Veterinary Pathology

Soft tissue sarcomas (STSs) are conventionally viewed as poorly immunogenic tumors; however, some human STSs have recently been reported to elicit an immune response, thus representing potential candidates for immunotherapy. Data regarding immune cell infiltrates in canine STSs are limited and reported without tumor-type stratification. The aim of this study was to retrospectively assess tumor-infiltrating lymphocytes (TILs) in canine STSs of 5 different histotypes. Eighty-seven canine STSs were collected: 22 perivascular wall tumors (PWTs), 19 liposarcomas, 17 fibrosarcomas, 16 myxosarcomas, and 13 leiomyosarcomas. The tumors were graded and immunolabeled for CD3, CD20, and FoxP3, and slides were scanned. T-cell, B-cell, Treg, and total TIL densities were quantified with QuPath software and expressed as cells/mm². The B/T-cells ratio and Treg/T-cell proportions were calculated. Total TIL densities were higher in PWTs and myxosarcomas (median = 225 and 303, respectively). PWTs had higher T-cell density but lower Treg proportion (median = 152 and 7.6% respectively). Myxosarcomas had higher Treg densities and B/T-cell ratios (median = 24.4 and 1.57, respectively). No association with grade was found among STSs as a group. In myxosarcomas, higher grade was significantly associated with higher total TILs, and CD20+ and FoxP3+ cell densities (p < .05). The results suggest that PWTs and myxosarcomas may represent the most immunogenic STS types. Myxosarcomas elicit a B-cell and Treg-rich immune response; PWTs stimulate a T-cell-rich and Treg-poor reaction. The immune system response may contribute to the more aggressive behavior of myxosarcomas and the more indolent course of PWTs.

Classification schemes regarding canine subcutaneous mast cell tumors (csMCTs) remain elusive, lack consensus, and are prone to interobserver variability and bias. This observational study aimed to assess the reproducibility and the prognostic significance of volume-weighted mean nuclear volume ( v v ¯ ), a stereological estimation offering insights into nuclear size and its variability, in csMCTs. Thirty csMCTs were selected with information regarding outcome, and v v ¯ was estimated using the “point-sampled intercept” method. Interobserver and intraobserver v v ¯ reproducibility yielded concordance coefficients near or above 0.90. Regarding previously reported risk factors (pattern, mitotic count, and multinucleated cells), no statistically significant differences were identified between patterns and clinical outcome, nor between patterns and v v ¯ ; however, the infiltrative pattern was represented more in the poorer outcome group and had higher v v ¯ values. When comparing v v ¯ and clinical outcome, a statistically significant difference emerged. Cases with poorer outcomes had higher v v ¯ values ( x ~ = 192.9) than cases with more favorable outcomes ( x ~ = 120.5), and this association was statistically significant on both univariable and multivariable analyses. This study suggests that v v ¯ is highly reproducible and is associated with clinical outcome in csMCTs.

Dermoid cysts are developmental abnormalities usually located at the dorsal body midline. Histologically, these lesions reduplicate the skin, with associated collagen bundles, adnexal structures, and are filled by keratin and hairs. While these cysts have well-recognized breed and anatomical predispositions in dogs, the information in feline patients is restricted to scattered case reports and anecdotal data. Through a multi-institutional retrospective case series, we aim to describe the clinical and demographic features of this condition in cats. We retrieved a total of 220 cases. The average age at the time of diagnosis was 5.5 years, with 59.5% (131/220) being males. Domestic short hairs were the most represented breed, accounting for 56.4% (124/220) of the cases. The average cyst diameter was 1.4 cm, and 99.5% (219/220) of the cases the cysts were cutaneous and subcutaneous with the most common anatomical location being the neck (55.9%; 123/220). One dermoid cyst was in the mesentery. In most cases, no associated inflammatory lesions were reported (66.4%; 146/220). The anatomical location of the dermoid cyst did not differ significantly among sexes (Chi-square test, P = .840), breeds (Chi-square test, P = .999), ages (Chi-square test, P = .627), or other histological findings related to the cyst (Chi-square test, P = .363).

Synovial lipomatosis is an uncommon, intra-articular, fat-containing, proliferative lesion with unknown etiology that is rarely reported in dogs. A retrospective study spanning 13 years was conducted to search for cases of canine synovial lipomatosis. Among 188 synovial biopsies of major diarthrodial joints (ie, shoulder, elbow, carpus, hip, stifle, and tarsus) from 186 dogs, 4 cases (2.1%) of synovial lipomatosis were identified. One case occurred in a stifle with chronic lateral patellar luxation. The other 3 cases had microscopic evidence of synovitis (eg, synovial hyperplasia, lymphoplasmacytic infiltrates, hemosiderin-laden macrophages, myxomatous changes, fibrosis, and increased vascularity) but lacked a clinical history of arthritis. Immunohistochemistry for HMGA2 was negative, suggesting canine synovial lipomatosis is a non-neoplastic proliferative lesion, yet the cause remains unknown.

The factors that determine the appearance of the different pathologic forms associated with bovine paratuberculosis are not fully understood, but new research suggests a critical role of innate immunity. Toll-like receptors (TLRs) trigger the recognition of invading pathogens by innate immune cells and the onset of specific immune responses. The aim of this work was to assess, immunohistochemically, the expression of TLR1, TLR2, TLR4, and TLR9 in intestinal samples of 20 cows showing different types of paratuberculous lesions: uninfected controls, focal lesions, paucibacillary, and multibacillary diffuse forms. The majority of labeled cells were morphologically consistent with macrophages. A differential cell count was performed in the intestinal lamina propria, gut-associated lymphoid tissue, and mesenteric lymph node. TLR9 immunolabeling between the different types of lesions was compared using a complete H-score. Focal and diffuse paucibacillary forms contained significantly increased TLR2-expressing macrophages outside of the lesions compared with the controls and diffuse multibacillary forms, and moderate TLR9 immunolabeling within granulomas. In the multibacillary granulomatous lesions, the expression of TLR1 and TLR4 was observed as well as increased TLR9 expression compared with the rest of the groups. Differences in the predominance of one type or another of TLR allows us to elucidate the importance of the innate immune response and its possible role in the development of the different types of paratuberculosis lesions.

The present study aimed to evaluate the histologic, histochemical, and immunohistochemical changes in buffalo livers with cystic echinococcosis. Noninfected and infected livers were collected from the freshly slaughtered buffalo at the Aligarh abattoir. Small pieces of both infected and noninfected livers (n = 5) were cut and processed for histologic and histochemical studies. Immunohistochemistry was performed using rabbit anti-CD3, CD19, and CD117 antibodies. The results revealed the presence of brood capsules and germinal and laminated membranes surrounded by a fibrous adventitial layer, followed by moderate and diffused infiltration of eosinophils, monocytes, neutrophils, lymphocytes, and marked focal infiltration of mast cells. The infected livers also had mild dilation of central veins and sinusoids, mild and focal necrosis of hepatic tissue, and congestion of central and portal veins. Periodic acid-Schiff reaction revealed marked glycogen depletion in the infected liver. Masson’s trichrome stain showed marked deposition of collagen fibers in the portal area, adventitial layer, and between the hepatocytes compared with the noninfected liver, where deposition was found only in the portal area. The T-cell response was more pronounced than the B-cell response in infected liver. Thus, it can be concluded that hydatid cyst infection causes several pathological and biochemical changes and increased infiltration of inflammatory cells in the infected livers, suggesting the involvement of nonspecific immune responses against hydatid cysts. The T-cell response was more pronounced than B-cells, indicating the involvement of cell-mediated immunity against cystic echinococcosis. These findings may help to understand the local immune responses to cystic echinococcosis.

Intoxication of sheep and cattle by Astylus atromaculatus recently occurred in Uruguay and Argentina in association with severe drought. Although the disease was experimentally reproduced in sheep in the 1970s, there is limited information on clinical and pathologic findings of sheep experimentally intoxicated by this beetle. Here, we described the clinical, gross, and microscopic findings in 3 sheep orally dosed with A. atromaculatus (treatment group, TG) and in 2 control sheep (control group, CG) dosed with distilled water. Anorexia, lethargy, ruminal stasis, reluctance to move, prolonged recumbency, and bruxism were observed in the TG but not the CG sheep. Gross postmortem lesions were only observed in TG sheep and included fibrinonecrotizing enteritis affecting the duodenum, jejunum, and ileum, and multifocal hemorrhages in rumen, omasum, and abomasum. Microscopically, all 3 TG animals had multifocal necrosis in the small intestine; the lesions were most severe in the jejunum. Multifocal necrosis was seen in the mucosa of the rumen, omasum, and abomasum. No significant gross or microscopic abnormalities were observed in the 2 CG sheep. The study supports the role of A. atromaculatus in acute, fatal gastrointestinal disease like that previously described in experimental and spontaneous cases in sheep.

Atypical Bacillus cereus strains (currently classified as Bacillus tropicus) capable of causing anthrax-like disease in humans and animals have emerged in the last 2 decades. These emerging strains are characterized by the acquisition of virulence plasmids pBCXO1 and pBC210, which are homologous to the pXO1 and pXO2 virulence plasmids of Bacillus anthracis, the agent of anthrax. The aim of this study was to describe the gross, histologic, microbiologic, and molecular features of an outbreak of anthrax-like septicemia caused by B. tropicus in red kangaroos (Macropus rufus). Three red kangaroos from a wild animal preserve were found dead with no premonitory clinical signs. No changes in husbandry were reported prior to the outbreak. The peracute disease process was characterized by severe splenomegaly, associated with fibrinonecrotizing splenitis in all affected animals, in addition to segmental suppurative enteritis in 2 kangaroos and cutaneous excoriations, with underlying necrotizing cellulitis and lymphadenitis, in 1 kangaroo. Numerous intralesional, gram-positive and capsulated bacilli were identified as the formerly known B. cereus group via bacteriologic culture. Whole-genome sequencing from one of the bacterial isolates (designated 11844) revealed numerous anthrax-like virulence factors, including the pBCXO1 and pBC210 virulence plasmids. This isolate also had a close phylogenetic relationship with other B. tropicus strains carrying these virulence plasmids, including B. tropicus (formerly B. cereus) G9241. This is the first report of B. tropicus leading to anthrax-like disease in kangaroos. This disease form carries significant public health risks due to potential zoonotic transmission.

“Pix” is one of the most common skin defects that reduce the quality of crocodilian leather. The name is derived from their resemblance to pit marks made by an ice pick. Histologically, each “pix” is associated with a focal dermal accumulation of immune cells, specifically lymphocytes and histiocytes. Consequently, these defects have been termed lymphohistiocytic proliferative cutaneous lesions (LPCLs). In farmed American alligators (Alligator mississippiensis), LPCLs have been associated with seropositivity against West Nile virus (WNV) and the presence of viral genome in the skin. Despite this association, the nature and pathogenesis of LPCLs remain unclear. Using immunohistochemistry and in situ hybridization, we unravel the microanatomy of LPCLs of alligators and localize WNV genome within the lesions. Our results show that LPCL lesions consist of de novo follicular aggregates of lymphocytes segregated into B- and T-cell zones, like tertiary lymphatic follicles of mammals and birds. Furthermore, the presence of WNV genome was highlighted by in situ hybridization in the macrophages of LPCLs, gut-associated lymphoid tissues, and the spleen. Our results suggest that LPCLs may form in American alligators’ skin as part of a generalized lymphofollicular proliferation, likely as an immune response against WNV infection.

Collection of coral for histologic examination requires holding of samples in seawater for a time before they are fixed for histologic processing. This could adversely affect the interpretation of morphologic changes during histologic examinations. We evaluated the microscopic morphology of Porites evermanni and Montipora capitata held (0–120 minutes) in seawater prior to fixation in Z-Fix formulated with raw or artificial seawater. We saw no evident effects of treatments on microscopic morphology. However, among 88 statistical comparisons, and after accounting for false discovery rate, holding time prior to fixation was associated with a significant increase in degree of mucosity of basal body walls.

Hepatic hemosiderosis is a common nutritional disease affecting numerous bird species. The aim of this study was to validate the use of pinch hepatic biopsies to quantify the hepatic iron in Asian glossy starlings (AGS) and compare clinical measures usable for antemortem disease monitoring. Following euthanasia, serum, liver pinch biopsies, and whole liver sections were sampled from 21 AGS. Liver biopsies and sections were stained with Perl’s Prussian blue, and the area containing iron pigment was calculated by image analysis. The mean percent iron-stained area per whole hepatic section was 62.52% ± 21.81%, while the mean percent iron-stained area in biopsies was 71.47% ± 19.77%. Positive correlations were found between hepatic iron concentration and percentage iron in whole sections (r = 0.92; P < .0001), percentage iron in biopsies (r = 0.91; P < .0001), and other clinico-pathological features. Hemosiderin in hepatic biopsies was significantly higher than in whole hepatic section assessed by image analysis (P < .0001).

Coiled-coil domain containing 85c (Ccdc85c) is a causative gene for genetic hydrocephalus found in hemorrhagic hydrocephalus (hhy) mice. The Ccdc85c knockout (KO) rat has subcortical heterotopia with frequent brain hemorrhage as seen in hhy mice. In this study, we report aberrant alpha-smooth muscle actin (α-SMA) expression in the wall of lateral ventricle of the Ccdc85c KO rats. The α-SMA-positive cells were distributed at the dorsal, medial, and lateral regions of the lateral ventricle of KO rats. The expression of α-SMA was first observed on postnatal day 20 (P20) and became noticeably stronger at P26 when hydrocephalus was prominent. Double immunofluorescence showed co-expression of α-SMA with nestin, vimentin, and glial fibrillary acidic protein in the ventricular lining of KO rats. Therefore, we conclude that α-SMA-positive cells may represent an immature subpopulation of cells at adult age around the lateral ventricle of Ccdc85c KO rats.

Neoplasms are only sporadically reported in New World primates and rarely in owl monkeys (Aotus spp.), specifically. Previous reports of neoplasms in owl monkeys are primarily restricted to lymphoma induced by Herpesvirus saimiri infection, although other tumors in the central nervous, genitourinary, gastrointestinal, and endocrine systems have been sporadically reported. Herein, we describe 3 previously unreported neoplasms in owl monkeys (Aotus nancymaae) including a pericardial mesothelioma in a 6-year-old male, a nephroblastoma in a 2-year-old male, and a cervical neoplasm with characteristics of an epithelioid trophoblastic tumor in a 4-year-old female, all occurring in the same closed breeding colony at a research facility in central Texas. Reporting of spontaneously occurring neoplasms in research colony populations is important for identifying potential animal models of human diseases and for improving colony management and species health.

This report describes cardiovascular and renal soft tissue mineralization and renal intratubular crystals in 13 out of 16 guinea pigs that were given very hard drinking water for 9 months. These animals, aged 14 to 20 months, were experimentally naïve. No clinical symptoms were observed, but 1 guinea pig was found dead in its cage. Necropsy did not reveal any gross findings; however, histologic examination revealed mineralization and crystal formations. Despite no known changes in the feed sourcing or formulation, the possibility that the incident was feed-related was considered. The most recent analysis of the feed obtained from the manufacturer during this period, which was conducted by an accredited laboratory authorized by the Ministry of Agriculture and Forestry, was appropriate. No similar lesions were reported at other centers using the same feed; however, drinking water analysis for total dissolved solids revealed extremely hard water, with elevated levels of calcium and calcium carbonate and low magnesium levels, due to a malfunctioning water treatment system. After installing a new system to balance calcium and magnesium, no new cases appeared over the next 2 years. It was determined that the mineralization and crystal formations were most likely caused by water hardness. This study demonstrates that mineralization typically attributed to feed in guinea pigs can also result from high calcium content in drinking water, highlighting the importance of water analysis in such cases.

Acute pancreatitis (AP) is a life-threatening condition, with a higher mortality rate in men than women and in which estrogens might play a protective role. This study aimed to investigate sex-dependent differences in a mouse model of caerulein-induced AP. Thirty-six C57BL/6J mice (19 females and 17 males) were treated intraperitoneally with phosphate-buffered saline or caerulein, and sacrificed 12 hours, 2 days, or 7 days after the last injection. Blood was collected for amylase, lipase, and glucose determination. Severity and extent of inflammation, apoptosis, and acinar to ductal metaplasia (ADM) in pancreatic tissue were scored histologically and total macrophages, major histocompatibility complex (MHC)-II+ cells, M2 macrophages, T and B cells, neutrophils, apoptosis, and ADM were marked immunohistochemically and quantified by digital image analysis. Serum amylase had a peak at 12 hours, without differences between the sexes. In females, pancreatitis reached a peak at 12 hours with a fast recovery while, in males, the peak was delayed to day 2 with residual apoptosis still present. Macrophages were the main inflammatory cell population, followed by T cells, B cells and neutrophils, without differences between sexes. In males, CD206+ cells and apoptosis were higher at both days 2 and 7, and cytokeratin-19+ (CK19+) ADM was higher at day 7 compared with females. The results of this study revealed a faster onset and resolution of caerulein-induced AP in female mice compared with male mice, supporting a sex-dependent modulation of acute pancreatitis.

The kidney plays an important role in iron homeostasis and mesangial cells (MCs) are phagocytic cells important for glomerular homeostasis. Sickle hemoglobin (HbS) modulators are promising clinical candidates for treatment of sickle cell disease. Although they prevent disease pathophysiology of HbS polymerization and red blood cell (RBC) sickling by increasing hemoglobin oxygen affinity, higher oxygen affinity can also cause transient tissue hypoxia with compensatory increases in erythropoiesis and subsequent increases in RBC turnover. CD-1 mice treated with an HbS modulator for 2 weeks developed higher RBC mass, increased erythropoiesis, and, by 1 month, deposition of intracellular pigments in renal tubular and parietal epithelium. In addition, in mice treated for 26 weeks, pigment was observed in MCs, which was accompanied by glomerular cell aggregates (MC hypercellularity) and tubulo-interstitial inflammation. The pigment was confirmed by Perl’s iron staining and transmission electron microscopy (TEM) to be iron-containing proteins. Glomerular cell aggregates were confirmed to be MCs by TEM, and Ki-67 immunolabeling suggested that MC hypercellularity was due to proliferation. Collectively, these findings, along with iron-containing proteins in livers and spleens, suggested that iron overload secondary to increased RBC turnover led to increased renal iron reabsorption. While both MC hypercellularity and tubulo-interstitial inflammation were thought to be responses to long-term accumulation of iron, the former was considered a homeostatic response to eliminate iron, and maintain glomerular structure and function, while the latter was more consistent with an iron-catalyzed oxidative stress response. To our knowledge, this is the first report of MC hypercellularity in a preclinical toxicity study.