A 5-year-old, spayed female cat was referred because of a mass in the cranial mediastinum noted on thoracic radiographs. A thymoma was diagnosed following ultrasound and biopsy of the mass. Treatment was initiated with coarse-fraction radiation therapy using external-beam therapy (four fractions of 5 Gy). The mass responded, but granulocytopenia developed. Bone marrow examination showed a myeloid to erythroid ratio of approximately 1:1, with a left shift within the myeloid line. These findings, as well as the lack of toxic changes within the peripheral blood neutrophils, suggested immune-mediated destruction of peripheral granulocytes. Immune suppression with prednisone and cyclosporine was instituted. After 7 weeks, the neutrophil count returned to normal. The tumor was removed, and cyclosporine was reduced and eventually discontinued 3 weeks postsurgery.
Culturing many obligate intracellular bacteria is difficult or impossible. However, these organisms have numerous adaptations allowing for infection persistence and immune system evasion, making them some of the most interesting to study. Recent advancements in genome sequencing, pyrosequencing and Phi29 amplification, have allowed for examination of whole-genome sequences of intracellular bacteria without culture. We have applied both techniques to the model obligate intracellular pathogen Anaplasma marginale and the human pathogen Anaplasma phagocytophilum, in order to examine the ability of phi29 amplification to determine the sequence of genes allowing for immune system evasion and long-term persistence in the host. When compared to traditional pyrosequencing, phi29-mediated genome amplification had similar genome coverage, with no additional gaps in coverage. Additionally, all msp2 functional pseudogenes from two strains of A. marginale were detected and extracted from the phi29-amplified genomes, highlighting its utility in determining the full complement of genes involved in immune evasion.
A male purebred Barbados Blackbelly, twin to another male, was born with obvious external congenital abnormalities. Male twinwas normal. Abnormal lamb had a short torso and spinal column. At 1 week, the lamb was presented with dyspnea and lethargy,and euthanasia was elected. Lamb was submitted for full necropsy and for additional diagnostics. After postmortem, in additionto histopathology, radiographs and computed tomographs were obtained. Lamb had congenital vertebral and occipital malformations,meningitis, hypospadias, and renal malformation. To authors’ knowledge, hypospadias and vertebral malformation inthe Barbados Blackbelly breed has not been reported. Apparently, Barbados Blackbelly breed harbors closely related genetics, thusvigilant monitoring for genetic malformations is suggested.
Evaluation of the effect of formalin fixation on skin specimens in dogs and catsSkin and subcutaneous tissues are the origin of most common neoplasms affecting dogs, accounting for approximately one third of all tumors encountered in the species.Surgical excision is frequently the best chance for a cure; determining factors influencing the success of excision are vital for surgical management of cases.This work examined the shrinkage of skin of various lengths from three sites in formalin for both dogs and cats.Tissues were measured on the animal (initial measurement), at the time of excision (post-removal), and after formalin fixation (post-fixation).While shrinkage after tissue removal was found in samples from the thorax, abdomen, and rear leg in dogs and from the rear leg in cats, no significant shrinkage due to formalin fixation was detected in any sample except for the thoracic samples from the dog.Therefore, when determining where to make incisions to effect a surgical cure, initial measurements should take into account tissue shrinkage effects.
Abstract We recently described the incidence of a SCID disease in a litter of Jack Russell terriers. In this study, we show that the molecular defect in these animals is faulty V(D)J recombination. Furthermore, we document a complete deficit in DNA-dependent protein kinase activity that can be explained by a marked diminution in the expression of the catalytic subunit DNA-dependent protein kinase catalytic subunit (DNA-PKcs). We conclude that as is the case in C.B-17 SCID mice and in Arabian SCID foals, the defective factor in these SCID puppies is DNA-PKcs. In mice, it has been clearly established that DNA-PKcs deficiency produces an incomplete block in V(D)J recombination, resulting in “leaky” coding joint formation and only a modest defect in signal end ligation. In contrast, DNA-PKcs deficiency in horses profoundly blocks both coding and signal end joining. Here, we show that although DNA-PKcs deficiency in canine lymphocytes results in a block in both coding and signal end joining, the deficit in both is intermediate between that seen in SCID mice and SCID foals. These data demonstrate significant species variation in the absolute necessity for DNA-PKcs during V(D)J recombination. Furthermore, the severity of the V(D)J recombination deficits in these three examples of genetic DNA-PKcs deficiency inversely correlates with the relative DNA-PK enzymatic activity expressed in normal fibroblasts derived from these three species.
Histopathology tissue archives can be an important source of specimens for retrospective studies, as these include samples covering a large number of diseases. In veterinary medicine, archives also contain samples from a large variety of species and may represent naturally-occurring models of human disease. The formalin-fixed, paraffin-embedded (FFPE) tissues comprising these archives are rich resources for retrospective molecular biology studies and pilot studies for biomarkers, as evidenced by a number of recent publications highlighting FFPE tissues as a resource for analysis of specific diseases. However, DNA extracted from FFPE specimens are modified and fragmented, making utilization challenging. The current study examines the utility of FFPE tissue samples from a veterinary diagnostic laboratory archive in five year intervals from 1977 to 2013, with 2015 as a control year, to determine how standard processing and storage conditions has affected their utility for future studies. There was a significant difference in our ability to obtain large amplicons from samples from 2015 than from the remaining years, as well as an inverse correlation between the age of the samples and product size obtainable. However, usable DNA samples were obtained in at least some of the samples from all years tested, despite variable storage, fixation, and processing conditions. This study will help make veterinary diagnostic laboratory archives more useful in future studies of human and veterinary disease.
Performing a mitosis count (MC) is the diagnostic task of histologically grading canine Soft Tissue Sarcoma (cSTS). However, mitosis count is subject to inter- and intra-observer variability. Deep learning models can offer a standardisation in the process of MC used to histologically grade canine Soft Tissue Sarcomas. Subsequently, the focus of this study was mitosis detection in canine Perivascular Wall Tumours (cPWTs). Generating mitosis annotations is a long and arduous process open to inter-observer variability. Therefore, by keeping pathologists in the loop, a two-step annotation process was performed where a pre-trained Faster R-CNN model was trained on initial annotations provided by veterinary pathologists. The pathologists reviewed the output false positive mitosis candidates and determined whether these were overlooked candidates, thus updating the dataset. Faster R-CNN was then trained on this updated dataset. An optimal decision threshold was applied to maximise the F1-score predetermined using the validation set and produced our best F1-score of 0.75, which is competitive with the state of the art in the canine mitosis domain.
Histopathology tissue archives can be an important source of specimens for retrospective studies, as these include samples covering a large number of diseases.In veterinary medicine, archives also contain samples from a large variety of species and may represent naturally-occurring models of human disease.The formalin fixed, paraffin-embedded (FFPE) tissues comprising these archives are rich resources for retrospective molecular biology studies and pilot studies for biomarkers, as evidenced by a number of recent publications highlighting FFPE tissues as a resource for analysis of specific diseases.However, DNA extracted from FFPE specimens are modified and fragmented, making utilization challenging.The current study examines the utility of FFPE tissue samples from a veterinary diagnostic laboratory archive in five year intervals from 1977-2013, with 2015 as a control year, to determine how standard processing and storage conditions has affected their utility for future studies.There was a significant difference in our ability to obtain large amplicons from samples from 2015 than from the remaining years, as well as an inverse correlation between the age of the samples and product size obtainable.However, usable DNA samples were obtained in at least some of the samples from all years tested, despite variable storage, fixation, and processing conditions.This study will help make veterinary diagnostic laboratory archives more useful in future studies of human and veterinary disease.
Summary A one‐year‐old, Thoroughbred colt presented for evaluation due to a one month history of fever of unknown origin and progressive weight loss. On initial presentation, the horse was febrile and showed signs localised to the respiratory tract. These included bilaterally increased bronchovesicular sounds and a moderate, diffuse interstitial pattern on thoracic radiographs. A transtracheal wash yielded mucopurulent debris, culture of which grew small numbers of Staphylococcus epidermidis and Aspergillus spp. The horse was discharged with a diagnosis of bronchointerstitial pneumonia and placed on antibiotic therapy. Ten days after initial presentation, he developed abdominal pain that was unresponsive to on‐farm treatment. The horse was febrile, displayed increased respiratory rate and effort, and showed moderate signs of abdominal pain. On rectal examination, a firm, 8–10 cm mass was palpated on midline. The colt was admitted to the hospital and scheduled for exploratory laparotomy, but died a short time later before surgery could be performed. This report describes the clinical, diagnostic and histopathological findings of a case of alimentary lymphoma in a yearling colt.
The definitive diagnosis of canine soft-tissue sarcomas (STSs) is based on histological assessment of formalin-fixed tissues. Assessment of parameters, such as degree of differentiation, necrosis score and mitotic score, give rise to a final tumour grade, which is important in determining prognosis and subsequent treatment modalities. However, grading discrepancies are reported to occur in human and canine STSs, which can result in complications regarding treatment plans. The introduction of digital pathology has the potential to help improve STS grading via automated determination of the presence and extent of necrosis. The detected necrotic regions can be factored in the grading scheme or excluded before analysing the remaining tissue. Here we describe a method to detect tumour necrosis in histopathological whole-slide images (WSIs) of STSs using machine learning. Annotated areas of necrosis were extracted from WSIs and the patches containing necrotic tissue fed into a pre-trained DenseNet161 convolutional neural network (CNN) for training, testing and validation. The proposed CNN architecture reported favourable results, with an overall validation accuracy of 92.7% for necrosis detection which represents the number of correctly classified data instances over the total number of data instances. The proposed method, when vigorously validated represents a promising tool to assist pathologists in evaluating necrosis in canine STS tumours, by increasing efficiency, accuracy and reducing inter-rater variation.
