Investigation of glucagon-like peptide-1 response to six oral carbohydrates in ponies.

Glucagon-like peptide-1 (GLP-1), the principal incretin in horses, may play a role in the pathophysiology of insulin dysregulation (ID). This study aimed to describe its concentration in response to three preserved forages and four dynamic tests for ID in ponies. Twelve adult ponies of mixed ID status were given a meal of hay, soaked hay or haylage, an in-feed oral glucose test (OGT), oral sugar test (OST), an oral test using a proprietary breakfast cereal (WEET) or a combined glucose-insulin tolerance test (CGIT) weekly in a randomised cross-over study. Glucose, insulin and GLP-1 concentrations were measured before and following each intervention. Ponies were designated ID or non-ID and insulin resistant (IR) or non-IR according to OGT and CGIT results, respectively. All interventions apart from the CGIT provoked a GLP-1 response within 30 min. The OGT and WEET interventions, (containing the greatest dose of non-structural carbohydrate, 1.06 and 1 g/kg BW, respectively), resulted in a greater area under the curve (AUC) for GLP-1 compared to all other interventions (P < 0.001). No difference in GLP-1 response was detected according to ID or IR status, despite there being strong positive correlations (rs [95 % CI]) between GLP-1 and insulin concentrations measured at individual time points (0.67 [0.62 - 0.71]; P < 0.001) and as AUC (0.66 [0.49-0.79], P < 0.001). These data do not support of the use of GLP-1 as an adjunctive diagnostic test for ID or IR, as defined by conventional intravenous or oral dynamic tests.

Comparison of one novel and four established diagnostic tests for insulin dysregulation in ponies.

Several tests have been advocated for diagnosis of insulin dysregulation (ID). Tests using simple sugars may not reflect the response to naturally ingested carbohydrates. This study aimed to evaluate agreement between the oral glucose test (OGT), the oral sugar test (OST), a novel oral test using a proprietary cereal (WEET), the IV combined glucose-insulin tolerance test (CGIT) and fasted basal insulin (FI) for diagnosis of ID. Each of the five tests above was performed on a group of six normal and six insulin dysregulated mixed-breed ponies in a randomised crossover study. Area under the curve (AUC) and maximum concentration of insulin from OGT, OST and WEET showed strong to very strong bivariate correlations (r = 0.85-0.94, and r = 0.87-0.92, respectively; P ≤ 0.001) and were significantly different between tests (associated with dose of carbohydrate) and between CGIT-positive and -negative ponies. Dichotomous results showed substantial agreement between OST and both WEET (κ = 0.65; P = 0.02) and OGT (κ = 0.67; P = 0.01) and between CGIT and both OST (κ = 0.63; P = 0.03) and OGT (κ = 0.67; P = 0.01), and no agreement between FI, which had low sensitivity, and all other tests (κ = 0.15 - 0.31; P > 0. 05). Palatability of WEET was variable, resulting in one pony being excluded for analysis of WEET data. Further work on development of an oral test using a more palatable feedstuff and appropriate cut-offs or diagnostic thresholds for tests of ID is warranted.

Comparison of fasted basal insulin with the combined glucose-insulin test in horses and ponies with suspected insulin dysregulation.

Fasting horses for measurement of basal serum insulin concentration (fasting insulin; FI) has been recommended to standardise testing for insulin dysregulation (ID), yet limited data exist comparing it to dynamic tests. This study aimed to compare FI with the combined glucose-insulin test (CGIT) in horses suspect for ID. We hypothesised that FI would have poor sensitivity for detecting ID compared to CGIT using conventional cut-offs. Records were retrieved from CGITs performed in horses fasted for approximately 8h. Serum insulin and glucose concentrations were measured before and for 150min following an IV bolus of glucose followed by insulin. Correlations between FI and CGIT values were assessed. Youden's index analysis was used to determine the optimal cut-off for FI. Logistic regression and Mann-Whitney U tests were used to determine factors affecting the results. CGITs (n=130) from 62 horses were evaluated. Compared to CGIT, sensitivity and specificity of FI for diagnosis of ID were 14.6% and 100% at a cut-off of 20µIU/mL and 63.4% and 87.2% at a cut-off of 5.2µIU/mL, respectively. FI was significantly correlated with insulin at 45min (rs=0.66) and 75min (rs=0.72); area under the curve for insulin (AUCinsulin; rs=0.67); glucose at 45min (rs=0.53); and AUCglucose (rs=0.50). Obesity was significantly associated with increased odds of a positive CGIT and horses with a positive CGIT were significantly older (P<0.05). In conclusion, FI correlated well with CGIT results and had adequate sensitivity and specificity at lower cut-offs, despite poor sensitivity at conventional cut-off values. Further research to derive cut-off values relevant to the fasting period is warranted.

Insulinaemic and glycaemic responses to three forages in ponies.

Reduction of the hyperinsulinaemic response to feeding is central to the management of insulin dysregulation (ID). The aim of this study was to compare insulinaemic and glycaemic responses to soaked hay, dry hay and haylage in ponies. Twelve ponies of mixed breeds were maintained under identical management conditions. A randomised four-way crossover trial was conducted, in which fasted animals were fed a meal of 0.25% body weight as dry matter intake soaked hay, dry hay or haylage, or administered an oral glucose test (OGT). Blood glucose and serum insulin concentrations were measured before and at 2h following OGT, and regularly for 5h following forage meals. Median and interquartile range (IQR) area under the curve (AUC) for insulin (AUCi) was greater for haylage (median 6495; IQR 17352) vs. dry hay (2932; IQR 5937; P=0.019) and soaked hay (1066; IQR 1753; P=0.002), and greater for dry hay vs. soaked hay (P=0.002). The AUC for glucose (AUCg) was lower for soaked hay (1021; IQR 99) vs. dry hay (1075; IQR 105; P=0.002) and haylage (1107; IQR 221; P=0.003). Six ponies were classified as having ID based on the OGT. AUCi was greater in ID vs. non-ID ponies after all forages. In contrast, there was no detectable effect of ID status on AUCg. On an equivalent dry matter basis, soaked hay produced the lowest insulinaemic and glycaemic responses to feeding, while haylage produced the highest responses. The insulinaemic effects of all forages were greater in ponies with ID. These data support the practice of soaking hay with water to reduce postprandial insulinaemic responses in ponies.

Comparison of two blood sampling techniques for the determination of coagulation parameters in the horse: Jugular venipuncture and indwelling intravenous catheter.

BACKGROUND: Evaluation of coagulation status is an important component of critical care. Ongoing monitoring of coagulation status in hospitalised horses has previously been via serial venipuncture due to concerns that sampling directly from the intravenous catheter (IVC) may alter the accuracy of the results. Adverse effects such as patient anxiety and trauma to the sampled vessel could be avoided by the use of an indwelling IVC for repeat blood sampling. OBJECTIVES: To compare coagulation parameters from blood obtained by jugular venipuncture with IVC sampling in critically ill horses. STUDY DESIGN: Prospective observational study. METHODS: A single set of paired blood samples were obtained from horses (n = 55) admitted to an intensive care unit by direct jugular venipuncture and, following removal of a presample, via an indwelling IVC. The following coagulation parameters were measured on venipuncture and IVC samples: whole blood prothrombin time (PT), fresh plasma PT and activated partial thromboplastin time (aPTT) and stored plasma antithrombin activity (AT) and fibrinogen concentration. D-dimer concentration was also measured in some horses (n = 22). Comparison of venipuncture and IVC results was performed using Lin's concordance correlation coefficient. Agreement between paired results was assessed using Bland Altman analysis. RESULTS: Correlation was substantial and agreement was good between sample methods for all parameters except AT and D-dimers. MAIN LIMITATIONS: Each coagulation parameter was tested using only one assay. Sampling was limited to a convenience sample and timing of sample collection was not standardised in relation to when the catheter was flushed with heparinised saline. CONCLUSIONS: With the exception of AT and D-dimers, coagulation parameters measured on blood samples obtained via an IVC have clinically equivalent values to those obtained by jugular venipuncture.

Evaluation of a Chemiluminescent Immunoassay for Measurement of Equine Insulin.

BACKGROUND: Many diagnostic tests for insulin dysregulation use reference intervals established with an insulin radioimmunoassay (RIA) that is no longer available. A chemiluminescent immunoassay (CLIA) is commonly used for the measurement of serum insulin concentration in clinical practice but requires further validation, especially at clinically relevant reference intervals. OBJECTIVES: To evaluate the CLIA for measurement of equine insulin and compare it to the previously validated, but now unavailable RIA. SAMPLES: Equine serum samples (n = 78) from clinical and experimental studies. METHODS: In this experimental study, performance of the CLIA was evaluated using standard variables, including comparison with the RIA. Continuous and binary outcomes were analyzed. RESULTS: The CLIA showed good intra-assay (coefficient of variation [CV], 1.8-2.4%) and interassay (CV, 3-7.1%) precision. Acceptable recovery on dilution (100 ± 10%) was achieved only at dilutions <1:1. Recovery on addition was acceptable. Comparison of the CLIA and RIA showed strong positive correlation (r = 0.91-0.98), with fixed and proportional bias. At 3 diagnostic cutoffs, sensitivity of CLIA compared with RIA ranged from 67 to 100% and specificity from 96 to 100%. CONCLUSIONS AND CLINICAL IMPORTANCE: The CLIA is a highly repeatable assay which is suitable for within- and between-horse comparisons. Dilution of high concentration samples should be performed with charcoal-stripped serum (CSS) and at the lowest dilution factor possible. At concentrations commonly used for diagnosis of insulin dysregulation (≤100 µIU/mL), results from the CLIA tend to be lower than from the RIA and should be interpreted accordingly. Further standardization of equine insulin assays is required.

Virological and serological investigation of Equid herpesvirus 1 infection in New Zealand.

Infection with equid herpesvirus 1 (EHV-1) may be asymptomatic, or may result in respiratory disease, abortion, neonatal death, or neurological disease. The aim of this study was to estimate the prevalence of EHV-1 infection, including differentiation between genotypes with aspartic acid (D) and asparagine (N) at position 752 of the DNA polymerase sequence, within a selected population of New Zealand horses. The second aim was to determine the predictive value of serology for detection of latently infected horses. Retropharyngeal lymph nodes (RLN) and trigeminal ganglia (TG) were dissected from 52 horses at slaughter and tested for the presence of EHV-1 DNA using magnetic bead, sequence-capture enrichment followed by nested PCR. Sera were tested for EHV-1 antibody using type-specific glycoprotein G ELISA. Overall, 17/52 horses tested positive for EHV-1 DNA. All but one positive PCR results were obtained from RLN samples. Fifteen of the EHV-1 positive horses harboured EHV-1 with N752 genotype, one of which was additionally infected with the D752 genotypes of the virus. Our data comprise the first detection of EHV-1 with D752 genotype in New Zealand and suggest that the "neurovirulent" variant of EHV-1 had been present in New Zealand for at least two years before the first reported outbreak of EHM. All sampled horses tested positive for EHV-4 antibody, and 11/52 tested positive for EHV-1 antibody. The strength of agreement between results of EHV-1 PCR and EHV-1 serology was "fair" (Kappa 0.259, 95% CI: -0.022-0.539), which was likely a reflection of low levels of both EHV-1 antibody in sera and EHV-1 DNA in tissues tested.

Distal aortic aneurysm presumed to be secondary to an infected umbilical artery in a foal.

CASE HISTORY: A 3-month-old female Warmblood foal was presented after displaying signs of colic with pyrexia for 5 days. CLINICAL AND PATHOLOGICAL FINDINGS: The foal continued to show signs of colic, frequently passed urine, and was pyrexic with an elevated white blood cell count. The umbilical stalk was thickened but there was no evidence of purulent material. Exploratory laparotomy revealed an enlarged left umbilical artery remnant tightly adhered to the bladder wall. The left umbilical artery continued to an aneurysm involving the distal aorta. The foal was subject to euthanasia and post-mortem examination confirmed a spherical aortic aneurysm, in the dorsal midline caudal to the kidneys that contained a large thrombus. Histopathological examination revealed inflammation and necrosis of the tunica intima and tunica media of the left umbilical artery with suppuration and bacterial colonies evident in the periarterial tissues. DIAGNOSIS: Infected aortic aneurysm presumably caused by an umbilical artery infection. CLINICAL RELEVANCE: A previously undetected umbilical infection appears to have resulted in an unusual delayed complication causing signs of colic in a foal. Veterinarians should be aware of this condition, and the possibility that it may be a cause of signs of colic in foals. Diagnosis based on ultrasonography should be possible, but may require sedation, visceral analgesia and careful examination.

A study of neonatal cryptosporidiosis of foals in New Zealand.

AIM: To assess the occurrence of Cryptosporidium oocysts in faecal specimens from foals, and investigate an outbreak of neonatal cryptosporidiosis in foals revealed in the course of the study. METHODS: Faecal specimens from foals received by a diagnostic veterinary laboratory in New Zealand between 2006 and 2007 were submitted to Massey University and tested microscopically for the presence of Cryptosporidium oocysts. The Cryptosporidium isolates in the oocyst-positive specimens were genetically identified to species level. In addition, specimen submission data from the participating laboratory for 2005-2007 were examined. In the course of the study, the identification of one Cryptosporidium-positive specimen triggered an on-farm investigation. RESULTS: Twelve faecal specimens submitted by the participating laboratory between 2006 and 2007 were tested further, and three were positive for C. parvum. Specimen submission records indicated a total of 67 faecal specimens were tested for Cryptosporidium by the participating laboratory between 2005 and 2007; 12 (18%) were positive. The on-farm investigation on a broodmare farm revealed a high incidence of neonatal diarrhoea in foals; C. parvum was the only enteropathogen found in the faeces of 4/4 affected foals examined. The diarrhoea in all those foals was self-limiting, manifesting during the second week of life, resembling foal heat diarrhoea, and accompanied by a short but intense period of shedding oocysts. CONCLUSIONS AND CLINICAL RELEVANCE: The fact that Cryptosporidium parasites were identified in 18% of faecal specimens from foals analysed for this agent in 2005-2007 by the participating laboratory indicated that infection with this agent in foals is not uncommon. Collectively, the results of this and previous studies performed in New Zealand indicate C. parvum is a cause of diarrhoea in newborn foals, potentially accounting for a proportion of cases empirically diagnosed as foal heat diarrhoea. It is therefore advisable to take precautions when handling diarrhoeic foals, until this potentially zoonotic agent is ruled out in the laboratory.

Suture exostosis causing obstruction in the nasolacrimal duct in three horses.

CASE HISTORY: Three horses were presented with facial swelling and epiphora. CLINICAL FINDINGS: All horses had facial swellings and radiographic findings consistent with exostosis at the lacrimomaxillary suture, and ipsilateral epiphora. Positive contrast dacryocystorhinography demonstrated that the site of obstruction of the nasolacrimal duct was located where the duct traverses the lacrimomaxillary suture. DIAGNOSIS: Lacrimomaxillary suture exostosis. CLINICAL RELEVANCE: Lacrimomaxillary suture exostosis should be considered as a differential diagnosis in cases of epiphora in horses. Dacryocystorhinography, preferably retrograde and normograde, may be used as an aid to diagnosis.