T-cell receptor and immunoglobulin gene polymorphisms and resistance to Haemonchus contortus in sheep

SUMMARY: DNA extracted from animals originating from three large half-sib sheep families derived from a single, highly resistant ram were probed with ovine T-cell receptor α and β chain and immunoglobulin lambda light chain variable and constant region cDNA probes. When genomic DNA was digested with TaqI, 19 polymorphic bands were detected with the lambda light chain immunoglobulin variable region probe and 6,4 and 5 polymorphic bands were detected with the T-cell receptor α and β probes and the lambda immunoglobulin constant region probes, respectively. All animals had been phenotypically assessed for Haemonchus contortus resistance by faecal egg counts and using best linear unbiased statistical methods, no statistically significant associations were found between RFLP banding patterns detected at these loci and faecal egg counts. The results show that germline polymorphism at these loci does not account for a significant proportion of the variation in resistance to Haemonchus contortus in this flock. ZUSAMMENFASSUNG: T-Zellrezeptor und Immunoglobulin-Genpolymorphismus und Resistenz von Schafen gegen Haemonchus contortus DNS aus Tieren von drei gro?en Halbgeschwister-Schaffamilien aus einem einzelnen hochgradig resistenten Schafbock wurden mit cDNS-Sonden aus ovinen T-Zellrezeptor α- und β-Ketten und Immunoglobulin λ leichten variablen Ketten und konstanten Regionen untersucht. Nach Verdau der genomischen DNS mit TaqI 19 wurden polymorphe Banden mit der λ leichte Ketten Immunoglobulin variablen Region-Sonde entdeckt, 6, 4 und 5 polymorphe Bande wurden mit T-Zellrezeptor α- und β-Sonden und mit der Sonde der λ Immunoglobulin konstanten Region gefunden. Alle Tiere wurden ph?notypisch in Hinblick auf Haemonchus contortus-Resistenz mittels f?kaler Eiproben untersucht. Statistische Analyse mittels linearer unverzerrter Pr?diktion ergab keine statistisch signifikanten Zusammenh?nge zwischen RFLP Bandmustern und f?kalen Eizahlen. Genetischer Polymorphismus an diesen Loci scheint nicht für einen signifikanten Teil der Variabilit?t in der Resistenz gegen Haemonchus contortus verantwortlich zu sein.     

Mastocytemia in dogs with acute inflammatory diseases

Nineteen dogs were identified that had mastocytemia (mast cells in venous blood samples) not associated with mast cell neoplasia. The first 10 dogs were identified by examination of blood films from dogs with suspected parvovirus enteritis (8), fibrinous pericarditis and pleuritis secondary to thoracic lacerations (1), and renal insufficiency of unknown cause (1). Because of the apparent association with acute enteritis, blood films from 52 suspected canine parvovirus cases were examined retrospectively and 8 mastocytemic dogs were found. An additional 52 canine blood films were randomly selected from the same retrospective time period and 1 mastocytemic dog was found that had pneumothorax, pelvic fractures, and hemorrhagic septic abdominal effusion secondary to renal hemorrhage and traumatized intestines. All mastocytemic dogs had acute inflammatory leukograms the day that mast cells were first detected: neutropenia with toxic neutrophils (4), neutropenia with a left shift (8), total neutrophil count within reference interval but with a left shift (5), or neutrophilia with a left shift (2). All dogs except the renal insufficiency case had circulating toxic neutrophils. Five dogs were mastocytemic on more than 1 day. The pathogenesis of the mastocytemia associated with the acute inflammatory disease was not determined.     

Nuclear receptor and target gene mRNA abundance in duodenum and colon of dogs with chronic enteropathies

Nuclear receptors (NR), such as constitutive androstane receptor (CAR), pregnane X receptor (PXR) and peroxisome proliferator-associated receptors alpha and gamma (PPAR, PPARγ) are mediators of inflammation and may be involved in inflammatory bowel disease (IBD) and food responsive diarrhea (FRD) of dogs. The present study compared mRNA abundance of NR and NR target genes [multi drug-resistance gene-1 (MDR1), multiple drug-resistance-associated proteins (MRD2, MRD3), cytochrome P450 (CYP3A12), phenol-sulfating phenol sulfotransferase (SULT1A1) and glutathione-S-transferase (GST A3-3)] in biopsies obtained from duodenum and colon of dogs with IBD and FRD and healthy control dogs (CON; n = 7 per group). Upon first presentation of dogs, mRNA levels of PPAR, PPARγ, CAR, PXR and RXR in duodenum as well as PPARγ, CAR, PXR and RXR in colon were not different among groups (P > 0.10). Although mRNA abundance of PPAR in colon of dogs with FRD was similar in both IBD and CON (P > 0.10), PPAR mRNA abundance was higher in IBD than CON (P < 0.05). Levels of mRNA of MDR1 in duodenum were higher in FRD than IBD (P < 0.05) or CON (P < 0.001). Compared with CON, abundances of mRNA for MRP2, CYP3A12 and SULT1A1 were higher in both FRD and IBD than CON (P < 0.05). Differences in mRNA levels of PPAR and MRP2 in colon and MDR1, MRP2, CYP3A12 and SULT1A1 in duodenum may be indicative for enteropathy in FRD and (or) IBD dogs relative to healthy dogs. More importantly, increased expression of MDR1 in FRD relative to IBD in duodenum may be a useful diagnostic marker to distinguish dogs with FRD from dogs with IBD.     

Variations on brain microglial gene expression of MMPs, RECK, and TIMPs in inflammatory and non-inflammatory diseases in dogs

Matrix metalloproteinases (MMPs), MMP inhibitors (TIMPs, tissue inhibitors of matrix metalloproteinases), and the membrane-anchored glycoprotein RECK (reversion-inducing cysteine-rich protein with Kazal motifs) contribute to the pathogenesis of many CNS diseases. To assess the potential pathogenetic roles of microglial MMP, TIMP, and RECK generation in extracellular matrix breakdown, opening of the blood brain barrier (BBB) and subsequent recruitment of leukocytes in the CNS, twenty-four dogs suffering from spontaneously occurring different intracranial and extracranial (control group) diseases were examined. Microglia cells were isolated ex vivo by density gradient centrifugation and their expressions of MMP-2, MMP-9, MMP-12, MMP-13, MMP-14, TIMP-1, TIMP-2, and RECK were examined via quantitative real-time polymerase chain reaction (qPCR). Zymography on CNS tissues in selected cases was performed to assess differences at the protein level. Dogs were grouped in different disease categories according to histopathological examinations, in groups with or without inflammatory reactions, and in groups with/without contrast enhancement in advanced diagnostic imaging as a function of BBB breakdown. The results showed a significant up-regulation of MMP-9 in dogs with inflammation in the nervous system compared to dogs with non-inflammatory diseases. An increased expression of MMP-9 might lead to a facilitated invasion of white blood cells. Furthermore, down-regulation of MMP-13 was found in dogs with contrast enhancement. Zymographical data reflected MMP-2 qPCR data. In conclusion, differential expression of MMPs and their inhibitors, but not of RECK, which might crucially influence the pathogenesis of a given disease, could be demonstrated in canine microglia. This reflects a further pathway in the microglial repertoire to respond to various disease conditions in the CNS, a characteristic that might be of particular relevance as a target for specific treatments.     

An update on aspects of viral gastrointestinal diseases of dogs and cats

Viruses commonly cause gastrointestinal illnesses in dogs and cats that range in severity from mild diarrhoea to malignant neoplasia. Perpetual evolution of viruses is reflected in changing disease patterns, so that familiar viruses are sometimes discovered to cause new or unexpected diseases. For example, canine parvovirus (CPV) has regained the ability to infect felids and cause a panleucopenia-like illness. Feline panleucopenia virus (FPV) has been shown to cause fading in young kittens and has recently been implicated as a possible cause of feline idiopathic cardiomyopathy. Molecular scrutiny of viral diseases sometimes permits deeper understanding of pathogenesis and epizootiology. Feline gastrointestinal lymphomas have not, in the past, been strongly associated with retroviral infections, yet some of these tumours harbour retroviral proviruses. Feline leukaemia virus (FeLV) may play a role in lymphomagenesis, even in cats diagnosed as uninfected using conventional criteria. There is strong evidence that feline immunodeficiency virus (FIV) can also be oncogenic. The variant feline coronaviruses that cause invariably-fatal feline infectious peritonitis (FIP) arise by sporadic mutation of an ubiquitous and only mildly pathogenic feline enteric coronavirus (FECV); a finding that has substantial management implications for cat breeders and veterinarians. Conversely, canine enteric coronavirus (CECV) shows considerable genetic and antigenic diversity but causes only mild, self-limiting diarrhoea in puppies. Routine vaccination against this virus is not recommended. Although parvoviruses, coronaviruses and retroviruses are the most important known viral causes of canine and feline gastrointestinal disease, other viruses play a role. Feline and canine rotaviruses have combined with human rotaviruses to produce new, reassortant, zoonotic viruses. Some companion animal rotaviruses can infect humans directly. Undoubtedly, further viral causes of canine and feline gastrointestinal disease await discovery.     

Evaluation of gastrointestinal permeability and mucosal absorptive capacity in dogs with chronic enteropathy

OBJECTIVE: To assess intestinal mucosal function by measuring permeability and absorptive capacity in dogs with chronic enteropathy (CE) before and after treatment and to determine whether those variables were correlated with clinical disease activity or histologic scoring of intestinal biopsy specimens. ANIMALS: 29 dogs with CE. PROCEDURE: Dogs were designated as having dietresponsive CE or CE requiring glucorticoid treatment. Severity of clinical signs was assessed by calculating the canine inflammatory bowel disease activity index (CIBDAI). Histologic severity of intestinal infiltration was assessed before and after 4 weeks of treatment in the diet-responsive group and before and after 10 weeks of treatment in the glucocorticoid group. Gastrointestinal permeability and mucosal absorptive capacity were assessed by use of intragastric administration of a solution containing lactulose, rhamnose, xylose, 3-O-methylglucose, and sucrose. Urine was collected 6 hours after administration of the sugar solution to determine urinary lactulose-to-rhamnose (L:R), xylose-to-methylglucose (X:M), and sucrose-to-methylglucose (S:M) ratios. RESULTS: Median CIBDAI scores decreased significantly in both groups of dogs after treatment. However, the median histologic grade of intestinal biopsy specimens did not change with treatment in either group. There were no significant differences in L:R, X:M, or S:M ratios after treatment in either group and no significant correlations between L:R, X:M, or S:M ratios and CIBDAI or histologic scores. CONCLUSIONS AND CLINICAL RELEVANCE: Results of tests for intestinal permeability and mucosal absorptive capacity were not useful indicators of clinical disease activity as assessed by the CIBDAI or the sever ity of infiltration as indicated by histologic evaluation.     

Insulin-like growth factor I concentration in dogs with inflammatory and neoplastic liver diseases

Liver diseases are known to influence the serum concentration of insulin-like growth factor I (IGF-I) in humans, but such an effect has rarely been investigated in dogs. The aim of this study was to investigate serum IGF-I concentrations in dogs with primary liver diseases, in comparison with levels in healthy dogs and dogs with non-hepatic diseases. For this purpose, IGF-I serum concentrations were measured (DSL-5600 kit) in 36 dogs with various liver diseases and compared with 22 healthy controls and 20 dogs with non-hepatic diseases. The results showed that dogs with liver diseases had significantly lower IGF-I serum concentrations (P < 0.001) than clinical healthy dogs or dogs with non-hepatic diseases. But the results also indicate that the aetiology of liver disease has no influence on IGF-I serum concentration.     

Laboratory procedures for the diagnosis of gastrointestinal tract diseases of dogs and cats

An increasing number of laboratory tests are available for diagnosis of gastrointestinal tract diseases in dogs and cats. Use of these tests can lead to more accurate and rapid diagnoses. This review discusses laboratory tests, both new and old, and the role they currently play in the evaluation of animals presented with gastrointestinal problems. A minimum database helps assess the severity of the disorder, detect extra-gastrointestinal causes of problems and assists in formulating diagnostic and therapeutic plans.

Faecal examination remains one of the most important diagnostic procedures in the investigation of gastrointestinal problems. Zinc sulphate faecal flotation is an excellent routine screening technique for helminth and protozoal infections, including giardiasis. Rectal cytology can assist in the diagnosis of large bowel disorders. Interpretation of faecal immunodiagnostic tests is hampered by insufficient knowledge of test sensitivities and specificities. Routine faecal cultures are not warranted and faecal occult blood tests are rarely indicated.

Serum tests for gastric inflammation are now under development. The serum trypsin-like immunoreactivity test remains the gold standard technique for the diagnosis of exocrine pancreatic insufficiency. Breath hydrogen tests can be helpful in assessing the functional relevance of mild abnormalities in small-bowel biopsy specimens. Subnormal concentrations of serum cobalamin appear to be more specific indicators of gastrointestinal disease in cats than in dogs. Tests for small intestinal bacterial overgrowth remain controversial and assessment of gastrointestinal permeability has yet to prove its value in the diagnostic assessment of companion animals with gastrointestinal problems. Faecal alpha₁-protease inhibitor (α₁-PI) shows promise for the diagnosis of protein-losing enteropathy.     

Laboratory procedures for the diagnosis of gastrointestinal tract diseases of dogs and cats

An increasing number of laboratory tests are available for diagnosis of gastrointestinal tract diseases in dogs and cats. Use of these tests can lead to more accurate and rapid diagnoses. This review discusses laboratory tests, both new and old, and the role they currently play in the evaluation of animals presented with gastrointestinal problems. A minimum database helps assess the severity of the disorder, detect extra-gastrointestinal causes of problems and assists in formulating diagnostic and therapeutic plans. Faecal examination remains one of the most important diagnostic procedures in the investigation of gastrointestinal problems. Zinc sulphate faecal flotation is an excellent routine screening technique for helminth and protozoal infections, including giardiasis. Rectal cytology can assist in the diagnosis of large bowel disorders. Interpretation of faecal immunodiagnostic tests is hampered by insufficient knowledge of test sensitivities and specificities. Routine faecal cultures are not warranted and faecal occult blood tests are rarely indicated. Serum tests for gastric inflammation are now under development. The serum trypsin-like immunoreactivity test remains the gold standard technique for the diagnosis of exocrine pancreatic insufficiency. Breath hydrogen tests can be helpful in assessing the functional relevance of mild abnormalities in small-bowel biopsy specimens. Subnormal concentrations of serum cobalamin appear to be more specific indicators of gastrointestinal disease in cats than in dogs. Tests for small intestinal bacterial overgrowth remain controversial and assessment of gastrointestinal permeability has yet to prove its value in the diagnostic assessment of companion animals with gastrointestinal problems. Faecal alpha1-protease inhibitor shows promise for the diagnosis of protein-losing enteropathy.     

Evaluation of resting cortisol concentration testing in dogs with chronic gastrointestinal signs

BackgroundResting cortisol concentrations are routinely measured in dogs with chronic gastrointestinal signs to rule out hypoadrenocorticism based on a concentration >2 μg/dL (>55 nmol/L).Hypothesis/ObjectivesTo assess the cross‐sectional prevalence of hypoadrenocorticism in a group of dogs with chronic gastrointestinal signs presented to a referral internal medicine service.AnimalsTwo‐hundred and eighty‐two client‐owned dogs with chronic gastrointestinal signs and with resting cortisol concentration testing performed.MethodsRetrospective review of medical records (final diagnosis, resting cortisol concentration, and adenocorticotropic hormone [ACTH] stimulation test results) of a referral population of dogs between May 2013 and September 2017.ResultsResting cortisol concentration was <2 μg/dL (<55 nmol/L) in 79 patients (28%). Repeated resting cortisol concentration measurements were performed in 28 dogs, and in 8, resting cortisol concentrations remained <2 μg/dL (<55 nmol/L). Post‐ACTH cortisol concentration was <2 μg/dL (<55 nmol/L) in 1 dog, consistent with a diagnosis of hypoadrenocorticism and giving a prevalence estimate of hypoadrenocorticism in this population of dogs of 0.3% (95% confidence interval [95CI], 0.03‐1.5%). In 19 dogs with an initial resting cortisol concentration <2 μg/dL (<55 nmol/L), hypoadrenocorticism was excluded based on a repeat resting cortisol concentration >2 μg/dL (>55 nmol/L). Overall, the most common diagnosis was chronic primary inflammatory enteropathy (176/282, 62.4%), followed by extragastrointestinal neoplasia (17/282, 6%), protein‐losing enteropathy, pancreatitis and megaesophagus (10/282, 3.5% each).Conclusions and Clinical ImportanceAlthough dogs with hypoadrenocorticism can present with chronic gastrointestinal signs, it was the final diagnosis in only 1 of 282 dogs presenting to a referral internal medicine service for signs of chronic enteropathy. Repeated resting cortisol concentration may be considered as a test to try and exclude hypoadrenocorticism.     

Genetic diversity of growth hormone receptor gene in cattle

Growth hormone receptor (GHR) belongs to a member of the cytokine receptor superfamily. Polymorphism of presence or absence of an approximately 1.2 kbp LINE-1 element is observed in bovine GHR gene. The present study was carried out for estimating the genetic diversity and the origin of the LINE-1 element in 10 European, Southeastern Asian and East Asian cattle breeds or populations. Genotyping of the LINE-1 revealed predominant LINE-1 presence in European breeds (0.917∼0.991), absence in the Bos taurus indicus populations (0.000∼0.017), and intermediate presence in Northeast Asian cattle (0.417∼0.522). From genetic features of LINE families, LINE-1 of GHR could be attributed to the same origin in both European and Asian cattle, and Asian LINE-1 may not be derived from recent introgression. This result suggested that LINE-1 in bovine GHR gene could have arisen in an ancestral population of Bos taurus taurus .     

Fecal alpha1-proteinase inhibitor concentration in dogs with chronic gastrointestinal disease

Background: Fecal α₁‐proteinase inhibitor (α₁‐PI) clearance is a reliable, noninvasive marker for protein‐losing enteropathy in human beings. An assay for use in dogs has been developed and validated. Objective: The aim of this study was to evaluate fecal α₁‐PI concentration in dogs with chronic gastrointestinal disease, compared with healthy dogs, and to assess its correlation with serum albumin concentration. Methods: Fecal samples were collected from 2 groups of dogs. Group 1 consisted of 21 clinically healthy client‐owned dogs without signs of gastrointestinal disease. Group 2 consisted of 16 dogs referred for investigation of suspected gastrointestinal disease. On the basis of gastric and duodenal biopsies, group 2 was further subdivided into dogs with normal histology (n = 9) and those with histologic abnormalities (n = 7: inflammatory bowel disease, n = 3; lymphangiectasia, n = 4). An ELISA was used to measure α₁‐PI concentrations in fecal extracts. Results: Fecal α₁‐PI concentrations, expressed as μg/g of feces, were not significantly different between groups 1 and 2 as a whole. However, fecal α₁‐PI concentrations (median, minimum‐maximum) were significantly higher in dogs with gastrointestinal diseases associated with histologic abnormalities (60.6 μg/g, 7.4–201.7 μg/g) compared with dogs with normal histology (3.8 μg/g, 0.7–74.0 μg/g) and control dogs (9.9 μg/g, 0.0–32.1 μg/g). There was no significant correlation between fecal α₁‐PI and serum albumin concentrations in dogs with gastrointestinal disease. Conclusions: Increased fecal α₁‐PI concentration may signal the need to obtain gastrointestinal biopsies for a final diagnosis. Fecal α₁‐PI concentration may be a useful test for early detection of protein‐losing enteropathy before decreases in serum albumin concentration can be detected.     

Quantification of mucin gene expression in tracheobronchial epithelium of healthy dogs and dogs with chronic bronchitis

OBJECTIVE: To develop a real-time PCR assay for the quantification of mucin gene expression in tracheobronchial brushing specimens from dogs and compare mucin gene expression in specimens from dogs with naturally occurring chronic bronchitis with that in specimens from healthy dogs. ANIMALS: 7 healthy dogs and 5 dogs with chronic bronchitis. PROCEDURES: Primers that were designed to span the predicted intron-exon boundaries of a canine MUC5AC-like gene were used to develop a real-time PCR assay for quantification of expression of that gene. Total mRNA was isolated from tracheobronchial brushing specimens obtained from dogs with and without bronchitis during anesthesia; MUC5AC-like gene expression in those samples was quantified by use of the real-time PCR assay. RESULTS: The PCR assay was sensitive and specific for the target sequence, the predicted amino acid sequence of which had greatest homology with human, porcine, and rat MUC5AC. The assay was able to quantify the target over a wide dynamic range. Dogs with chronic bronchitis had a 3.0-fold increase in the quantity of MUC5AC-like mRNA, compared with healthy dogs. CONCLUSIONS AND CLINICAL RELEVANCE: The ability to measure mucin gene expression from tracheobronchial brushing specimens collected from client-owned dogs during routine bronchoscopy should prove to be a useful tool for the study of bronchitis in dogs and expand the usefulness of airway inflammation in dogs as a model for bronchitis in humans.