus samples and 36 ventriculus samples. However, orientation problems were encountered in fixed tissue sections from 8 proventriculus samples and 2 ventriculus samples. In these improperly oriented sections, all mucosal layers were present but there was artifactual expansion or thinning of layers, absence of a distinct mucosal surface, or odd orientation in which muscularis was surrounded on multiple sides by mucosa. Tissue sections from 12 proventriculus samples and 1 ventriculus sample contained only mucosa, whereas the section from 1 ventriculus sample contained only muscularis.

Histologic abnormalities in biopsy specimens were uncommon. Two pigeons had biopsy samples containing lesions that consisted of mild hemorrhage in the koilin and markedly dilated proventricular glands surrounded by an intense lymphocytic infiltrate with few heterophils. Six pigeons, including the 1 for which the biopsy site was identified, had histologic lesions evident in necropsy samples. These lesions, which were possibly related to the biopsy procedure, included mild to marked fibrosis with granulomas and intramural perivascular inflammation in 1 pigeon; mild lymphohistiocytic inflammation and fibroplasia in the deep mucosa, submucosa, and muscularis of the ventriculus with few intraluminal heterophils in a second pigeon; and mild lymphohistiocytic interglandular inflammation in the isthmus as well as focally extensive glandular hyperplasia, disorganization, herniation, and lymphohistiocytic inflammation surrounding foreign material and extending into the muscularis of the ventriculus in a third pigeon. These lesions were small and unlikely to have been clinically important given that the associated pigeons had had no clinical signs suggestive of disease.

Lesions believed to be unrelated to the biopsy procedure were identified in 2 pigeons. In necropsy samples from 1 pigeon, the lesions included glandular ectasia with periglandular fibrosis, mild lymphocytic inflammation, and intraluminal larvated ova. In necropsy samples from the second pigeon, lesions suggestive of focal atherosclerosis were identified. Lesions suggestive of focal lymphohistiocytic air sacculitis were also detected in 1 pigeon and may or may not have been related to experimental manipulation; however, these lesions were mild and considered clinically unimportant.

Discussion

Gastroscopy of the proventriculus and ventriculus of healthy pigeons via an oral approach in the study reported here provided excellent visibility of the lumen and mucosal surface of the proventriculus and cranial portion of the ventriculus. With appropriate positioning of the pigeons, close monitoring of anesthesia, and conservative irrigation with saline solution, the gastroscopy procedure was safe.

In addition to controlling the ingress and egress ports, irrigation with saline solution may be more easily controlled by use of a bag that is not suspended high above the table, therefore reducing the pressure on the solution; the flow control on the IV infusion line to set the maximum infusion rate; or a predetermined volume for the size and species of bird. Air insufflation by use of flexible gastroscopes is traditionally used in veterinary gastroscopy. However, achieving distension of the lumen of avian gastric compartments through insufflation with air is challenging because of the reverse Trendelenburg position applied to a bird during the procedure, the presence of its crop, and its short esophagus, which allow the air to escape.

The ability to distend the lumen by use of saline solution instead is important to allow safe collection of biopsy specimens. In addition, irrigation with saline solution allows flushing and cleaning of the avian upper gastrointestinal tract, which often contains ingesta. This technique provides superior visibility to the air-insufflation technique and allows for more accurate collection of biopsy specimens from small birds. Air insufflation may be appropriate in some situations in which collection of biopsy specimens from an avian patient is not planned or indicated, such as foreign body identification and retrieval, and in larger avian species in which flexible gastroscopes can be used.

Positioning of pigeons in dorsal recumbency offers several advantages when irrigation with saline solution is performed. It allows the crop to expand with excess fluid administration, and monitoring of the crop size allows detection of overinfusion and removal of excess fluid before it pools into the oral cavity. The anatomy of the crop needs to be considered when performing this procedure. In avian species lacking a crop or with a rudimentary crop, the importance of diligence during infusion with saline solution cannot be overemphasized.

Careful positioning was important to reduce the risk of complications associated with the procedure in the present study. Placing the endotracheal tube such that the distal opening was facing the dorsal aspect of the trachea appeared to reduce the risk of tracheal obstruction. Tracheal obstruction was appreciated as a decrease in the end-tidal concentration of carbon dioxide with manipulation of the head and neck during gastroscopy. In some situations, turning the pigeon's head slightly laterally resulted in elevation of the glottis, reducing the risk of aspiration associated with overflow of saline solution.

In most avian species weighing > 400 g, particularly species with longer necks than others, a temporary ingluviotomy is recommended when a rigid endoscope is used, as this should allow a more thorough examination of the caudal compartments of the upper gastrointestinal tract.9 This approach would likely provide a more complete evaluation of the ventriculus in pigeons and may also reduce the risk of saline solution aspiration, given that it is a more invasive surgical procedure than gastroscopy via an oral approach.

Endoscopic biopsy of the proventriculus can be safely performed by use of a technique that differs from that recommended for small mammals. For cats and dogs, it is recommended that the jaws of the biopsy forceps be directed perpendicular to the mucosal surface of the stomach. It is also preferable to deflate the stomach lumen to obtain better quality samples.1 Although endoscopic biopsy could be performed safely in the ventriculus because of its thick muscular layer, the initial attempt to obtain a biopsy specimen from the proventriculus of a pigeon in the study reported here resulted in perforation, most likely attributable to the thinner muscle layer of the proventricular wall, compared with that of the mammalian stomach. We strongly recommend collection of proventriculus biopsy specimens with the jaws of the biopsy forceps pressed nearly parallel to the mucosa and the lumen distended to reduce the risk of grasping the full thickness of the proventriculus. This approach was not associated with perforation in any of the pigeons in which biopsy was subsequently performed. Smaller (3F) biopsy forceps may need to be used in smaller birds to reduce the risk of proventricular perforation. However, the size of the biopsy sample obtained with 3F biopsy forceps is approximately a third the size of that obtained with 5F forceps, and a different telescope system is required. The risk of proventricular perforation may be reduced in birds with a luminal mass lesion.

Biopsy specimens collected from the proventriculus of pigeons in the present study had a minimal degree of crush artifact and were of diagnostic quality. Needles or cotton-tipped applicators were not used for specimen transfer because of a previous report11 of damage to biopsy specimens when cotton-tipped applicators, rather than sterile saline solution, were used. Although biopsy specimens were submitted for histologic evaluation in tissue cassettes in the present study, 2 specimens for 1 pigeon were lost during transfer or histologic processing. Particularly for proventriculus specimens, the thin wall and pale color can make tissues difficult to see and automatic tissue processors may have rigorous vacuum cycles that can cause stress to fragile tissues. Therefore, placing such specimens between sponges or using a gel designed for this purpose is recommended at the time of collection to minimize the loss of valuable specimens.

When tissues are embedded in paraffin, they must be carefully oriented so that the correct face of the tissue will be sectioned. With any small tissue specimens, difficulties can arise in identifying the correct surface or in achieving a perfectly oriented section. If the tissue is tangentially sectioned, this can result in artifacts that include narrowing, expansion, or complete absence of 1 or more layers. Similarly, if the incorrect face of the tissue is sectioned, this can result in only 1 layer being included in the section, regardless of whether a full-thickness specimen was originally obtained. These problems are not unique to the present study but are a consistent problem with endoscopic collection of biopsy specimens and general collection of particularly small biopsy specimens.

The usefulness of obtaining biopsy specimens of ventricular mucosa is uncertain and would need to be further explored in birds with known ventricular diseases. Full-thickness mucosal specimens were uncommonly obtained from the ventriculus of pigeons in the present study, and most ventriculus specimens contained koilin only or koilin and superficial epithelium. Few diseases are manifested within the koilin, and mucosal diseases might or might not extend to the superficial mucosa. Additional evaluation is required to determine whether presence of ventricular disease or a grossly visible lesion would allow more successful specimen collection or whether an alternative biopsy technique is required for this compartment of the avian gastrointestinal tract.

Hemorrhage attributable to the biopsy procedure in the present study was minor and had no effect on PCV and plasma total solids concentration measured 24 hours later; however, evidence of short-term melena was observed. Whenever evidence of hemorrhage or an ongoing decrease in PCV is identified following collection of biopsy specimens from birds, further investigation is warranted. The proventriculus appeared to yield specimens of higher diagnostic quality than did the ventriculus. In clinical practice, multiple biopsies should be performed to maximize the opportunity for obtaining adequate, representative specimens; however, whether this can be achieved is dependent on the size of the bird. In the present study, 2 biopsy specimens/pigeon were considered diagnostic in most situations for evaluation of the proventricular mucosa. Additional research is needed to evaluate the reported gastroscopy and biopsy technique in birds with suspected proventricular or ventricular disorders and in avian species other than pigeons.

Acknowledgments

Supported by the Pamela de Journo Fund.

Presented in abstract form at the ExoticsCon, San Antonio, Tex, September 2015.

The authors thank Lisa Reno and Ethan Karstedt for technical assistance and Dr. Deborah Keys for statistical assistance.

Footnotes

a.

Nutriblend Gold, Purina Mills LLC, St Louis, Mo.

b.

Torbugesic (10 mg/mL), Fort Dodge Animal Health, Overland Park, Kan.

c.

Isoflo, Abbott Laboratories, Abbott Park, Ill.

d.

VT-5000, BASi Vetronics, Bioanalytical Systems Inc, West Lafayette, Ind.

e.

Doppler ultrasonic flow detector model 811-B, Parks Medical Electronics Inc, Aloha, Ore.

f.

DRE Echo vital signs monitor, DRE Medical Inc, Louisville, Ky.

g.

Temperature therapy pad and T/PUMP classic model TP650, Gaymar Industries, Orchard Park, NY.

h.

HOPKINS telescope 64019BA, Karl Storz, Tuttlingen, Germany.

i.

Examination sheath 67065CV, Karl Storz, Tuttlingen, Germany.

j.

Cold light fountain XENON 20132101-1, Karl Storz, Tuttlingen, Germany.

k.

HD camera head 22220055-3, Karl Storz, Tuttlingen, Germany.

l.

IMAGE 1 HUB HD camera control unit 22201011U110, Karl Storz, Tuttlingen, Germany.

m.

Biopsy forceps 67161Z, Karl Storz, Tuttlingen, Germany.

n.

Beuthanasia-D, Schering Plough Animal Health, Summit, NJ.

o.

SAS, version 9.3, SAS Institute, Cary, NC.

References

  • 1. Sum S, Ward CR. Flexible endoscopy in small animals. Vet Clin North Am Small Anim Pract 2009;39: 881902.

  • 2. Langlois I. The anatomy, physiology, and diseases of the avian proventriculus and ventriculus. Vet Clin North Am Exot Anim Pract 2003;6: 85111.

    • Search Google Scholar
    • Export Citation
  • 3. Anjos BL, Diefenbach A, Rissi DR, et al. Anaplastic ventricular adenocarcinoma in a blue and gold macaw (Ara ararauna). Braz J Vet Pathol 2012;5: 7880.

    • Search Google Scholar
    • Export Citation
  • 4. De Voe R, Degernes L, Karli K. Dysplastic koilin causing proventricular obstruction in an eclectus parrot (Eclectus roratus). J Avian Med Surg 2003;17: 2732.

    • Search Google Scholar
    • Export Citation
  • 5. Gibbons PM, Busch MD, Tell LA, et al. Internal papillomatosis with intrahepatic cholangiocarcinoma and gastrointestinal adenocarcinoma in a peach-fronted conure (Aratinga aurea). Avian Dis 2002;46: 10621069.

    • Search Google Scholar
    • Export Citation
  • 6. Leach MW, Paul-Murphy J, Lowenstine LJ. Pet bird medicine. Three cases of gastric neoplasia in psittacines. Avian Dis 1989;33: 204210.

    • Search Google Scholar
    • Export Citation
  • 7. Yonemaru K, Sakai H, Asaoka Y, et al. Proventricular adenocarcinoma in a Humboldt penguin (Spheniscus humboldti) and a great horned owl (Bubo virginianus); identification of origin by mucin histochemistry. Avian Pathol 2004;33: 7781.

    • Search Google Scholar
    • Export Citation
  • 8. Campbell TW, Turner O. Carcinoma of the ventriculus with metastasis to the lungs in a sulphur-crested cockatoo (Cacatua galerita). J Avian Med Surg 1999;13: 265268.

    • Search Google Scholar
    • Export Citation
  • 9. Divers SJ. Avian diagnostic endoscopy. Vet Clin North Am Exot Anim Pract 2010;13: 187202.

  • 10. Mejia-Fava J, Divers SJ, Jiménez D, et al. Diagnosis and treatment of proventricular nematodiasis in an umbrella cockatoo (Cacatua alba). J Am Vet Med Assoc 2013;242: 11221126.

    • Search Google Scholar
    • Export Citation
  • 11. Stahl SJ, Hernandez-Divers SJ, Cooper TL. Evaluation of trans-cutaneous pulmonoscopy for examination and biopsy of the lungs of ball pythons and determination of preferred biopsy specimen handling and fixation procedures. J Am Vet Med Assoc 2008;233: 440445.

    • Search Google Scholar
    • Export Citation

Contributor Notes

Address correspondence to Dr. Sladakovic (izidora.s@gmail.com).

Dr. Ellis’ present address is Antech Diagnostics, 1111 Marcus Ave, Lake Success, NY 11042.