Polymerase chain reaction tests for the identification of Ross River, Kunjin and Murray Valley encephalitis virus infections in horses

OBJECTIVE: To develop and validate specific, sensitive and rapid diagnostic tests using RT-PCR for the detection of Ross River virus (RRV), Kunjin virus (KV) and Murray Valley encephalitis virus (MVEV) infections in horses. METHODS: Primer sets based on nucleotide sequence encoding the envelope glycoprotein E2 of RRV and on the nonstructural protein 5 (NS5) of KV and MVEV were designed and used in single round PCRs to test for the respective viruses in infected cell cultures and, in the case of RRV, in samples of horse blood and synovial fluid. RESULTS: The primer pairs designed for each of the three viruses amplified a product of expected size from prototype viruses that were grown in cell culture. The identity of each of the products was confirmed by nucleotide sequencing indicating that in the context used the RT-PCRs were specific. RRV was detected in serums from 8 horses for which there were clinical signs consistent with RRV infection such that an acute-phase serum sample was taken and submitted for RRV serology testing. The RRV RT-PCR was analytically sensitive in that it was estimated to detect as little as 50 TCID50 of RRV per mL of serum and was specific in that the primer pairs did not amplify other products from the 8 serum samples. The RRV primers also detected virus in three independent mosquito pools known to contain RRV by virus isolation in cell culture. Samples from horses suspected to be infected with KV and MVEV were not available. CONCLUSION: Despite much anecdotal and serological evidence for infection of horses with RRV actual infection and associated clinical disease are infrequently confirmed. The availability of a specific and analytically sensitive RT-PCR for the detection of RRV provides additional opportunities to confirm the presence of this virus in clinical samples. The RT-PCR primers for the diagnosis of KV and MVEV infections were shown to be specific for cell culture grown viruses but the further validation of these tests requires the availability of appropriate clinical samples from infected horses.     

Myasthenia gravis and masticatory muscle myositis in a dog

A 21-month-old, castrated male Vizsla was presented for pelvic limb weakness, difficulty opening his mouth, ptyalism, voice change, and urinary incontinence. Myasthenia gravis and masticatory myositis were diagnosed. The unusual clinical findings, diagnosis, treatment, and case outcome are described, followed by a brief discussion of myasthenia gravis and masticatory myositis.     

Growth,nutrient utilization,and body composition of dairy calves fed milk replacers containing different amounts of protein

Male Holstein calves < 1 wk of age were allowed a 2-wk adaptation period after purchase, and then were blocked by BW and assigned randomly within block to either a baseline slaughter group or one of four experimental groups (n = 8 to 9 per group). Treatments were isocaloric milk replacers (12.5% solids) fed at 12% of BW that contained 16.1, 18.5, 22.9, or 25.8% CP (DM basis) from whey protein sources. After a 6-wk feeding period, all calves were slaughtered and the weights and chemical composition of the viscera-free carcasses (VFC; including head, hide, feet, and tail) were determined. Gain of BW (0.38, 0.45, 0.56, and 0.62 kg/d) and gain:feed ratio (0.51, 0.59, 0.71, and 0.78) increased linearly (P < 0.001) as dietary CP increased; rate of change in body length, wither height, and heart girth also increased linearly (P < or = 0.05). Balance measurements conducted during wk 3 and 4 of the experimental period showed that both absorbed N (16.9, 20.0, 25.8, and 30.6 g/d) and retained N (7.6, 9.0, 13.2, and 15.6 g/d) increased linearly (P < 0.001) as dietary CP increased. Retained N as a percentage of absorbed N increased linearly (P < 0.01) as dietary CP increased (44.3, 44.7, 50.7, and 50.9%), whereas biological value was unaffected (71.1, 68.7, 69.5, and 67.3%; P = 0.26). Digestible energy and ME represented 94.5 and 89.7% of intake energy, respectively, and were not affected by dietary CP content. Plasma urea N concentration increased linearly (2.9, 3.3, 4.6, and 6.0 mg/dL) as dietary CP increased. Contents of water (68.2, 69.1, 70.2, and 70.5%; P < 0.001) and protein (19.6, 20.0, 20.0, and 20.2%; P < 0.10) in VFC increased linearly, whereas contents of fat (7.2, 6.2, 5.5, and 5.2%; P < 0.001) and ash (5.1, 5.2, 4.8, and 4.7%; P < 0.02) decreased linearly as dietary CP increased. Trends in visceral tissue composition were similar to those for VFC. The content of water in VFC tissue gain increased, whereas contents of fat and energy decreased, as dietary CP increased. Final VFC energy and gain of energy in VFC were not affected by dietary CP. At similar initial ME intakes, increasing dietary CP (i.e., increasing protein: energy) linearly increased ADG, gain:feed, N retention, and deposition of lean tissue in VFC, demonstrating that diet composition can markedly affect components of body growth in preruminant dairy calves.     

Effects of formaldehyde fixation on equine platelets using flow cytometric methods to evaluate markers of platelet activation

OBJECTIVE: To investigate the effects of formaldehyde fixation on equine platelets using flow cytometric methods to evaluate markers of platelet activation. SAMPLE POPULATION: Blood samples from 6 Thoroughbreds. PROCEDURE: The degree of fluorescence associated with binding of fluorescein isothiocyanate (FITC)-conjugated anti-human fibrinogen antibody and FITC-annexin V in unactivated and adenosine diphosphate (ADP)-, platelet activating factor (PAF)-, and A23187-activated platelet samples in unfixed and 0.5, 1.0, and 2.0% formaldehyde-fixed samples was assessed by use of flow cytometry. RESULTS: In samples incubated with FITC-anti-human fibrinogen antibody prior to fixation, addition of 2.0% formaldehyde resulted in a 30% increase in total fluorescence in ADP- and PAF-activated samples and a 60% increase in A23187-activated samples. Fixation for 24 hours prior to addition of antibody resulted in reduced fluorescence of samples containing antihuman fibrinogen antibody for all 3 concentrations of formaldehyde in PAF-activated samples. The addition of all 3 concentrations of formaldehyde after incubation with FITC-annexin V resulted in significant increases in fluorescence in unactivated and activated platelet samples. As length of fixation time increased, there was a gradual increase in fluorescence that was significant at 24 hours. CONCLUSIONS: Because fixation with 2.0% formaldehyde results in significant changes in fluorescence in activated platelet samples containing anti-fibrinogen antibody, lower concentrations of formaldehyde should be used to fix equine platelet samples. Formaldehyde-fixed platelet samples should be analyzed within 12 hours of fixation to avoid artifactual increases in fluorescence. Fixation of samples containing FITC-annexin V should be avoided because of significant increases in fluorescence that may interfere with interpretation of results.     

Comparison of a putative second serotype of chicken infectious anemia virus with a prototypical isolate I. Pathogenesis

CIAV-7 is a virus with similar pathogenic and physicochemical characteristics to, but antigenically distinct from, chicken infectious anemia virus (CIAV). The pathogenesis of CIAV-7 was evaluated in a comparative study with a representative isolate of CIAV, the Del-Ros strain. The pathogenesis of CIAV-7 was similar to Del-Ros on the basis of the clinical disease induced and gross and microscopic lesions, although CIAV-7 produced fewer and less severe lesions overall. A second comparative pathogenesis study was performed with Del-Ros and CIAV-7, both alone and in combination with infectious bursal disease virus (IBDV). In this study, the pathogenesis of CIAV-7 was similar to Del-Ros in clinical, gross, and microscopic lesions in the bone marrow. However, thymic lesions were less severe in CIAV-7-inoculated birds. The interaction between Del-Ros and IBDV was synergistic, whereas there was no observed potentiation of CIAV-7-induced disease by IBDV. Progeny from breeder flocks from several geographic locations in the eastern United States were challenged with CIAV-7 or Del-Ros to assess protection by maternal antibodies. Some progeny from all flocks had protection against CIAV-7 challenge, providing evidence for the presence of CIAV-7 in the field. Additionally, the number of birds protected against CIAV-7 or Del-Ros challenge varied within flocks, demonstrating that the agents are serologically distinct.     

Effect of lameness on milk yield in dairy cows

OBJECTIVE: To examine the relationship between lameness and milk yield in dairy cows. DESIGN: Cohort study. ANIMALS: 531 dairy cows. PROCEDURE: Cows affected with lameness were classified into 1 of 3 groups on the basis of type of diseases or lesions observed, including interdigital phlegmon (foot rot), papillomatous digital dermatitis (foot warts), or claw lesions. Cows not affected with lameness were classified as healthy. From Dairy Herd Improvement Association records, 305-day mature equivalent milk yield data were collected at the end of lactation or when the cow left the herd. Milk yield was compared between cows affected with lameness and healthy cows. RESULTS: 167 (31%) cows were affected with lameness during lactation. Lame cows had claw lesions (60%), papillomatous digital dermatitis (31%), or interdigital phlegmon (9%). Milk yield in lame cows with interdigital phlegmon (mean, 17,122 lb) was significantly less, compared with healthy cows (19,007 lb). CONCLUSIONS AND CLINICAL RELEVANCE: In this herd, interdigital phlegmon was associated with a 10% decrease in milk production. Lame cows with claw lesions or papillomatous digital dermatitis produced less milk than healthy cows, but the difference was not significant.     

Isolated canine and murine intestinal cells exhibit a different pattern of fuel utilization for oxidative metabolism

The amount and type of dietary fiber influences the end-products of fermentation and thus fuel availability to intestinal tissue. Metabolic fuel usage was studied in intestinal cells isolated from dogs consuming a commercial diet or from rats consuming either a commercial rat diet or dog diet to examine preferential fuel usage, the effect of diet, and species differences. Production of 14CO2 was measured by incubating cells in media containing either D-[U-14C]glucose, [1-14C]n-butyrate, L-[U-14C]glutamine, or [1-14C]propionate with or without competing substrates. The presence of a mixture of 5 mM each of glucose, butyrate, propionate, and acetate and 1 mM glutamine in the media decreased CO2 production from glucose, glutamine, and propionate by canine enterocytes (P < 0.05) and from glutamine and propionate by canine colonocytes (P < 0.05). The presence of glutamine in the media decreased glucose oxidation by murine enterocytes, regardless of the diet. Similarly, glutamine decreased glucose oxidation by murine colonocytes (P < 0.05), but only when the rats had consumed the rat diet. Regardless of diet, murine colonocytes oxidized more butyrate (P < 0.01) than did enterocytes, and murine enterocytes tended (P < 0.07) to oxidize more glucose than did colonocytes. The proportion of propionate in colonic contents was higher in dogs than in rats (P < 0.02), and the proportion of butyrate tended to be higher in contents from rats than in those from dogs (P < 0.08). Colonic and cecal wet weights were decreased (P < 0.05) when rats were fed the commercial dog diet. Preferred utilization of substrates by isolated canine enterocytes and colonocytes differed from that of murine intestinal cells. These differences were only partially overcome by feeding the same diet to each species.     

Restricted suckling of tropical dairy cows by their own calf or other cows' calves

The objective of this study was to compare restricted suckling of tropical cows by their own or another cow's calf with artificial rearing of the calves and no suckling. In Exp. 1, cows were mechanically milked twice daily, after which for 15 min they were either suckled by their own calf (Treatment O) or multiple-suckled by other cows' calves (Treatment M) or unsuckled, with the calves reared artificially (Treatment A). Machine milk yield was similar for the three treatments, but in the two suckling treatments the additional milk consumed by the calf increased (P = 0.02) total production (2,682, 2,634, and 2,336 kg/lactation for Treatments O, M, and A, respectively). Machine milk fat concentration was reduced (P = 0.05) by suckling (2.90, 3.07, and 3.20% for Treatments 0, M, and A, respectively), but the milk sampled just before suckling (to represent that taken by the calves) had a high fat concentration (mean 7.9%). Machine milk somatic cell count was also reduced (P = 0.05) by suckling, from 106,000/mL (Treatment A) to 85,000/mL (Treatment M) and 95,000 (Treatment O). Cows suckling their own calf lost more weight and body condition than cows whose calves were reared artificially, with multiple-suckled calves intermediate. Cows suckling their own calf had postpartum interval to first estrus increased (P = 0.01) by 31 d and conception rates to first service of 44% compared to 77% for the other two treatments (P = 0.01). The growth of the suckled calves was compared with that of the artificially reared calves, which were given recommended milk allowances. The artificially reared calves consumed more milk and concentrates, which were available ad libitum to all calves, and gained (P = 0.03) 0.07 kg/d more weight than suckled calves. A second experiment determined that suckling once daily did not reduce reproductive performance compared to artificial rearing. We conclude that suckling cows twice daily increases total milk production but reduces body weight in early lactation. Cows suckling their own calves have reduced reproductive performance compared to those suckling other calves or reared artificially.