Detection and quantification of equine herpesvirus-1 viremia and nasal shedding by real-time polymerase chain reaction.

Equine herpesvirus-1 (EHV-1) infection is common in young horses throughout the world, resulting in respiratory disease, epidemic abortion, sporadic myelitis, or latent infections. To improve on conventional diagnostic tests for EHV-1, a real-time polymerase chain reaction (PCR) technique was developed, using primers and probes specific for the EHV-1 gB gene. Amplification efficiencies of 100% +/- 5% were obtained for DNA isolated from a plasmid, infected peripheral blood mononuclear cells (PBMCs), and nasal secretions from infected ponies. The dynamic range of the assay was 8 log10 dilutions, and the lower limit of detection was 6 DNA copies. Fifteen ponies, seronegative for EHV-1, were experimentally infected with EHV-1, and nasal samples were used to quantify shedding of virus by both virus isolation and real-time PCR analysis. Virus isolation identified nasal shedding of EHV-1 in 12/15 ponies on a total of 25 days; real-time PCR detected viral shedding in 15/15 ponies on 75 days. Viremia was quantified using PBMC DNA, subsequent to challenge infection in 3 additional ponies. Viremia was identified in 1/3 ponies on a single day by virus isolation; real-time PCR detected viremia in 3/3 ponies on 17 days. When real-time PCR was used to analyze PBMC DNA from 11 latently infected ponies (documented by nested PCR), EHV-1 was not detected. We conclude that real-time PCR is a sensitive and quantitative test for EHV-1 nasal shedding and viremia and provides a valuable tool for EHV-1 surveillance, diagnosis of clinical disease, and investigation of vaccine efficacy.     

Prevalence of parasite eggs and cysts in faeces from dairy cows in Colombia

Summary When 1, 711 bovine faecal samples from 113 farms in eight dairy areas of Colombia were examined for the presence of helminth eggsFasciola hepatica eggs were found in the faeces from 60% of the farms and samples from animals kept above 2,000 m. Strongyle eggs were found in faeces from 82% of the farms and in 18% of the samples.

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Resumen Se examinaron 1.711 muestras fecales provenientes de 113 fincas, localizadas en ocho zonas lecheras de Colombia. Se encontraron huevos deFasciola hapatica, en el 60% de fincas y muestras fecales de animales sobre los 2,000 m.s.n.m. Se encontraron tambien huevos de estrogilideos en heces del 82% de fincas y en el 18% de las muestras examinadas.

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Résumé La recherche d’oeufs d’helminthes sur 1711 échantillons de fécès provenant de 113 fermes de 8 districts laitiers de Colombie a permis de noter la présence d’oeufs deFasciola hepatica dans 60 p. 100 des fermes et échantillons originaires d’animaux vivant à plus de 2,000 m. Des oeufs de strongles ont été notés dans les fécès provenant de 82 p. 100 des fermes et 18 p. 100 des échantillons.

     

Metastatic immune infiltrates correlate with those of the primary tumour in canine osteosarcoma

Our lack of understanding of the immune microenvironment in canine osteosarcoma (cOSA) has limited the identification of potential immunotherapeutic targets. In particular, our ability to utilize readily available tissue from a dog's primary tumour to predict the type and extent of immune response in their pulmonary metastatic lesions is unknown. We, therefore, collected 21 matched pairs of primary tumours and pulmonary metastatic lesions from dogs with OSA and performed immunohistochemistry to quantify T‐lymphocyte (CD3), FOXP3+ cell, B‐lymphocyte (Pax‐5), and CD204+ macrophage infiltration. We found that T‐lymphocytes and FOXP3+ infiltrates in primary tumours positively correlated with that of metastatic lesions (ρ = 0.512, P = 0.038 and ρ = 0.698, P = 0.007, respectively), while a strong trend existed for CD204+ infiltrates (ρ = 0.404, P = 0.087). We also observed T‐ and B‐lymphocytes, and CD204+ macrophages to be significantly higher in a dog's pulmonary metastasis compared to their primary tumour (P = 0.018, P = 0.018, P = 0.016, respectively), while FOXP3+ cells were only significantly higher in metastases when all primary tumour and metastasis lesions were compared without pairing (P = 0.036). Together, these findings suggest that the metastatic immune microenvironment may be influenced by that of the primary cOSA, and that primary tumour immune biomarkers could potentially be applied to predict immunotherapeutic responses in gross metastatic disease. We, therefore, provide a rationale for the treatment of cOSA pulmonary metastases with immunotherapeutics that enhance the anti‐tumour activity of these immune cells, particularly in dogs with moderate to high immune cell infiltration in their primary tumours.     

Demonstration of uniformity of calcium absorption in adult dogs and cats

A meta‐analysis was conducted to understand quantitative aspects of calcium (Ca) and phosphorus (P) absorption in adult dogs and cats. 34 studies in dogs and 14 studies in cats met the criteria for inclusion in the meta‐analysis. Intake and faecal excretion values of Ca and P were subjected to a modified Lucas test and subsequent regression analyses. According to the current scientific consensus, Ca true digestibility (absorption) should increase at low Ca intake and decrease at high Ca intake. If true, this should result in a nonlinear relationship between the percentage of Ca excreted and dietary Ca intake. The present meta‐analysis showed a highly significant linear relationship (p < 0.0001) between Ca intake and Ca excretion suggesting a lack of systematic quantitative adaptation in true Ca digestibility. This finding suggests either that the time period covered by standard digestion trials is too short to induce adaptation mechanisms or that dogs and cats at maintenance will not efficiently alter quantitative Ca absorption percentage according to the amount ingested. If the latter is true, a dietary Ca supply differing greatly from the recommended dietary intake might impair the health of cats and dogs when fed long term. The data plots for P intake and faecal excretion were less uniform suggesting other factors not just dietary intake influence faecal P excretion. In adult cats, the dietary Ca:P ratio strongly influenced the true digestibility of P, whereas this effect was less marked in adult dogs. Faecal P excretion was significantly correlated to faecal Ca excretion in both species (p < 0.0001), and surprisingly, the level of P intake did not appear to be an important determinant of true digestibility of P.     

Effects of wet corn gluten feed and intake level on diet digestibility and ruminal passage rate in steers

Twelve ruminally cannulated Jersey steers (BW = 534 kg) were used in an incomplete Latin square design experiment with a 2 x 2 factorial arrangement of treatments to determine the effects of wet corn gluten feed (WCGF) and total DMI level on diet digestibility and ruminal passage rate. Treatments consisted of diets formulated to contain (DM basis) steam-flaked corn, 20% coarsely ground alfalfa hay, and either 0 or 40% WCGF offered once daily for ad libitum consumption or limited to 1.6% of BW (DM basis). Two consecutive 24-d periods were used, each consisting of 18 d for adaptation, 4 d for collection, and a 2-d in situ period. Rumens of all steers were evacuated once daily at 0, 4, 8, and 12 h after feeding. Chromic oxide (10 g/[steer*d]) was fed as a digestibility marker, and steers were pulse-dosed with Yb-labeled alfalfa hay to measure ruminal particulate passage rate. Dacron bags containing 5 g of steam-flaked corn, WCGF, or ground (2-mm screen) alfalfa hay were placed into the rumens of all steers and removed after 3, 6, 12, or 48 h. Wet corn gluten feed increased percent apparent total-tract digestion of OM (P < 0.01), NDF (P < 0.01), and starch (P < 0.03), decreased (P < 0.01) ruminal total VFA concentration, increased (P < 0.01) ruminal NH3 concentration, and increased (P < 0.01) ruminal pH. Wet corn gluten feed also increased (P < 0.01) ruminal passage rate of Yb. Limit feeding decreased (P < 0.01) percent apparent total-tract digestion of both OM and NDF, ruminal total VFA concentration (P < 0.01), and ruminal fill (P < 0.01), but increased (P < 0.01) ruminal NH3 concentration. Apparent total-tract digestion of starch was not affected (P = 0.70) by level of DMI. A DMI level x hour interaction (P < 0.01) occurred for ruminal pH. Limit feeding increased ruminal pH before and 12 h after feeding, but decreased ruminal pH 4 h after feeding compared with diets offered ad libitum. A diet x DMI level interaction (P < 0.02) occurred for in situ degradation of alfalfa hay, with dietary addition of WCGF increasing (P < 0.02) the extent of in situ alfalfa hay degradation in steers fed for ad libitum consumption. This study suggests that WCGF increases OM and NDF digestion, and that limit feeding diets once daily might depress OM and NDF digestion, possibly due to decreased stability of the ruminal environment.     

Effects of some nonionic polyoxyethylene surfactants on uptake of ethirimol and diclobutrazol from suspension formulations applied to wheat leaves

The influence of a number of commercial nonionic polyoxyethylene surfactants on the foliar penetration and movement of two systemic fungicides, ethirimol and diclobutrazol, was studied in outdoor-grown wheat plants at different growth stages and post-treatment temperatures in two consecutive growing seasons. Both fungicides were applied as ca 0·2 μl droplets of aqueous suspension formulations containing 0·5 g litre^?1 of ¹⁴C-labelled active ingredient; surfactants were added to these suspensions at concentrations ranging from 0·2-10 g litre^?1. To achieve optimum uptake of each fungicide the use of surfactants with different physicochemical properties was required. For diclobutrazol, a lipophilic compound, uptake of radiolabel was best with surfactants of low mean molar ethylene oxide (E) content (5-6) but it was necessary to use concentrations of ca 5 g litre^?1 to attain this. The surfactant threshold concentration for uptake enhancement of radiolabel from ethirimol formulations (< 2 g litre^?1) was much lower than that for diclobutrazol but surfactants with E contents > 10 induced the greatest amount of uptake. For both fungicides, surfactants with an aliphatic alcohol hydrophobe were generally more efficient in promoting their uptake than those with a nonylphenol moiety. The sorbitan-based surfactant ‘Tween 20’ proved to be an effective adjuvant only for the ethirimol formulation; the uptake enhancing properties of the block copolymer ‘Synperonic PE/F68’ were weak. Uptake performance could not be related to the spreading properties of the respective formulations on the wheat leaf surface or to differences in solubilisation of the two fungicides by the surfactants. Although surfactants could substantially increase the amount of acropetal transport of radiolabel from both fungicides, none of those tested specifically promoted it; a constant proportion of the radioactive dose absorbed by a treated leaf was usually exported away from the site of application. The results are discussed in the light of current theories about the mode of action of surfactants as spray adjuvants.