Polymerase chain reaction and other laboratory techniques in the diagnosis of long incubation rabies in Australia

SUMMARY Blood and post-mortem tissues from a 10-years-old girl were submitted to the Australian Animal Health Laboratory. Clinical signs and histopathological lesions had suggested a diagnosis of rabies, but, an unusually long incubation period of at least 5 years did not encourage such a diagnosis. Serological examinations by the rapid fluorescent focus inhibition test revealed a dramatic increase in rabies virus-neutralising antibody during the 10-day period of hospitalisation. The results of a fluorescent antibody test on brain smears, and an immunoperoxidase test on formalin-fixed sections of brain were also consistent with a diagnosis of rabies. Attempts to isolate virus were unsuccessful. Polymerase chain reactions (PCRs) were conducted on a 10% suspension of a post-mortem sample from the patient's brain, using primers based on the published sequence of the Pasteur virus strain of rabies virus. 413 and 513 bp fragments from the nucleoprotein gene and a 403 bp fragment from the glycoprotein gene were amplified. Subsequent sequencing of these fragments, and comparison with equivalent regions of known rabies viruses, confirmed that the fragments originated from a virus belonging to the rabies virus serotype. This case demonstrated the advantage of using a range of laboratory techniques to obtain a definitive diagnosis. In particular, a PCR-based test may allow a diagnosis, even in the face of conditions that preclude virus isolation such as apparently occurred in this case. Finally, this case demonstrated that an unusually long incubation period should not discourage a tentative clinical diagnosis of rabies.     

Seminal Plasma Composition from Ejaculates Collected by Artificial Vagina and Electroejaculation in Guirra Ram

This study was conducted to evaluate changes in ram seminal plasma composition from ejaculates obtained using artificial vagina (AV) and electroejaculation (EE). To address this question, we assessed the effect of semen collection method on volume, sperm concentration, sodium concentration, potassium concentration, sodium/potassium ratio, total protein content and protein profile using sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and 2-D polyacrylamide gel electrophoresis. The main findings from this study were: (i) similar volume was obtained, while sperm concentration was significantly lower for EE method; (ii) potassium and sodium/potassium concentration ratio were not influenced by recovery method, while sodium concentration increased significantly when semen was recovered using EE; (iii) approximately 80% of the total relative seminal plasma protein is represented by four protein fractions of molecular weights around 15, 21, 24 and 50 kDa and there were not differences and (iv) focussing the two-dimensional SDS-PAGE gel on the 10–25 kDa rank, the image analysis software detected around 22 spots with isoelectric points ranging from 5.1 to 6.1. Two protein spots (15 kDa and 5.5 and 22 kDa and 5.2 for molecular weight and isoelectric point respectively) increased significantly when semen was recovered using EE. One spot protein with molecular weight around 25 kDa and isoelectric point of 5.2 were only found in the seminal plasma from the semen recovery by AV. As it was demonstrated, ejaculates obtained with EE modify the sodium concentration, alter two proteins concentration and induced the loss of one protein in seminal plasma.     

When less is more: heterogeneity in grass patch height supports herbivores in counter-intuitive ways

Herbivores are an integral part of the African landscape and have evolved with the vegetation to create the savanna landscape. Managers of these landscapes can benefit from a better understanding of how indigenous herbivores use the landscape to which they are adapted. In this study we observed which patches were frequently utilised, by doing regular monthly road counts, grass height observations and dung counts on selected short grass patches in the Kruger National Park. Smaller-framed impala and blue wildebeest (meso-herbivores) were most regularly seen on these nutritious patches, while from dung deposits it was clear that the even larger-framed buffalo (mega-herbivores) spent time there. This preference can be explained by considering the nutritional needs and food intake of the herbivores. Smaller-framed herbivores seem to be able to satisfy their dietary requirements on the high-quality forage patches, while larger-framed herbivores seem to supplement the quality forage by also spending foraging time on areas of higher grass biomass. From this insight we propose that range management should take herbivore preferences into account and allow herbivores to select and concentrate their foraging on the most nutritious forage. This approach is likely to decrease inputs while allowing animals to maintain or increase production.     

Systemic granulomatous disease in dairy cattle during a dicyandiamide feeding trial

CASE HISTORY: Mature, in-calf, non-lactating, Friesian or Friesian-cross cows were fed dicyandiamide (DCD) at daily doses of 0.15 g/kg (Group 1; n=31), 0.45 g/kg (Group 2; n=21) and 0.75 g/kg (Group 3; n=12), as part of a safety trial, which also included a control group (n=15). Daily health observations were carried out on each cow until Day 86 of the study. On Day 28 one cow from Group 3 was observed with signs of disease, and subsequently disease was noted in other cows.

CLINICAL FINDINGS: Clinical signs in the first case included depression, pyrexia (40.9°C), salivation and dehydration, in addition to progressive weight loss, followed by death on Day 32. Other cows from all treatment groups developed clinical signs of disease resulting in euthanasia of seven animals. Disease occurred in 10/12 (83%) cows in Group 3, 11/21 (52%) cows in Group 2, and 7/31 (23%) cows in Group 1. Clinical signs were variable and included dermatitis and pruritus of the head and neck, petechial haemorrhages, pyrexia, weight loss, thrombocytopenia, neutropenia, and regenerative anaemia.

PATHOLOGICAL FINDINGS: Gross findings included generalised lymphadenopathy, subcutaneous oedema, petechiation of mucosal and serosal surfaces, and gastrointestinal haemorrhage. Histologically, multiple organs and tissues contained inflammatory foci characterised by infiltrates of lymphocytes, plasma cells, macrophages and occasionally prominent multinucleated giant cells and eosinophils.

DIAGNOSIS: Multisystemic granulomatous and haemorrhagic syndrome resembling cell-mediated hypersensitivity, associated with DCD ingestion.

CLINICAL RELEVANCE:: This is the first report of toxicity in cattle associated with ingestion of DCD. The proportion of affected cows increased with increasing dose of DCD, but not all cattle in the high dose group developed disease, therefore additional factors may determine whether or not an individual cow will develop DCD-associated disease.     

Investigation of the index case herd and identification of the genotypes of Theileria orientalis associated with outbreaks of bovine anaemia in New Zealand in 2012

CASE HISTORY AND CLINICAL FINDINGS: On 7 September 2012 the Ministry for Primary Industries was notified of a dairy cow with regenerative anaemia (haematocrit (HCT) 0.08?L/L) in a herd of 465 Jersey-Friesian cross cows (index case herd) in the Northland region of New Zealand. Organisms consistent with Theileria spp. were present in red blood cells on a blood smear. No other causes of anaemia were detected following examination of affected cows. Blood samples collected from 29 randomly selected cows on 26 September 2012 showed that 24 (83%) were anaemic (HCT≤0.24 L/L) and therefore fitted the case definition for bovine anaemia associated with Theileria orientalis infection.

LABORATORY FINDINGS: Using a T. orientalis type-specific PCR assay that targeted the single subunit rRNA gene, all of six animals tested were positive for T. orientalis type Ikeda. Blood samples collected from clinically affected cattle in 11 subsequent outbreaks from throughout the North Island showed that T. orientalis Ikeda type was a common finding, but mixed infections with Chitose type were also identified. In addition, using a PCR assay that targeted the major piroplasm surface gene, T. orientalis type 5 was detected in one cow from the Waikato region.

DIAGNOSIS: The presence of T. orientalis type Ikeda, as well as type 5, was confirmed in cattle from outbreaks of bovine anaemia in herds throughout the North Island of New Zealand.

CLINICAL RELEVANCE: Two new types of T. orientalis were identified in this investigation, that were associated with a sudden rise in cases of bovine anaemia. The body of evidence showed that the Ikeda type was implicated as the cause of disease observed in this epidemic.     

Wintersweet (Chimonanthus praecox) toxicity in four goats

CASE HISTORY: A one-year-old female goat presented with acute onset of recumbency, seizures and vocalisation approximately 5 hours after being given access to branch trimmings from a neighbour's garden. The plant from which the pruned branches came was subsequently identified as wintersweet (Chimonanthus praecox). Three other goats kept in the same paddock displayed similar clinical signs over a period of 4 hours following the initial presentation.

CLINICAL FINDINGS: All four goats were ataxic, displayed tetanic seizures and were in lateral recumbency; they had dilated pupils and were hyperaesthetic, with elevated heart and respiratory rates. After symptomatic treatment, including sedation with diazepam, one of the three goats continued to deteriorate and was subjected to euthanasia. The remaining three goats recovered over 1–14 days with nursing care and physiotherapy.

DIAGNOSIS: Toxicity due to ingestion of wintersweet, which contains the alkaloid calycanthine.

CLINICAL RELEVANCE: Calycanthine is a central nervous system toxin, causing convulsions. Wintersweet shrubs are present in many New Zealand gardens. Practitioners should be aware that the seeds and flowers, and possibly the leaves, of this plant are highly toxic with signs of toxicity including ataxia, hyperaesthesia and seizures.     

Incidence and risk factors of heat-related illness in dogs from New South Wales,Australia (1997–2017)

Heat Related Illness (HRI) in dogs is expected to increase as heatwaves surge due to global warming. The most severe form of HRI, heat stroke, is potentially fatal in dogs. The current study investigated the incidence and risk factors for HRI in dogs in NSW, Australia, from 1997 to 2017. We identified 119 HRI cases during this period, with a fatality rate of 23%. Dog breeds at elevated risk of HRI were Australian Stumpy Tail Cattle Dog, British Bulldog, French Bulldog, Maremma Sheepdog, Italian Greyhound, Chow Chow, Airedale Terrier, Pug, Samoyed, English Springer Spaniel, Labrador Retriever, Golden Retriever, Cavalier King Charles Spaniel, Border Collie, Staffordshire Bull Terrier, and pooled non-Australian National Kennel Council breeds (which included the American and Australian Bulldog) when compared with cross breeds (i.e., the reference variable). As expected, HRI cases were more likely in December and January, during the Australian summer and during hotter years (e.g., 2016). There were no differences in the risk of HRI between males and females nor between desexed or un-desexed dogs; but older dogs were at increased risk of HRI. These findings underscore the need for data collection that will enable the incidence of HRI in dogs to be monitored and to better understand canine risk factors particularly as temperatures will continue to rise due to global warming. The risk of mortality from HRI underpins the need for education programs focussed on prevention and early identification of HRI so that owners present affected dogs to their veterinarian as promptly as possible.     

Bilateral oblique facial clefts,rudimentary eyes and hydrocephalus in an aborted equine foetus

Knowledge of congenital malformations and their causes in horses is generally sparse. Such conditions require more scientific attention to improve their diagnostics and inform prevention strategies. Here, a unique syndrome of bilateral oblique facial clefts (meloschisis), rudimentary eyes and hydrocephalus is reported in an equine foetus spontaneously aborted at gestation day 224. The cause of abortion was considered to be intrauterine death caused by umbilical cord torsions and subsequent compromised blood flow, but the aetiology of the malformation could not be determined. A detailed history, which includes exposure to a range of pharmaceutical compounds during the early stages of pregnancy, is provided and emphasizes the need for accurate recording of treatments in pregnant animals.