Diagnosing feline immunodeficiency virus (FIV) and feline leukaemia virus (FeLV) infection: an update for clinicians

With the commercial release in Australia in 2004 of a vaccine against feline immunodeficiency virus (FIV; Fel‐O‐Vax FIV®), the landscape for FIV diagnostics shifted substantially. Point‐of‐care (PoC) antibody detection kits, which had been the mainstay for diagnosing FIV infection since the early 1990s, were no longer considered accurate to use in FIV‐vaccinated cats, because of the production of vaccine‐induced antibodies that were considered indistinguishable from those produced in natural FIV infections. Consequently, attention shifted to alternative diagnostic methods such as nucleic acid detection. However, over the past 5 years we have published a series of studies emphasising that FIV PoC test kits vary in their methodology, resulting in differing accuracy in FIV‐vaccinated cats. Importantly, we demonstrated that two commercially available FIV antibody test kits (Witness? and Anigen Rapid?) were able to accurately distinguish between FIV‐vaccinated and FIV‐infected cats, concluding that testing with either kit offers an alternative to PCR testing. This review summarises pertinent findings from our work published in a variety of peer‐reviewed research journals to inform veterinarians (particularly veterinarians in Australia, New Zealand and Japan, where the FIV vaccine is currently commercially available) about how the approach to the diagnosis of FIV infection has shifted. Included in this review is our work investigating the performance of three commercially available FIV PoC test kits in FIV‐vaccinated cats and our recommendations for the diagnosis of FIV infection; the effect of primary FIV vaccination (three FIV vaccines, 4 weeks apart) on PoC test kit performance; our recommendations regarding annual testing of FIV‐vaccinated cats to detect ‘vaccine breakthroughs’; and the potential off‐label use of saliva for the diagnosis of FIV infection using some FIV PoC test kits. We also investigated the accuracy of the same three brands of test kits for feline leukaemia virus (FeLV) diagnosis, using both blood and saliva as diagnostic specimens. Based on these results, we discuss our recommendations for confirmatory testing when veterinarians are presented with a positive FeLV PoC test kit result. Finally, we conclude with our results from the largest and most recent FIV and FeLV seroprevalence study conducted in Australia to date.     

Does blood contamination of urine compromise interpretation of the urine protein to creatinine ratio in dogs?

AIMS: To determine the effect of contamination of urine with 0–5% blood, varying in haematocrit and protein concentrations, on the urine protein to creatinine ratio (UPC) in dogs, and to determine whether the colour of urine can be used to aid interpretation of UPC results.

METHODS: Urine samples were collected by free catch from 18 dogs, all of which had UPC?<0.2. Venous blood samples were also collected from each dog, and the blood from each dog was added to its own urine to produce serial concentrations of 0.125–5% blood. The colour of each urine sample was recorded by two observers scoring them as either yellow, peach, orange, orange/red or red. Protein and creatinine concentrations were determined, and dipstick analysis and sediment examination was carried out on each sample. Based on colour and dipstick analysis, samples were categorised as either having microscopic, macroscopic or gross haematuria. A linear mixed model was used to examine the effect of blood contamination on UPC.

RESULTS: The uncontaminated urine of all 18 dogs had a UPC?<0.2. Adding blood to the urine samples resulted in an increase in UPC at all contamination concentrations compared to the non-contaminated urine (p<0.001). None of the 54 samples with microscopic haematuria had UPC?>0.5. For 108 samples with macroscopic haematuria the UPC was >0.5 in 21 samples (19.4 (95% CI=13.1–27.9)%), and for 54 samples with gross haematuria 39 (72 (CI=59.1–82.4)%) had a UPC?>0.5. No samples had a UPC?>2.0 unless the blood contamination was 5% and only 3/18 (17%) samples at this blood contamination concentration had a UPC?>2.0.

CONCLUSIONS AND CLINICAL RELEVANCE: This study showed that while blood contamination of ≥0.125% does increase the UPC, if the urine remains yellow (microscopic haematuria), then there is negligible chance that a UPC?>0.5 will be solely due to the added blood. In that scenario, attributing the proteinuria present to the haematuria in the sample would be inappropriate. However blood contamination that results in discolouration of the urine sample from yellow (indicating macroscopic or gross haematuria) could increase the UPC above the abnormal range and would need to be considered as a differential for the proteinuria. Thus knowledge of urine colour, even if limited to simple colour scores (yellow, discoloured, red) could be utilised to aid interpretation of the UPC in samples with haematuria.     

Pyometra and unilateral uterine horn torsion in a sheep

A 13‐year‐old Romanov sheep presented for evaluation of vaginal discharge, depression and anorexia. Blood work demonstrated mature neutrophilia and marked hyperlactatemia. Transrectal and transabdominal ultrasound revealed echogenic fluid (pus) within the uterus. Purulent uterine fluid culture yield Escherichia coli. A diagnosis of pyometra was made. After medical treatment without clinical improvement, an exploratory laparotomy was performed, and a 360‐degree uterine left horn torsion was identified with the pyometra and multiple corpora lutea on both ovaries. Ovariohysterectomy was performed and the ewe recovered uneventfully.     

Endometrial Expression of IFNAR‐1 and Oxytocin Receptor (OTR) is not Improved by Prostaglandin Analogues when Compared to Progestagens in Ewes

The objective of this study was to investigate differences on the endometrial immunoexpression of type I IFN receptor subunit 1 (IFNAR1) and oxytocin receptor (OTR) during the time of maternal recognition of pregnancy in sheep, when oestrus is synchronized with either prostaglandin analogues (group PG) or conventional progestagens (group P). Plasma progesterone was measured from day 0 to 21 post‐coitus (pc) (day 0 = day of oestrus). Immunohistochemistry was performed in samples of uterine horns from pregnant sheep on days 9pc, 13pc, 15pc, 17pc and 21pc to locate IFNAR1 and OTR expression in different endometrial compartments. Mean levels of plasma progesterone were different between treatments, obtaining higher levels in the PG group than in the P group (p < 0.05). Comparing days of pregnancy, IFNAR1 protein expression was different in the luminal epithelium (LE) (p < 0.05), while OTR was different in the LE and in the superficial glandular epithelium (SG) (p < 0.05). Temporal variation on the expression of both proteins from day 9pc to 21pc has been evidenced. IFNAR1 and OTR expression did not show significant differences between treatments. However, the response observed in the endometrium was highly inconsistent when prostaglandin analogues were used. Therefore, the protocol based on prostaglandin analogues still needs to be optimized before being considered as a better alternative to progestagens for oestrous synchronization in sheep.     

The Effect of Glycerol Concentrations on the Post‐thaw In Vitro Characteristics of Cryopreserved Sex‐sorted Boar Spermatozoa

The objective of this study was to optimize protocols for the cryopreservation of sex‐sorted boar spermatozoa. In the experiment 1, we evaluated the effects of a standard boar sperm cryopreservation procedure (3% final glycerol concentration) on the in vitro characteristics of sex‐sorted sperm frozen at low sperm concentrations (20 × 10⁶ sperm/ml; S20 group). Non‐sorted spermatozoa frozen at 1000 × 10⁶ (C1000 group) and 20 × 10⁶ (C20 group) sperm/ml were used as the freezing control groups. In experiment 2, the effects of different final glycerol concentrations (0.16%, 0.5%, 1.0%, 2.0% and 3.0%) on post‐thaw quality of the S20 and C20 groups were evaluated. In both experiments, the samples were evaluated prior to freezing (5°C) and at 30, 90 and 150 min after thawing. Experiment 1 indicated that freezing sperm at low concentrations decreased (p < 0.05) the total motility (TM) and progressive motility (PM) at 90 and 150 min after thawing regardless of whether the sperm were sorted or not. However, the sperm membrane integrity was not affected at any evaluation step. Inexperiment 2, significant effects on the TM and PM because of increased glycerol concentrations in the S20 and C20 groups were observed only at 90 and 150 min after thawing. The samples frozen in 3% glycerol showed lower (p < 0.05) TM and PM values when compared to those frozen in the presence of 0.5% and 1% glycerol. In both experiments, non‐sorted control samples displayed higher percentages of spermatozoa with damaged DNA than sorted spermatozoa. In conclusion, the optimization of cryopreservation conditions by decreasing the glycerol concentrations can improve post‐thaw motility of sex‐sorted spermatozoa frozen at low concentrations.     

Comparative efficacy of flubendazole and a commercially available herbal wormer against natural infections of Ascaridia galli, Heterakis gallinarum and intestinal Capillaria spp. in chickens

The efficacy of a commercially available flubendazole-based product and a commercially available herbal product were compared against three species of helminth parasites of chickens: Ascaridia galli, Heterakis gallinarum and Capillaria spp. A total of 48 naturally infected chickens were used in the study with 16 birds in each of three treatment groups (untreated control; flubendazole; and a herbal product). One bird from each treatment group was necropsied on Day 0 prior to first treatment to confirm the parasite species present in the birds. Treatments were administered as labelled and the 45 remaining birds were necropsied on Day 12 and worm counts performed. Average worm counts in the two treated groups were compared to the untreated controls to calculate efficacy. Flubendazole (Group A) achieved an overall efficacy of 99.4% for the three parasite species. The herbal product (Group B) achieved efficacies ranging from less than zero to 11.6% for the three parasites, with worm counts not significantly different to the untreated controls. At present, commercially available herbal products claiming anthelmintic properties do not require licencing as veterinary medicinal products (Directive 2004/28/EC: see Article 17 and 33-38) and thus are not required to meet specific efficacy thresholds. Products which do not appear to deliver acceptable anthelmintic efficacy, are obviously a concern from many aspects but specifically from an animal welfare perspective.