Chlamydiae and atherosclerosis: can psittacine cases support the link?

Atherosclerosis is a common disease in pet birds, particularly in psittacines. Little is known about the role of risk factors predisposing birds to this disease. In our study, we tried to detect chlamydiae in formalin-fixed and paraffin-embedded atherosclerotic tissue from 103 pet birds to clarify their role in atherosclerosis. Methods used were polymerase chain reaction (PCR), sequencing, and immunohistochemistry. Histopathologic examination served to classify the extent of atherosclerotic lesions. In the PCR, 4 (3.9%) of 103 cases, all of them with advanced stages of atherosclerosis, were positive. Subsequent sequence analysis revealed high identities (94%-100%) with Chlamydophila psittaci in three cases. Interestingly, two of these birds came from C. psittaci-infected populations. Because of the low incidence (3.9%), the occurrence only in advanced stages, and the association with C psittaci-infected avian populations, a causal relationship between chlamydiae and atherosclerosis in pet birds is rather improbable.     

Detection of Mycobacterium avium subsp. paratuberculosis in ovine faeces by direct quantitative PCR has similar or greater sensitivity compared to radiometric culture

The aims of this study were to develop a new real-time quantitative PCR (QPCR) assay based on IS900 for detection and quantification of Mycobacterium avium subsp. paratuberculosis (MAP) DNA in faeces, and to use this to detect infected sheep. Both the C and S strains of MAP were detected by the QPCR assay, and no cross reactions were detected with 51 other species of mycobacteria including 10 which contained IS900-like sequences. One copy of IS900 fragment cloned into plasmid pCR2.1 and 1 fg of MAP genomic DNA were consistently detected, while in spiked faecal samples the detection limit was 10 viable MAP per gram of ovine faeces. A total of 506 individual ovine faecal samples and 27 pooled ovine faecal samples with known culture results were tested. The QPCR assay detected 68 of 69 BACTEC culture positive individual faeces and there was a strong relation between time to detection in culture and DNA quantity measured by QPCR (r= -0.70). In pooled faecal samples, QPCR also agreed with culture (kappa=0.59). MAP DNA was detected from some culture negative faecal samples from sheep exposed to MAP, suggesting that the QPCR has very high analytical sensitivity for MAP in faecal samples and detects non-viable MAP in ovine faeces. None of the faecal samples from 176 sheep that were not exposed to MAP were positive in QPCR. This is the first report of a direct faecal QPCR assay that has similar sensitivity to a gold standard radiometric culture assay.     

Sampling and repeatability of radiometric faecal culture in bovine Johne's disease

The repeatability of detection of Mycobacterium avium subsp. paratuberculosis (Map) within and between samplings from 16 paratuberculous dairy cows (13 subclinical; 3 clinical) was investigated by radiometric culture of quadrants of faecal dung pats collected on four to seven occasions over a 10-16-day period. Results were compared to serological status and to pathological and bacteriological findings in multiple tissues obtained at slaughter from 15 of the animals 2-6 weeks after the faecal samplings. From faecal samples taken on 77 occasions over the 2-week period, 296/308 (96%) quadrants were culture positive, with samples from all cattle showing evidence of faecal shedding of Map. Histological lesions typical of paratuberculosis were present in 14 of the 15 cows examined at slaughter, varying in severity from mild (two animals) to moderate (4) and advanced (8), and all predilection tissue sites yielded Map. The negative faecal samples were derived from a single animal that was culture positive in two quadrants on each of the first two (of four) sampling occasions (i.e. culture positive in only 4 of 16 collected quadrants). This animal was found to be histologically negative at slaughter, and culture positive from three of five predilection tissue sites. Faecal samples from cows with subclinical and clinical paratuberculosis, with lesion severity ranging from mild to severe at multiple predilection sites, produced faeces with relatively consistent concentrations of Map within samples. There was significant variation in concentrations of Map between samples in individual animals over a period of 2 weeks, but this did not affect the dichotomous positive-negative culture status for 15 of the 16 cattle. A faecal sample collected non-randomly per rectum thus provides a representative specimen for detection of Map by radiometric culture on a single sampling occasion.     

Relative adrenal insufficiency in dogs with sepsis

BACKGROUND: A syndrome of relative adrenal insufficiency has been identified in septic humans, and is associated with hypotension and death. Relative adrenal insufficiency is generally associated with basal serum cortisol concentration within or above the reference range and a blunted cortisol response to adrenocorticotropic hormone administration. It is unknown whether relative adrenal insufficiency occurs in septic dogs. HYPOTHESIS: That relative adrenal insufficiency occurs in septic dogs, and that relative adrenal insufficiency is associated with hypotension and mortality. ANIMALS: Thirty-three septic dogs admitted to a small animal intensive care unit. METHODS: Dogs were included in the study if they had a known or suspected infectious disease and had systemic inflammatory response syndrome. Dogs were excluded if they had disease or medication history expected to affect the hypothalamic-pituitary-adrenal axis. Serum cortisol and endogenous plasma adrenocorticotropic hormone concentrations were measured before, and serum cortisol concentration measured 1 hour after, intramuscular administration of 250 microg of cosyntropin/dog. The change in cortisol concentration (delta-cortisol) before and after cosyntropin administration was determined in each dog. RESULTS: Hypotension was associated with lower delta-cortisol values (OR 1.3; CI 1.0-1.9; P = .029). delta-Cortisol cutoff of 3.0 microg/dL was most accurate for predicting hypotension, survival to discharge, and 28-day survival. The rate of death in dogs with delta-cortisol < or = 3 microg/dL was 4.1 times that of dogs with delta-cortisol > 3 microg/dL (RR 4.1; CI 1.5-12.3; P = .01). CONCLUSIONS AND CLINICAL RELEVANCE: Delta-cortisol < or = 3 microg/dL after adrenocorticotropic hormone administration is associated with systemic hypotension and decreased survival in septic dogs.     

Ultrasound-guided core needle biopsy in dogs with thyroid carcinoma

Currently, a histological diagnosis of highly vascularized canine (c) thyroid carcinoma (TC) is primarily obtained following excisional biopsy (EB) through thyroidectomy. Non-EBs are contraindicated in unresectable invasive cTCs due to their highly vascularized nature, which subsequently, lack histological diagnosis. We hypothesised ultrasound-guided core needle biopsy (UGCNB) to be a safe biopsy technique to obtain an accurate histological diagnosis in unresectable TCs. Nine client-owned dogs with suspected naturally occurring TC, presented for surgical excision, were included. First, a UGCNB was taken from the cervical tumour, followed by EB. Haemorrhage following UGCNB was evaluated preoperatively and once the tumour was surgically exposed by visual inspection and ultrasonography. Histological analysis, including cell organisation, tumour capsular and vascular invasion, and immunohistochemistry were performed and compared between both biopsy specimens (i.e., UGCNB and EB) of the same dog. Pre- and peroperative visual inspection revealed minor, localised haemorrhage, subsequent to the UGCNB, in 7/9 dogs. Histology of the EBs confirmed TC in 8/9 dogs and was inconclusive in 1/9 dogs. Histology of the UGCNBs revealed neoplastic thyroid tissue in 7/9 UGCNBs and was inconclusive in 1/9 UGCNBs. The remaining UGCNB contained no mass related tissue and was, therefore, excluded. Histological parameters (i.e., cell organisation, tumour capsular and vascular invasion) were not concordant between 6/8 included UGCNBs and their respective EB. Immunolabelling for thyroglobulin and calcitonin was concordant between all eight included UGCNBs and their respective EB. The remaining evaluated immunohistochemical markers (i.e., cyclooxygenase-2 [COX-2], P-glycoprotein and vascular endothelial growth factor [VEGF]) were concordant between the included UGCNBs and the EBs in 6/8 dogs. To conclude, UGCNBs can be safely obtained in suspected cTCs and enable a reliable diagnosis of the thyroid origin, thyroid cell origin and potential therapeutic markers such as COX-2, P-glycoprotein and VEGF. Subsequently, UGCNB enables clinicians to establish an individually tailored treatment plan in dogs with unresectable TC.     

Dihydrooxadiazines: Octopaminergic system as a potential site of insecticidal action

Dihydrooxadiazines are structural analogs of octopamine and were compared with octopamine for their ability to compete with [³H]dihydroergocryptine ([³H]DHE) for binding sites on DHE-sensitive receptors, to stimulate adenylate cyclase activity in nervous system homogenates of Periplaneta americana L., and to modulate the action of the peptide proctolin on the oviducal muscles of Locusta migratoria L. [³H]DHE binding was inhibited by low concentrations (μM range) of octopamine, phentolamine, N-demethylchlordimeform (DCDM) and several dihydrooxadiazines. The tested dihydrooxadiazines acted as aminergic agonists in stimulating cyclic AMP production in cockroach nervous system homogenates and did not show additive effects with octopamine, whereas additivity was observed with 5-hydroxytryptamine. The relative potency of octopaminergic antagonists, including mianserin, cyproheptadine, phentolamine, and gramine, to block octopamine-mediated elevation of cyclic AMP production was similar to the rank-order potency of the same antagonists to inhibit dihydrooxadiazine-mediated elevation of cyclic AMP production. Octopamine, 2-(4-bromophenyl)-5,6-dihydro-4H-1,3,4-oxadiazine (4-Br-PDHO), and 8-Br-cyclic AMP caused increased phosphorylation of proteins that are phosphorylated by exogenously added cyclic AMP-dependent protein kinase. These results indicate that the dihydrooxadiazine-induced rise in cyclic AMP levels in homogenates of the cockroach nervous system results directly in activation of an endogenous cyclic AMP-dependent protein kinase. 4-Br-PDHO behaved similarly to octopamine in modulating the action of proctolin-induced contractions in locust oviducal muscles. These observations suggest that dihydrooxadiazines act as octopamine agonists and have an octopaminergic action in modulating the action of proctolin. Thus, it is proposed that dihydrooxadiazines exert at least part of their insecticidal and miticidal actions through interaction with the octopaminergic system.     

1-Octanol/Water Partition Coefficient (Kow) and pKa for Ionisable Pesticides Measured by apH-Metric Method

pK_a values for a wide range of commonly used ionisable pesticides, together with the log K_ow values of the most lipophilic form of each, have been measured using pH-metric techniques. Examples of acids, bases and multiprotic compounds from the major classes of herbicides, and a number of insecticides and fungicides that contain ionisable groups, are included. The pK_a and log K_ow values so obtained were generally in good agreement with values taken from the literature that were measured by other methods. The lower limit of log K_ow that could be measured by the pH-metric method lay below the -0·97 obtained for amitrole, but the method could not be applied to glyphosate for which shake-flask measurements indicated log K_ow below -3. The highest log K_ow obtained in this study was 5·12 for pentachlorophenol. The pH-metric technique offers a rapid and convenient method to determine pK_a and log K_ow for ionisable compounds, especially when utilising an automatic titration system linked to a dedicated computer.