Establishment of proliferative cell nuclear antigen gene as an internal reference gene for polymerase chain reaction of a wide range of archival and fresh mammalian tissues.

Polymerase chain reaction (PCR) from paraffin-embedded tissues provides a powerful tool to amplify DNA from a variety of recent and archival material. Because DNA from paraffin-embedded samples is more degraded than from fresh material, the amplification of reference genes is essential to exclude false-negative results. This study describes the use of the proliferative cell nuclear antigen (PCNA) gene as a reference gene in a range of animal species and in humans. The PCNA-PCR to amplify a fragment extending from exon 5 through exon 6 and including the intervening intron 6 gave a reproducible pattern, with a 280-base pair (bp) band from canine, equine, bovine, ovine, and caprine samples showing high sequence homology. Porcine, guinea pig, tiger, and lion samples, however, gave an additional fragment of approximately 197 bp. The whole intron 6 from these fragments is missing, possibly representing a pseudogene. In feline samples only the 197-bp fragment could be detected. This study shows that the PCNA gene is highly conserved across a broad range of animal species and is well suited as an internal control for PCR analysis in veterinary medicine.     

Chlamydiales in guinea-pigs and their zoonotic potential

The aim was to detect and characterize chlamydial infections in guinea-pigs (GP) with ocular disease, study their pathogenicity and zoonotic potential and to test for the presence of Acanthamoebae spp. in GP eyes and to investigate whether they could act as vectors for Chlamydia-like organisms. Overall 126 GP, of which 77 were symptomatic, were screened by clinical examination, cytology, gross pathology, histology, immunohistochemistry, polymerase chain reaction (PCR) and bacteriology. A new Chlamydiaceae-specific intergenic spacer rRNA gene PCR, designed to amplify this segment linking the 16S and 23S regions, was performed. DNA samples were also received from one owner including samples of his cat and rabbit. Guinea-pigs: 48 of 75 symptomatic, but only 11 of 48 asymptomatic GP were positive by PCR for Chlamydophila caviae guinea-pig inclusion conjunctivitis (GPIC) (P < 0.0001). Eighteen of 75 or 15/48, respectively, were positive for DNA from Chlamydia-like organisms. Acanthamoebae-DNA could be found in two GP, of which one was symptomatic. Owner, cat and rabbit: Samples of all three species were positive by PCR for C. caviae GPIC and the owner's one-day disposable contact lenses showed a positive PCR result for the Chlamydia-like organism Parachlamydia acanthamoebae. No Acanthamoebae-DNA could be detected. This study is the first to describe Chlamydia-like organisms in GP and to detect C. caviae GPIC in human, cat and rabbit. Therefore, C. caviae GPIC could pose a zoonotic potential. We believe that the finding of C. caviae GPIC in species other than GP is probably not unique.     

Cryptococcosis in Gilbert's and long-nosed potoroo

Two cases of fatal cryptococcosis are described, one of Cryptococcus neoformans infection in a Gilbert's potoroo (Potorous gilbertii) and one of Cryptococcus gattii infection in a long-nosed potoroo (Potorous tridactylus). The diagnoses were confirmed by culture and specific immunohistochemistry, respectively. The long-nosed potoroo tested positive using the latex cryptococcal antigen test (LCAT), whereas the Gilbert's potoroo had a negative LCAT result despite having advanced disease of some duration. In both cases, the clinical presentation was a progressive neurologic disease associated with a central nervous system infection. Pulmonary infection was also observed in the long-nosed potoroo. Specific treatment with antifungal agents was unsuccessful in the long-nosed potoroo.     

The relationship between digital perfusion pressure and hoof lamellar blood flow in isoflurane-anesthetized horses

Digital perfusion pressure (DPP) equals mean arterial pressure (MAP) at the hoof coronet minus digital interstitial pressure (DIP) within the hoof. To test whether lamellar blood flow (LBF) changes proportionately to DPP, anesthesia was induced and maintained with isoflurane in six horses to target a MAP of 60 mmHg. Arterial, venous, and hoof interstitial pressures were measured in each pelvic limb. LBF was measured using fluorescent microspheres during dobutamine infusions targeting either 60 (low), 80 (medium), or 100 (high) mmHg MAP. Following euthanasia, hoof lamina was collected for microsphere isolation. To reduce intra-individual variability, medium and high pressures and flows were divided by their respective low pressure and flow baseline values, yielding indexed variables of ΔLBF and ΔDPP. The ΔLBF correlated negatively with the ΔDPP. We conclude that LBF was not solely determined by passive pressure-flow relationships and that systemic hypertension may not effectively increase dermal LBF in horses.     

Effects of the fungal endophyte Acremonium coenophialum in fescue on pregnant mares and foal viability.

Effects of the endophyte Acremonium coenophialum in tall fescue on pregnant mares and foal viability were evaluated. Twenty-two mature pregnant mares were randomly chosen to graze either Kentucky-31 tall fescue that was free from A coenophialum (endophyte-free, EF) or tall fescue infected with A coenophialum (endophyte-present, EP) after the first 90 days of pregnancy through parturition. Concentrations of pyrrolizidine and ergopeptine alkaloids were significantly greater in EP grass, compared with EF pasture. Ten of 11 mares grazing EP pasture had obvious dystocia. Mean duration of gestation was significantly greater for the EP group, compared with the EF group. Foal survivability was severely reduced among mares grazing EP fescue with only 1 foal surviving the natal period. Udder development and lactation were low in mares grazing EP grass. The absence of clinical problems in mares grazing EF grass implicated the endophyte as the causative agent of reproductive problems and perinatal foal mortality in pregnant mares grazing endophyte-infected fescue grass. Caution should be exercised in allowing pregnant mares to graze pastures infected with the endophyte A coenophialum.     

Electroencephalographic alterations in a mare with presumed intermittent neuroglycopenia caused by severe hypoglycaemia-associated with nonislet cell tumour

The focus of this report was to document the electroencephalogram (EEG) findings associated with nonislet cell tumour-induced hypoglycaemia (NICTH) in a racing Thoroughbred mare subsequently diagnosed with renal carcinoma and carcinomatosis. We evaluated cortical cerebral activity and confirmed paroxysmal activity consistent with subclinical seizures by performing an EEG prior to onset of clinical signs of seizures. We showed that the electrical activity pattern progressed in a similar fashion as in humans with severe, progressive hypoglycaemia and presumed neuroglycopenia. The glycaemic threshold for abnormalities detected in EEG showed that abnormal electrical cortical activity appeared when blood glucose dropped to 44 mg/dL (2.4 mmol/L) and paroxysmal activity spread, becoming generalised when blood glucose dropped to 23 mg/dL (1.3 mmol/L). NICTH is an uncommon paraneoplastic phenomenon in several species. In horses, NICTH has been reported most frequently in primary renal tumours, followed by primary hepatic tumours. This report highlights the importance of investigating the possibility of hypoglycaemia in cases of horses that present altered state of consciousness with or without intermittent seizures. This case report also highlights that the diagnostic work-up for horses presenting with episodic hypoglycaemia should comprise investigation of possible neoplasia, as paraneoplastic hypoglycaemia should be included in the list of differentials of horses with hypoglycaemia.     

Investigations into the temporal development of epitheliocystis infections in brown trout: a histological study

Epitheliocystis in Swiss brown trout (Salmo trutta) is a chlamydial infection, mainly caused by Candidatus Piscichlamydia salmonis and Candidatus Clavichlamydia salmonicola. To gain a better understanding of the temporal development of infections in wild brown trout, we investigated epitheliocystis infections during the course of the summer and autumn months of a single year (2015), and compared this to sampling points over the span of the years 2012–2014. The survey focused on tributaries (Venoge and Boiron) of the Rhone flowing in to Lake Geneva. When evaluated histologically, epitheliocystis infections were found throughout the period of investigation with the exception of the month of June. Fifty to 86 animals per sampling were investigated. Highest prevalence and infection intensities were seen in September. A correlation between epitheliocystis infection and water temperatures was not evident. Interyear comparison revealed consistent levels of prevalence and infection intensities in late summer. The absence of infections in June, combined with the consistent interyear results, indicates seasonal fluctuation of epitheliocystis infections in brown trout with a reservoir persisting during winter months from which infections can re‐initiate each year. This could either be at levels below detection limits within the brown trout population itself or in an alternative host.     

Evaluation of twice-daily, low-dose trilostane treatment administered orally in dogs with naturally occurring hyperadrenocorticism

OBJECTIVE: To evaluate the effects of twice-daily oral administration of a low-dose of trilostane treatment and assess the duration of effects after once-daily trilostane administration in dogs with naturally occurring hyperadrenocorticism (NOH). DESIGN: Prospective study. ANIMALS: 28 dogs with NOH. PROCEDURES: 22 dogs received 0.5 to 2.5 mg of trilostane/kg (0.23 to 1.14 mg/lb) orally every 12 hours initially. At intervals, dogs were reevaluated; owner assessment of treatment response was recorded. To assess drug effect duration, 16 of the 22 dogs and 6 additional dogs underwent 2 ACTH stimulation tests 3 to 4 hours and 8 to 9 hours after once-daily trilostane administration. RESULTS: After 1 to 2 weeks, mean trilostane dosage was 1.4 mg/kg (0.64 mg/lb) every 12 hours (n = 22 dogs; good response [resolution of signs], 8; poor response, 14). Four to 8 weeks later, mean dosage was 1.8 mg/kg (0.82 mg/lb) every 12 or 8 hours (n = 21 and 1 dogs, respectively; good response, 15; poor response, 5; 2 dogs were ill). Eight to 16 weeks after the second reevaluation, remaining dogs had good responses (mean dosages, 1.9 mg/kg [0.86 mg/lb], q 12 h [n = 13 dogs] and 1.3 mg/kg [0.59 mg/lb], q 8 h [3]). At 3 to 4 hours and 8 to 9 hours after once-daily dosing, mean post-ACTH stimulation serum cortisol concentrations were 2.60 and 8.09 Pg/dL, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: In dogs with NOH, administration of trilostane at low doses every 12 hours was effective, although 2 dogs became ill during treatment. Drug effects diminished within 8 to 9 hours. Because of potential adverse effects, lower doses should be evaluated.