Describing the within laboratory and between laboratory agreement of a serum ELISA in a national laboratory network

Results from laboratory assays for detection of animal disease are often assessed for repeatability (agreement within laboratory) and reproducibility (agreement between laboratories). This work aimed to understand the strengths and limitations of available methods for describing these quantities. Five major veterinary laboratories in Australia volunteered to participate in a designed evaluation based on repeat testing of twenty bovine sera. Sampling was stratified so that ten of the sera were negative to the virus neutralisation test (VNT) for antibody to bovine herpes virus 1 (BHV-1) and the remaining ten sera were VNT positive. Each serum was divided into 50 replicates and each laboratory assayed one replicate of each serum on a weekly basis using a commercial ELISA for BHV-1. Laboratories were blinded to the identity of sera. The data on sample to positive control ratio (S/P) for these 1000 individual assays were collated, sources of variance analysed using a random effects model, and reliability coefficients (ρ) obtained from the variance estimates as quantitative measures of within and between laboratory agreement. Coefficient of variation (CV) was calculated for combinations of sera and laboratory. CV was found to be higher for sera with the lowest mean S/P values (VNT -ve sera). For VNT -ve sera, agreement of S/P within laboratory was low to moderate (ρ: 0.01-0.27) and the agreement between all labs was low (ρ=0.02). Reliability coefficients for VNT +ve sera were very high for agreement within laboratories (ρ: 0.63-0.92) and moderate for agreement between laboratories (ρ=0.52). As well, simulation demonstrated that sero-prevalence has a dramatic affect on the reliability coefficient if sampling were to be irrespective of VNT status. We conclude that there are some limitations with the available approaches for assessing agreement within and between laboratories. Although reliability coefficients have some drawbacks they are an attractive way of reducing reliance on subjective assessment of agreement.     

Farm and socio-economic characteristics of smallholder milk producers and their influence on technology adoption in Central Mexico

In order to identify the factors influencing adoption of technologies promoted by government to small-scale dairy farmers in the highlands of central Mexico, a field survey was conducted. A total of 115 farmers were grouped through cluster analysis (CA) and divided into three wealth status categories (high, medium and low) using wealth ranking. Chi-square analysis was used to examine the association of wealth status with technology adoption. Four groups of farms were differentiated in terms of farms' dimensions, farmers' education, sources of incomes, wealth status, management of herd, monetary support by government and technological availability. Statistical differences (p < 0.05) were observed in the milk yield per herd per year among groups. Government organizations (GO) participated little in the promotion of the 17 technologies identified, six of which focused on crop or forage production and 11 of which were related to animal husbandry. Relatives and other farmers played an important role in knowledge diffusion and technology adoption. Although wealth status had a significant association (p < 0.05) with adoption, other factors including importance of the technology to farmers, usefulness and productive benefits of innovations together with farmers' knowledge of them, were important. It is concluded that the analysis of the information per group and wealth status was useful to identify suitable crop or forage related and animal husbandry technologies per group and wealth status of farmers. Therefore the characterizations of farmers could provide a useful starting point for the design and delivery of more appropriate and effective extension.     

Cataracts are not associated with retinal detachment in the Bichon Frise in the UK--a retrospective study of preoperative findings and outcomes in 40 eyes

Objective The aim of this study was to evaluate whether the Bichon Frise population in the UK is at the same risk of developing retinal detachment in association with cataract formation and following phacoemulsification as described in reports from the USA. Procedures The medical records of Bichon Frises which were presented for cataract assessment and of those which were treated with phacoemulsification at Willows Referral Service between 1997 and 2009 were reviewed. Results Forty eyes (26 dogs) with unilateral or bilateral cataracts were included in the study. There was no evidence of retinal detachment associated with the cataracts at initial presentation. Phacoemulsification was performed on 34 eyes (20 dogs). Clinically evident lens‐induced uveitis was treated preoperatively in 17/34 eyes. Artificial lens implantation was carried out in 30/34 eyes; automated anterior vitrectomy was performed in 7/34 eyes. The mean follow‐up time was 16.6 months (range 1.5–73 months). At the last re‐examination, 31/34 eyes (91.2%) were visual. Three eyes (8.8%) were blind – two (in the same dog) because of presumptive bilateral optic nerve disease and one because of uveitis and secondary glaucoma. There was no evidence of retinal detachment following phacoemulsification in any of the 34 eyes. Conclusion This study suggests that the Bichon Frise population in the UK does not appear to have a predisposition for retinal detachment in association with cataract formation or following cataract surgery. Prophylactic random transscleral laser retinopexy or transscleral cryopexy cannot therefore be routinely recommended for Bichon Frises with cataracts in the UK.     

Identification and localization of the structural proteins of anguillid herpesvirus 1

ABSTRACT: Many of the known fish herpesviruses have important aquaculture species as their natural host, and may cause serious disease and mortality. Anguillid herpesvirus 1 (AngHV-1) causes a hemorrhagic disease in European eel, Anguilla anguilla. Despite their importance, fundamental molecular knowledge on fish herpesviruses is still limited. In this study we describe the identification and localization of the structural proteins of AngHV-1. Purified virions were fractionated into a capsid-tegument and an envelope fraction, and premature capsids were isolated from infected cells. Proteins were extracted by different methods and identified by mass spectrometry. A total of 40 structural proteins were identified, of which 7 could be assigned to the capsid, 11 to the envelope, and 22 to the tegument. The identification and localization of these proteins allowed functional predictions. Our findings include the identification of the putative capsid triplex protein 1, the predominant tegument protein, and the major antigenic envelope proteins. Eighteen of the 40 AngHV-1 structural proteins had sequence homologues in related Cyprinid herpesvirus 3 (CyHV-3). Conservation of fish herpesvirus structural genes seemed to be high for the capsid proteins, limited for the tegument proteins, and low for the envelope proteins. The identification and localization of the structural proteins of AngHV-1 in this study adds to the fundamental knowledge of members of the Alloherpesviridae family, especially of the Cyprinivirus genus.     

Bovine neonatal pancytopenia in German Holstein calves

Profiles of blood cell counts were evaluated for 15 calves from three different farms. These calves showed petechia in the mucous membranes and in the skin and prolonged secondary bleeding after puncture. The clinical course of the disease could be observed in eleven calves. With exception of one case, the blood cell counts indicated a severe anaemia, leukocytopenia and thrombocytopenia. Out of these 15 calves, six calves survived and the other nine calves died or had to be euthanized due to the severity of the disease. Necropsy of these nine calves revealed petechia in the skin, subcutis, muscles, in inner organs and all serous membranes. Pathohistological examination showed a depletion of the bone marrow and lymphatic tissue in eight calves. These findings confirmed the diagnosis of bovine neonatal pancytopenia (BNP) for eight of these nine calves. Bluetongue virus serotype 8 was tested negatively using PCR. Bovine virus diarrhoea virus (BVDV) was negatively tested using immunofluorescence and cell culture and salmonella species were negatively tested in seven dissected calves. A cluster of toxins was negatively tested in one of the dissected calves. All 15 calves had high antibody titres for BVDV. The BVDV-antibody titres from twelve dams with affected calves were positive in six cases and not detectable in the other six cases. In three of the six dams with not detectable BVDV-antibody titres, calves were fed with colostrum of a further dam with high BVDV-antibody titres. In the further three dams without detectable BVDV-antibody titres, we could not ascertain which colostrum has been fed to the calves. BVDV-specific antigen could not be detected in any of the samples from the calves and dams tested. Using the activity of the gamma-glutamyl-transferase, we assumed a sufficient supply with colostrum for the examined calves.The cause for the occurrence of these BNP cases was due to bone marrow depletion.The reason for the bone marrow depletion remained unclear. However, it was obvious that the BNP described here is highly likely caused by colostrum from cows with positive BVDV-antibody titres.     

First morphological characterization of 'Candidatus Mycoplasma turicensis' using electron microscopy

At least three haemotropic mycoplasmas have been recognized in cats: Mycoplasma haemofelis (Mhf), 'Candidatus Mycoplasma haemominutum' (CMhm) and 'Candidatus M. turicensis' (CMt). The latter was originally identified in a Swiss pet cat with haemolytic anaemia and shown to be prevalent in domestic cats and wild felids worldwide using molecular methods. So far, there has been no confirmatory morphological evidence of the existence of CMt presumably due to low blood loads during infection while CMhm has only been characterized by light microscopy with discrepant results. This study aimed to provide for the first time electron microscopic characteristics of CMt and CMhm and to compare them to Mhf. Blood samples from cats experimentally infected with CMt, CMhm and Mhf were used to determine copy numbers in blood by real-time PCR and for transmission and scanning electron microscopy. High resolution scanning electron microscopy revealed CMt and CMhm to be discoid-shaped organisms of 0.3 μm in diameter attached to red blood cells (RBCs). In transmission electron microscopy of CMt, an oval organism of about 0.25 μm with several intracellular electron dense structures was identified close to the surface of a RBC. CMhm and CMt exhibited similar morphology to Mhf but had a smaller diameter. This is the first study to provide morphological evidence of CMt thereby confirming its status as a distinct haemoplasma species, and to present electron microscopic features of CMhm.     

Effect of dietary soy on serum thyroid hormone concentrations in healthy adult cats

OBJECTIVE: To compare effects of short-term administration of a soy diet with those of a soy-free diet on serum thyroid hormone concentrations in healthy adult cats. ANIMALS: 18 healthy adult cats. PROCEDURE: Cats were randomly assigned to receive either a soy or soy-free diet for 3 months each in a crossover design. Assays included CBC, serum biochemical profile, thyroid hormone analysis, and measurement of urinary isoflavone concentrations. RESULTS: Genistein, a major soy isoflavone, was identified in the urine of 10 of 18 cats prior to dietary intervention. Compared with the soy-free diet, cats that received the soy diet had significantly higher total thyroxine (T4) and free T4 (fT4) concentrations, but unchanged total triiodothyronine (T3) concentrations. The T3/fT4 ratio was also significantly lower in cats that received the soy diet. Although the magnitudes of the increases were small (8% for T4 and 14% for fT4), these changes resulted in an increased proportion of cats (from 1/18 to 4/18) that had fT4 values greater than the upper limit of the laboratory reference range. There was no significant effect of diet on any other measured parameter. CONCLUSIONS AND CLINICAL RELEVANCE: Short-term administration of dietary soy has a measurable although modest effect on thyroid hormone homeostasis in cats. Increase in T4 concentration relative to T3 concentration may result from inhibition of 5'-iodothyronine deiodinase or enhanced T3 clearance. Soy is a common dietary component that increases serum T4 concentration in cats.     

Echocardiographic estimation of mean left atrial pressure in a canine model of acute mitral valve insufficiency

High mean left atrial pressure (MLAP) due to canine degenerative mitral valve disease is associated with clinically relevant morbidity and mortality. The ability to noninvasively measure MLAP would assist in the diagnosis and treatment of disease. Doppler echocardiography allows measurement of early transmitral blood flow (E) and the velocity of the mitral valve annulus (Ea). The ratio of early mitral inflow velocity to early mitral annular velocity (E: Ea) correlates well with MLAP in human subjects. We sought to determine the ability of E: Ea to predict MLAP in dogs with experimentally induced mitral regurgitation. Nine anesthetized purpose-bred dogs underwent placement of a Swan-Ganz catheter into the left atrium and recording of MLAP. Simultaneous transthoracic echocardiographic and hemodynamic studies were performed after acute chordae tendineae rupture and during IV infusion with nitroprusside (2.5-5.0 microg x kg(-1) x min(-1)) or hydralazine (1-1.5 mg/kg). Mitral regurgitant fraction, measured by single-plane angiography and thermodilution, ranged from 17% to 81%. MLAP increased from 5.4 +/- 2.5 mm Hg to 17.4 +/- 9.4 mm Hg after creation of mitral valve regurgitation (MR; P = .018). Forty sets of echocardiographic measurements were obtained from 7 dogs, and E, as well as E: Ea, were linearly related to MLAP. The R2 value for the linear regression equation containing E: Ea as the dependent variable (0.83) was greater than that for E (0.73). The 95% confidence intervals were calculated for predicting MLAP = 20 mm Hg from E:Ea, and E:Ea >9.1 or <6.0 indicated a 95% probability that MLAP was >20 mm Hg or <20 mm Hg, respectively. Echocardiography can be used to predict MLAP in isoflurane-anesthetized dogs with experimentally induced acute mitral valve insufficiency.     

Postoperative assessment of surgical clip position in 16 dogs with cancer: a pilot study

Metallic hemoclips or surgical staples were inserted in 16 tumor-bearing dogs at the time of surgical resection of the tumor. Orthogonal radiographs were taken immediately postoperatively and after wound healing to visualize the location and number of hemoclips or metallic staples. A shift in hemoclip/staple position was identified in nine dogs, mainly from positioning during radiography. In three dogs, an absolute shift in marker position was identified. Based on this study, it appears that the placement of surgical clips is potentially useful in identifying the tumor bed, which may be of benefit in establishing radiation treatment fields.