Development and analytical validation of a radioimmunoassay for the quantification of alpha1 -proteinase inhibitor in serum and feces from the common marmoset (Callithrix jacchus).

BACKGROUND: The objective of this study was to develop and analytically validate a radioimmunoassay (RIA) for the measurement of alpha1 -proteinase inhibitor (α1 -PI) concentrations in serum and feces from the common marmoset. METHODS: Serum samples (n = 30) and 3-day fecal samples (n = 30) were obtained from healthy marmosets. An RIA was established and validated by determination of sensitivity, working range, dilutional parallelism, spiking recovery, and intra- and interassay variability. A reference interval for mα1 -PI in serum and feces was established. RESULTS: Sensitivity and upper limit of the working range were 0.75 and 100.62 µg/L, respectively. Observed-to-expected (O/E) ratios for serial dilutions ranged from 89.9% to 123.0% (mean ±SD: 106.0 ± 11.5%) for 8 serum samples, and from 90.6% to 132.7% (mean ±SD: 107.6 ± 19.2%) for 4 fecal samples. O/E ratios for spiking recovery ranged from 97.6% to 104.4% (mean ±SD: 101.3 ± 3%) for 4 serum samples, and from 97.5% to 101.4% (mean ±SD: 99.2 ± 1.8%) for 4 fecal samples and 3 different spiking concentrations. Coefficients of variation (CV) for intra-assay variability for 8 serum samples ranged from 1.7% to 10.6% and 2.2% to 5.1% in the 8 fecal samples. The interassay CV for eight serum samples ranged from 1.3% to 9.9%, and from 1.0% to 6.7% in the 8 fecal samples. The reference interval in serum was determined to be 1047-1484 µg/L. The reference interval in serum was determined to be 1047-1484 µg/L. The reference interval for the 3-day mean fecal concentration, and 3-day maximum fecal concentration were determined to be 32.4-124.4 µg/g and 39.1-158.7 µg/g of feces, respectively. CONCLUSION: The developed assay is sensitive, linear, accurate, precise, and reproducible. Further studies are needed to determine the clinical utility of this assay.

Piecemeal endoscopic polypoidectomy for the management of a canine pharyngeal hemangiosarcoma.

A 7-year-old castrated male Maltese dog was presented for increased respiratory sounds, inability to bark, dysphagia, and hyporexia. Radiographs revealed an ovoid, opaque mass in the caudal nasopharynx. An airway examination and computed tomography scan were followed by endoscopic polypoidectomy. The mass was a hemangiosarcoma, and the patient survived > 13 months.

Polypoïdectomie endoscopique fragmentaire pour la gestion d'un hémangiosarcome pharyngé canin. Un chien Bichon maltais mâle castré âgé de 7 ans a été présenté pour des sons respiratoires accrus, l'incapacité de japper, de la dysphagie et de l'hyporexie. Les radiographies ont révélé une masse ovoïde opaque dans le nasopharynx caudal. Un examen des voies respiratoires et une tomodensitométrie ont été suivis d'une polypoïdectomie endoscopique. La masse était un hémangiosarcome et le patient a survécu >13 mois.(Traduit par Isabelle Vallières).

The fecal microbiome of dogs with exocrine pancreatic insufficiency.

Exocrine pancreatic insufficiency (EPI) in dogs is a syndrome of inadequate synthesis and secretion of pancreatic enzymes. Small intestinal bacterial dysbiosis occurs in dogs with EPI, and is reversed with pancreatic enzyme therapy. However, there are no studies evaluating the fecal microbiome of dogs with EPI. The objective of this study was to evaluate the fecal microbiome of dogs with EPI. Three day pooled fecal samples were collected from healthy dogs (n = 18), untreated (n = 7) dogs with EPI, and dogs with EPI treated with enzyme replacement therapy (n = 19). Extracted DNA from fecal samples was used for Illumina sequencing of the bacterial 16S rRNA gene and analyzed using Quantitative Insights Into Microbial Ecology (QIIME) and PICRUSt was used to predict the functional gene content of the microbiome. Linear discriminant analysis effect size (LEfSe) revealed significant differences in bacterial groups and functional genes between the healthy dogs and dogs with EPI. There was a significant difference in fecal microbial communities when healthy dogs were compared to treated and untreated dogs with EPI (unweighted UniFrac distance, ANOSIM P = 0.001, and 0.001 respectively). Alpha diversity was significantly decreased in untreated and treated EPI dogs when compared to the healthy dogs with respect to Chao1, Observed OTU, and Shannon diversity (P = 0.008, 0.003, and 0.002 respectively). The families Bifidobacteriaceae (P = 0.005), Enterococcaceae (P = 0.018), and Lactobacillaceae (P = 0.001) were significantly increased in the untreated and treated dogs with EPI when compared to healthy dogs. In contrast, Lachnospiraceae (P < 0.001), and Ruminococcaceae (P < 0.01) were significantly decreased in dogs with EPI. Dogs with EPI (before treatment) had significant increases in functional genes associated with secretion system, fatty acid metabolism, and phosphotransferase system. In contrast, healthy dogs had a significant increase in genes related to phenylalanine, tyrosine and tryptophan biosynthesis, transcription machinery and sporulation. In conclusion, this study shows that the fecal microbiome of dogs with EPI (both treated and untreated) is different to that of healthy dogs.

Fecal Concentrations of N-methylhistamine in Common Marmosets (Callithrix jacchus).

Chronic lymphocytic enteritis (CLE) is a frequent disease in common marmosets. However, no diagnostic test for early detection of CLE is available. Mast cells have an important role in gastrointestinal disease. The purpose of this study was to measure fecal concentrations of N-methylhistamine (NMH), a breakdown product of histamine metabolism, in common marmosets. A previously established NMH gas chromatography-mass spectrometry assay for canine feces and urine was used, and partial validation was performed. The reference intervals (n = 30) established for fecal NMH concentrations in common marmoset were 118.2 ng/g or less for a single fecal sample, 121.7 ng/g or less for the 3-d mean, and less than or equal to 167.5 ng/g for the 3-d maximum. Considerable day-to-day variation was observed in fecal NMH concentrations; the mean %CV was 42.2% (minimum, 7.1%; maximum, 141.4%). Fecal NMH concentrations were measured in 14 marmosets for which necropsy reports were available; 7 of the 8 marmosets with CLE and the 1 animal with lymphoma and ulcerative enteritis had increased fecal NMH concentrations. Increased fecal NMH concentrations may serve as a potential marker for CLE; however, further studies exploring the role of mast cells in marmosets with CLE are needed.

Serum Cobalamin and Folate Concentrations in Common Marmosets (Callithrix jacchus) with Chronic Lymphocytic Enteritis.

Serum cobalamin and folate concentrations can serve as surrogate markers of gastrointestinal disease in dogs and cats, where they can have diagnostic, therapeutic, and prognostic implications. Chronic disease of the gastrointestinal tract, particularly chronic lymphocytic enteritis (CLE), occurs frequently in captive common marmosets. The aims of this study were to validate a commercially available assay for measuring serum cobalamin and folate concentrations in common marmosets, to establish reference intervals for these analytes in healthy marmosets, and to measure serum concentrations in common marmosets with CLE. The commercial assay was linear, accurate, precise, and reproducible for the measurement of serum cobalamin and folate concentrations in common marmosets. In healthy marmosets, the serum cobalamin concentration ranged from 322 to 2642 pg/mL (n = 35) and serum folate concentration from 54.8 to 786.4 ng/mL (n = 37). Low serum folate concentrations were moderately sensitive (greater than 70%) for CLE, and low serum cobalamin concentrations were moderately (greater than 70%) specific for CLE. Both serum cobalamin and folate concentrations were relatively unchanged in marmosets during 120 to 220 d. Serum cobalamin and folate concentrations were stable for approximately 7 y when samples were stored at -80 °C. Additional studies are warranted to further study the clinical implications of low serum cobalamin and folate concentrations in common marmosets.

Urine-Filled Large Prostatic Cystic Structure in Two Unrelated Male Miniature Dachshunds.

A 1 yr old intact male miniature dachshund presented for posturing to urinate without voiding and nocturia. Physical examination revealed congenital reproductive abnormalities and a fluid-filled structure caudal to the urinary bladder. The dog was diagnosed with a prostatic cyst and underwent an exploratory laparotomy with an attempt to remove the cyst. Twelve weeks later, the dog returned with recurring clinical signs, and the cyst was found to have returned back to its original size. A second intact male miniature dachshund presented at 7 mo of age for stranguria. Physical examination revealed congenital reproductive abnormalities and a fluid-filled structure on rectal palpation, much like the first dog. The dog was diagnosed with a prostatic cyst and underwent an exploratory laparotomy. An attempt was made to close communication between the prostate and cyst. The dog re-presented 3 wk later for recurrence of clinical signs, and the prostatic cyst was found to have increased in size. Both dogs were euthanized because of recurrence of clinical signs. This report describes the presence of prostatic cysts in two young dogs with congenital abnormalities of the genital and reproductive tracts and the similarities seen in human boys diagnosed with prostatic utricles.

Randomized placebo controlled clinical trial of an enteric coated micro-pelleted formulation of a pancreatic enzyme supplement in dogs with exocrine pancreatic insufficiency.

BACKGROUND: Pancreatic enzyme supplements for the treatment of exocrine pancreatic insufficiency (EPI) in dogs can be uncoated or enteric coated. Enteric coated supplements might be advantageous. HYPOTHESIS/OBJECTIVES: Enteric coated enzyme supplements are superior to uncoated supplements in dogs with clinical EPI. ANIMALS: Eleven dogs with naturally occurring EPI that were apparently free from other diseases. METHODS: Randomized, blinded, controlled cross-over clinical trial comparing a novel micro-encapsulated enteric coated enzyme supplement to a commercially available uncoated product in dogs with clinical EPI. Search of serum canine serum trypsin-like immunoreactivity concentration ≤ 2.5 µg/L in the Gastrointestinal Laboratory database was used to identify dogs with EPI. RESULTS: There was no difference -4.46% (95% CI: -7.97%--0.96%; P = .15) in the % acid hydrolysis fecal fat (primary outcome) between the enteric coated formulation (median: 11.8%; range 6.4%-17.0%) and the uncoated pancreatic enzyme replacement product (median: 17.5%; range: 5.2%-24.9%) in the 11 dogs that completed the study. Other variables did not differ between treatments. CONCLUSIONS AND CLINICAL IMPORTANCE: This study, which had low statistical power, did not detect a difference between formulations.

Purification and partial characterization of α1-proteinase inhibitor in the common marmoset (Callithrix jacchus).

Fecal alpha1-proteinase inhibitor (α1-PI) concentration has been to diagnose enteric protein loss in dogs and cats. Chronic lymphocytic enteritis is commonly seen in the marmoset (Callithrix jaccus) and is characterized by hypoalbuminemia. As a prelude to immunoassay development for detecting enteric protein loss, marmoset serum α1-PI was purified using immunoaffinity chromatography and ceramic hydroxyapatite chromatography. Partial characterization was performed by reducing gel electrophoresis and enzyme inhibitory assays. Protein identity was confirmed with peptide mass fingerprinting and N-terminal amino acid sequencing. Molecular mass, relative molecular mass, and isoelectric point for marmoset α1-PI were 54 kDa, 51,677, and 4.8-5.4, respectively. Trypsin, chymotrypsin, and elastase inhibitory activity were observed. N-terminal amino acid sequence for marmoset α1-PI was EDPQGDAAQKMDTSHH. In conclusion, marmoset α1-PI was successfully purified from serum with an overall yield of 12% using a rapid and efficient method. Purified marmoset α1-PI has characteristics similar to those of α1-PI reported for other species.