Development of maternal and foetal immune responses in cattle following experimental challenge with Neospora caninum at day 210 of gestation

This study examined the immunological responses of pregnant cattle and their foetuses following an experimental challenge with live Neospora caninum tachyzoites at day 210 of gestation. Animals were bled prior to and weekly throughout the experiment and sacrificed at 14, 28, 42 and 56 days post inoculation (dpi). At post mortem examination, samples of lymph nodes and spleen were collected from both dam and foetus for immunological analysis. Subcutaneous (sc) inoculation over the left prefemoral (LPF) lymph node of pregnant cattle at day 210 of gestation, led to the vertical transmission of parasites by 14 dpi, however no foetal deaths were observed in the infected animals. Foetuses from infected dams mounted Neospora-specific humoral and cell-mediated immune (CMI) responses by 14 dpi. These responses involved anti-Neospora IgG, antigen-specific lymphocyte proliferation, and the production of the cytokines IFN–γ, interleukin (IL)-4 and IL-10. There was also evidence of innate immunity during the response against Neospora from infected dams, with statistically significant (p < 0.05) increases in mean expression of toll like receptors (TLR)-2 on 56 dpi in maternal spleen, LPF, right prefemoral (RPF), left uterine (LUL) and right uterine (RUL) lymph nodes and TLR-9 in retropharyngeal (RLN), LPF and RPF lymph nodes from 28 dpi. Statistically significant (p < 0.05) increases in mean TLR-9 were detected in spleen samples from foetuses of infected dams, compared to the foetuses from control animals. Our results show that vertical transmission of the parasite occurred in all infected dams, with their foetuses showing effective Neospora-specific cell mediated, humoral and innate immune responses.     

Experimental infection with equine herpesvirus type 1 (EHV-1) induces chorioretinal lesions

Equine herpesvirus myeloencephalitis (EHM) remains one of the most devastating manifestations of equine herpesvirus type 1 (EHV-1) infection but our understanding of its pathogenesis remains rudimentary, partly because of a lack of adequate experimental models. EHV-1 infection of the ocular vasculature may offer an alternative model as EHV-1-induced chorioretinopathy appears to occur in a significant number of horses, and the pathogenesis of EHM and ocular EHV-1 may be similar. To investigate the potential of ocular EHV-1 as a model for EHM, and to determine the frequency of ocular EHV-1, our goal was to study: (1) Dissemination of virus following acute infection, (2) Development and frequency of ocular lesions following infection, and (3) Utility of a GFP-expressing virus for localization of the virus in vivo. Viral antigen could be detected following acute infection in ocular tissues and the central nervous system (experiment 1). Furthermore, EHV-1 infection resulted in multifocal choroidal lesions in 90% (experiment 2) and 50% (experiment 3) of experimentally infected horses, however ocular lesions did not appear in vivo until between 3 weeks and 3 months post-infection. Taken together, the timing of the appearance of lesions and their ophthalmoscopic features suggest that their pathogenesis may involve ischemic injury to the chorioretina following viremic delivery of virus to the eye, mirroring the vascular events that result in EHM. In summary, we show that the frequency of ocular EHV-1 is 50-90% following experimental infection making this model attractive for testing future vaccines or therapeutics in an immunologically relevant age group.     

Phenotypic characterisation of the cellular immune infiltrate in placentas of cattle following experimental inoculation with Neospora caninum in late gestation

Despite Neospora caninum being a major cause of bovine abortion worldwide, its pathogenesis is not completely understood. Neospora infection stimulates host cell-mediated immune responses, which may be responsible for the placental damage leading to abortion. The aim of the current study was to characterize the placental immune response following an experimental inoculation of pregnant cattle with N. caninum tachyzoites at day 210 of gestation. Cows were culled at 14, 28, 42 and 56 days post inoculation (dpi). Placentomes were examined by immunohistochemistry using antibodies against macrophages, T-cell subsets (CD4, CD8 and γδ), NK cells and B cells. Macrophages were detected mainly at 14 days post inoculation. Inflammation was generally mild and mainly characterized by CD3⁺, CD4⁺ and γδ T-cells; whereas CD8⁺ and NK cells were less numerous. The immune cell repertoire observed in this study was similar to those seen in pregnant cattle challenged with N. caninum at early gestation. However, cellular infiltrates were less severe than those seen during first trimester Neospora infections. This may explain the milder clinical outcome observed when animals are infected late in gestation.     

Prevalence of and risk factors for isolation of meticillin‐resistant Staphylococcus spp. from dogs with pyoderma in northern California,USA

Background – Canine pyodermas associated with meticillin‐resistant Staphylococcus spp. (MRS) have increased in prevalence over the past decade. Hypothesis/Objectives – To compare the prevalence of MRS isolation from dogs with superficial pyoderma at a primary care clinic (PCC) and those at a tertiary care facility (VMTH) in California, USA, and identify associated risk factors. Animals – Client‐owned dogs from the VMTH (80 dogs) and the PCC (30 dogs). Methods – Aerobic bacterial culture and antibiotic susceptibility were performed on swab specimens collected from dogs, and meticillin resistance was determined using microdilution methods according to Clinical and Laboratory Standards Institute guidelines. A mecA gene PCR assay was used to confirm meticillin resistance when possible. Results – Of 89 staphylococcal isolates from the VMTH, 34 (38.2%) were meticillin resistant. In 31 dogs, pyoderma persisted, and one or more follow‐up isolates were obtained. The species isolated and drug susceptibility changed unpredictably during treatment. Of 33 PCC isolates, nine (27.3%) were meticillin resistant. Multiple drug resistance was identified in 41 of 53 (77.3%) MRS isolates from the VMTH and five of nine from the PCC. The sensitivity and specificity of PCR for the detection of meticillin resistance was 34 of 39 (87%) and 86 of 87 (99%), respectively. Risk factors for meticillin resistance for both sites were antibiotic treatment within the last year (P = 0.001), and for VMTH, hospitalization of dogs within the last year (P = 0.001). Conclusions and clinical importance – The prevalence of meticillin resistance was not different between VMTH and PCC isolates (P = 0.29). Previous antimicrobial therapy was an important risk factor for the isolation of MRS at both sites.     

Structural Characterization of New Microcystins Containing Tryptophan and Oxidized Tryptophan Residues

Microcystins are cyclic peptides produced by cyanobacteria, which can be harmful to humans and animals when ingested. Eight of the (more than) 90 microcystin variants presently characterized, contain the amino acid tryptophan. The well-researched oxidation products of tryptophan; kynurenine, oxindolylalanine, and N-formylkynurenine, have been previously identified in intact polypeptides but microcystin congeners containing oxidized tryptophan moieties have not been reported. Liquid chromatography-tandem mass spectrometric analysis of an extract of Microcystis CAWBG11 led to the tentative identification of two new tryptophan-containing microcystins (MC‑WAba and MC-WL), as well as eight other microcystin analogs containing kynurenine, oxindolylalanine and N‑formylkynurenine (Nfk). Investigation of one of these congeners (MC‑NfkA) by nuclear magnetic resonance spectroscopy was used to verify the presence of Nfk in the microcystin. Liquid chromatography-mass spectrometry analysis of a tryptophan oxidation experiment demonstrated that tryptophan-containing microcystins could be converted into oxidized tryptophan analogs and that low levels of oxidized tryptophan congeners were present intracellularly in CAWBG11. MC-NfkR and MC-LNfk were detected in standards of MC-WR and MC-LW, indicating that care during storage of tryptophan-containing microcystins is required.     

WHOLE‐BODY BIODISTRIBUTION OF 3′‐DEOXY‐3′‐[18F]FLUOROTHYMIDINE (18FLT) IN HEALTHY ADULT CATS

Positron emission tomography/computed tomography (PET/CT) utilizing 3′‐deoxy‐3′‐[¹⁸F]fluorothymidine (¹⁸FLT), a proliferation tracer, has been found to be a useful tool for characterizing neoplastic diseases and bone marrow function in humans. As PET and PET/CT imaging become increasingly available in veterinary medicine, knowledge of radiopharmaceutical biodistribution in veterinary species is needed for lesion interpretation in the clinical setting. The purpose of this study was to describe the normal biodistribution of ¹⁸FLT in adult domestic cats. Imaging of six healthy young adult castrated male cats was performed using a commercially available PET/CT scanner consisting of a 64‐slice helical CT scanner with an integrated whole‐body, high‐resolution lutetium oxy‐orthosilicate (LSO) PET scanner. Cats were sedated and injected intravenously with 108.60 ± 2.09 (mean ± SD) MBq of ¹⁸FLT (greater than 99% radiochemical purity by high‐performance liquid chromatography). Imaging was performed in sternal recumbency under general anesthesia. Static images utilizing multiple bed positions were acquired 80.83 ± 7.52 (mean ± SD) minutes post‐injection. Regions of interest were manually drawn over major parenchymal organs and selected areas of bone marrow and increased tracer uptake. Standardized uptake values were calculated. Notable areas of uptake included hematopoietic bone marrow, intestinal tract, and the urinary and hepatobiliary systems. No appreciable uptake was observed within brain, lung, myocardium, spleen, or skeletal muscle. Findings from this study can be used as baseline data for future studies of diseases in cats.