Clare Knottenbelt

Evaluation of a rapid assay for canine C-reactive protein

Ms McGrottys current address is Broadleys Veterinary Hospital, Craig Leith Road, Broadleys, Stirling FK7 7LE THE acute phase response is a defence mechanism which occurs in response to tissue damage or inflammation, and includes production of acute phase proteins (APPS) by the liver. Monitoring serum concentrations of APPS provides valuable information on a hosts innate immune response. C-reactive protein (cRP) is the most sensitive APP in dogs, with serum concentrations increasing over 100 times within 24 to 48 hours of stimulation (Conner and others 1988). This increase has been shown to occur in dogs with pyometra, bacterial infections, and following surgery or trauma (Conner and others 1988). In human medicine, CRP assays are used as an aid to diagnosis, prognosis and to assess response to treatment (Pepys and Baltz 1983, Kushner and Mackiewicz 1987). Studies using an ELISA validated for canine CRP determined that healthy animals have a CRP concentration of below 10 mg/litre, while those with an acute phase response have a CRP of above 10 mg/litre (Eckersall and others 1989). Although a test for CRP should provide a valuable addition to diagnostic investigations in canine medicine, it has had limited application due to a lack of availability. A rapid qualitative assay for canine CRP has recently been produced, based on immunochromatography and utilising antibody to canine CRP. The aim of this study was to assess the accuracy of this new method of determining CRP concentrations. Canine serum samples submitted to the University of Glasgow Veterinary School Diagnostic Services for routine diagnostic tests were assayed for CRP concentrations by ELISA. Samples were also obtained from six healthy animals as part of routine health assessment befire undergoing anaesthesia. Selected samples were then further analysed using the CRP Rapid Assay Pack (EVL) by sample dilution followed by immunochromatography on a prepared test strip according to the manufacturers instructions. One drop of serum was diluted in 1 ml of buffer solution and mixed thoroughly. One drop of the mixture was added to the sample zone, followed by three drops of buffer. The results were read after five

Association between Key-Gaskell syndrome and infection by Clostridium botulinum type C/D

There is growing evidence that equine dysautonomia is a toxicoinfection with Clostridium botulinum type C. The possibility that feline dysautonomia has the same aetiology was investigated by attempting to detect botulinum type C neurotoxin in the food, faeces and the contents of the ileum of affected cats, and by serology. The toxin was detected directly in four of eight affected cats and after enrichment in seven of them, and in their dried food. No toxin was detected in healthy control cats or in their tinned food. Recent exposure to the organism was assessed by the detection of immunoglobulin A (IgA) in the faeces of healthy control cats and affected cats. The levels of IgA antibodies to the toxin and to surface antigens of C botulinum type C in the faeces of the affected cats 14 weeks after the outbreak were significantly higher than in the faeces of the control cats.

Haptoglobin concentrations in a canine hospital population

MAVROGENIS, A. P., KOUMAS, A., KAKOYIANNIS, C. K. & TALIOTIS, C. H. (1995) Use of somatic cell counts for the detection of subclinical mastitis in sheep. Small Ruminant Research 17, 79-84 MENZIES, P. I. & RAMANOON, S. Z. (2001) Mastitis of sheep and goats. Veterinary Clinics ofNorth America: Food Animal Practice 17, 333-358 MONTGOMERY, D. C. (1991) Design and Analysis of Experiments. 3rd edn. New York, Wiley MORONI, P. & CUCCURU, C. (2001) Relationship between mammary gland infections and some milk immune parameters in Sardinian breed ewes. Small Ruminant Research 41, 1-7 SARATSIS, P., ALEXOPOULOS, C., TZORA, A. & FTHENAKIS, G. C. (1999) The effect of experimentally induced subclinical mastitis on the milk yield of dairy ewes. Small Ruminant Research 32, 205-209 SCHMIDT-MADSEN, P. (1975) Fluoro-opto-electronic cell counting on milk. Journal ofDairy Research 42, 227-239 WINTER, P. & COLDITZ, I. G. (2002) Immunological responses of the lactating ovine udder following experimental challenge with Staphylococcus epidermidis. Veterinary Immunology and Immunopathology 89, 57-65

Outbreak of dysautonomia (Key-Gaskell syndrome) in a closed colony of pet cats

Six of eight pet cats in a closed colony developed overt signs of dysautonomia over a period of seven days; two of them died and one was euthanased. Dysautonomia was confirmed histopathologically in two of these cats, and in the others the diagnosis was based on the characteristic clinical and radiographic findings. In the two apparently unaffected cats abnormal oesophageal motility was demonstrated by fluoroscopy, suggesting that there may be a subclinical form of the disease. The surviving cats had higher and more variable heart rates (mean 165 bpm) than the non-survivors (mean 121 bpm).

Monitoring the response of canine hyperadrenocorticism to trilostane treatment by assessment of acute phase protein concentrations

BACKGROUND Acute phase proteins (APPS) include haptoglobin (Hp), C-reactive protein (CRP) and serum amyloid A (SAA). Increased Hp concentrations may be induced by endogenous or exogenous glucocorticoids in dogs. OBJECTIVES To assess whether control of hyperadrenocorticism (HAC) affects the concentrations of Hp, CRP, SAA, alkaline phosphatase (ALKP) and cholesterol, to determine whether these analytes can be used to assess control of HAC following trilostane treatment, and whether a combination of these tests offers a valid method of assessing disease control. METHODS Hp, CRP, SAA, ALKP and cholesterol were assessed in 11 dogs with spontaneous HAC before and after treatment with trilostane. Adequate control of HAC was defined as post-ACTH cortisol less than 150 nmol/l. RESULTS Significant reductions in Hp, ALKP, cholesterol and SAA (P<0.05) but not of CRP were found after control of HAC. Only Hp, cholesterol and ALKP were moderately informative (Se & Sp>0.7) of disease control when compared to adrenocorticotropin or corticotropin (ACTH) stimulation test. SAA and CRP were unhelpful (Se & Sp<0.7). The analysis of the combination of the analytes did not improve the correlation with ACTH stimulation test. CLINICAL RELEVANCE Relying on these analytes does not provide additional information over ACTH stimulation test results when assessing control of HAC treated with trilostane.

Determination of nicotine exposure in dogs subjected to passive smoking using methanol extraction of hair followed by hydrophilic interaction chromatography in combination with Fourier transform mass spectrometry

There has been no previous assessment of the level of nicotine exposure in companion animals as a result of passive smoking. A method was developed for the determination of nicotine in dog hair where extraction was carried out by sonication in methanol. The levels of nicotine obtained by extraction with methanol were found to be comparable to the lengthier method involving digestion of the hair in 1M NaOH. The methanol extracts were injected directly onto a ZICHILIC column coupled to an Exactive high resolution Fourier Transform mass spectrometer. Endogenous nicotine was quantified against (2)H(4)-nicotine spiked into the extraction medium, linearity was found over a wide range with the calibration curve having a slope close to 1 indicating an equal response for nicotine and the deuterated internal standard, precision was determined to be ±1.9%. Nicotine was present in widely varying amounts in the hair of dogs belonging to smokers and was found to be absent from the hair of dogs belonging to non-smokers. In addition to nicotine, nicotine N-oxides, cotinine, nornicotine and nornicotine N-oxide could be detected in the hair of dogs belonging to smokers. The nicotine N-oxides were only observed in methanol extracts suggesting these compounds are not stable to the NaOH digestion process.