Studies on the association of malondialdehyde as a biomarker for oxidative stress and degree of malignancy in dogs with mammary adenocarcinomas.

BACKGROUND: Mammary adenocarcinomas are one of the most common tumour diseases in bitches. The relationship between oxidative stress and the degree of malignancy of the tumour has not been sufficiently researched in veterinary medicine. OBJECTIVES: The main objective was to investigate the potential role of MDA as a practice-relevant biomarker for the assessment of systemic oxidative stress and to determine whether this parameter can indicate the malignancy grade of a mammary adenocarcinoma. METHODS: In the present pilot study, MDA plasma concentrations were analysed in 55 bitches with (n = 28) and without (n027) malignant adenocarcinomas of the mammary gland using two different measurement methods and the relationship to tumour size was investigated. RESULTS: The mean MDA concentration measured by enzyme-linked immunosorbent assay (ELISA) was 289 ng/mL (range 365-634 ng/mL) in dogs with grade 1 adenocarcinoma (n = 13), 288.5 ng/mL (range 85-752 ng/mL) in dogs with grade 2 adenocarcinoma (n = 10), 332 ng/mL (range 239-947 ng/mL) in dogs with grade 3 (n = 5) adenocarcinoma and 293 ng/mL (range 175-549 ng/mL) in dogs without a mammary tumour (n = 27). When MDA was measured by HPLC, the average MDA concentration in the study group (n = 11) was 0.24 µmol/L (range 0.16-0.37) and that of the control group (n = 15) was 0.27 µmol/L (range 0.16-1.62). Thus, there were no significant differences between the study group with malignant adenocarcinomas and the control group in both examination methods (p > 0.05). Furthermore, there was no correlation between the MDA concentrations and the approximate volume of the mammary tumour. CONCLUSION: The results highlight the challenges of providing a prognosis for the malignancy of a mammary adenocarcinoma based on MDA concentrations in plasma using ELISA or HPLC. As a result, histopathological examination remains the gold standard for diagnosing and differentiating adenocarcinomas of the mammary gland.

Cutaneous apocrine gland neoplasia in 16 captive African wild dogs (Lycaon pictus).

This retrospective study documents the occurrence of single and multiple cutaneous apocrine gland tumours (CATs) on the dorsal midline of 16 captive African wild dogs (AWDs, Lycaon pictus) derived from 161 submissions to diagnostic laboratories in South Africa, France and Germany between 1997 and 2022. Animals included in the study came from zoological institutions in South Africa (n = 2), France (n = 5) and Germany (n = 1) and ranged from 5 to 14 years of age. Fifteen affected animals were female (94%) and one was male. CATs presented as raised, hairless, multilobular, grey firm masses, consistently located along the dorsal midline. Apart from a single cutaneous apocrine adenoma and a cystadenoma occurring concurrently with two non-cystic adenocarcinomas, neoplasms were consistent with malignant cutaneous apocrine adenocarcinomas with lymphatic spread and visceral metastases. Advanced age and female sex were identified as risk factors. A genetic component or association with the increasing use of GnRH agonist contraceptives was suspected but could not be established. This study highlights the need for close clinical monitoring of AWDs over the age of 5 years for the development of CATs along the dorsal midline and supports early surgical intervention. More research is needed to determine the role of inbreeding, endocrine changes and husbandry factors that may play a role in the development of CATs on the dorsal midline of AWDs.

Immobility-associated thromboprotection is conserved across mammalian species from bear to human.

Venous thromboembolism (VTE) comprising deep venous thrombosis and pulmonary embolism is a major cause of morbidity and mortality. Short-term immobility-related conditions are a major risk factor for the development of VTE. Paradoxically, long-term immobilized free-ranging hibernating brown bears and paralyzed spinal cord injury (SCI) patients are protected from VTE. We aimed to identify mechanisms of immobility-associated VTE protection in a cross-species approach. Mass spectrometry-based proteomics revealed an antithrombotic signature in platelets of hibernating brown bears with heat shock protein 47 (HSP47) as the most substantially reduced protein. HSP47 down-regulation or ablation attenuated immune cell activation and neutrophil extracellular trap formation, contributing to thromboprotection in bears, SCI patients, and mice. This cross-species conserved platelet signature may give rise to antithrombotic therapeutics and prognostic markers beyond immobility-associated VTE.

Macrophages in dermal disease progression of phospholipase D4-deficient Fleckvieh calves.

A new gene defect in Fleckvieh calves leads to a syndrome with partial phenotype overlap with bovine hereditary zinc deficiency. A mutation in a gene encoding phospholipase D4 (PLD4), an endosomal exonuclease, causes the disorder. In mice, PLD4 activity indirectly regulates the Toll-like receptor 9 (TLR9) pathway via degradation of microbial DNA. PLD4 absence thus results in visceral macrophage activation comparable to human macrophage activation syndrome. In this study, disease progression and the role of macrophages in affected calves were monitored clinically, clinicopathologically, and histologically over time. Breeding data identified 73 risk matings of heterozygous carriers resulting in 54 potentially PLD4-deficient calves born on farms. PLD4 status was examined via 5'-exonuclease assay, detecting 6 calves carrying the defect. These were purchased and monitored daily until final necropsy. The calves developed progressive skin lesions starting with small scaling areas terminating in severe crusting dermatitis, especially in areas with mechanical exposure. Histological and immunohistochemical analyses indicated that macrophages with cytoplasmic vacuolation increased considerably in skin sections obtained weekly during the disease course. Macrophage increase correlated with increased dermal lesion severity. Macrophage activation was confirmed by prominent phagocytic activity in the superficial dermis using electron microscopy. Dermal mRNA abundance of CCL2 and CCL3 measured by quantitative polymerase chain reaction verified macrophage activation. Further increase in mRNA of downstream molecule MyD88 and cytokine IL12b connected bovine PLD4 deficiency to increased TLR9 pathway activation. In contrast to human macrophage activation syndrome, the main feature of bovine PLD4 deficiency was local disease in organs with contact to microbial DNA (skin, intestine, lungs).

A case series of canine cutaneous inverted papilloma with one case showing evidence of recurrence.

BACKGROUND: This article describes a case of multiple, recurrent, cutaneous inverted papillomas (CIPs) in a German shepherd dog, combined with a retrospective follow-up assessment of eight cases. METHODS: A 3-year-old, black, female German shepherd dog presented with four rapidly growing, alopecic, domed, hyperpigmented, shiny nodules, compatible with CIP. The dog was managed for pruritus, associated with atopic dermatitis, with long-term prednisolone therapy and this therapy was continued after the surgical excision. Twelve months after the initial presentation the dog was represented for two new CIP lesions, coinciding with an increased dosage of prednisolone during an allergic flare-up period. RESULTS: Histopathological evaluation was conducted on both the original and subsequent lesions from the case with recurrence. Eight cases of CIP, based on histopathological findings, were identified retrospectively from the authors' diagnostic pathology service over the previous two years. All nine cases had lesions that exhibited varying degrees of inverted epithelial hyperplasia, multiple, endophytic, papillary epidermal projections, a cup-shaped base with central hyperkeratosis and active viral pathological findings (koilocytes). The submitting veterinarians were contacted and follow-up regarding recurrence and concurrent medications was obtained. CONCLUSIONS AND CLINICAL IMPORTANCE: CIP is uncommonly reported, typically as a single lesion with no previous reports of recurrence, although one group of dogs with severe combined immunodeficiency developed invasive malignancies. In seven of eight retrospective cases no recurrence of CIP was recorded. The authors speculate that the recurrence in the German shepherd dog may have been associated with chronic (although low-dose) glucocorticoid administration.

Histomorphology and Immunohistochemistry of a Congenital Nephromegaly Demonstrate Concurrent Features of Heritable and Acquired Cystic Nephropathies in a Girgentana Goat (Capra falconeri).

Polycystic kidney diseases (PKD) represent frequent congenital and adult nephropathies in humans and domestic animals. This report illustrates an uncommon state of congenital PKD in a girgentana goat (Capra falconeri). A stillborn female goat kid was submitted for postmortem examination and underwent macroscopic and microscopic examination. The kidneys showed a bilateral nephromegaly and a perpendicular polycystic altered texture of the renal parenchyma. Renal tissue sections were comprehensively investigated by histopathology (overview and special stains), immunohistochemistry (CD10, CD117, pan-cytokeratin, cytokeratin 7, E-cadherin, Pax2, Pax8, and vimentin), and electron microscopy (SEM, TEM). Histopathology of renal tissue sections revealed polycystic alterations of the renal parenchyma as well as conspicuous polypoid proliferates/projections of the renal tubular epithelium, which showed clear cell characteristics. Furthermore, epithelial projections were indicative for epithelio-mesenchymal-transition, cellular depolarization, and strong expression of differentiation markers Pax2, Pax8, and CD10. Ultrastructural morphology of the projections was characterized by numerous diffusely distributed, demarked round cytoplasmic structures and several apico-lateral differentiations. Additionally, hepatic malformations comprising biliary duct proliferation with saccular dilation and bridging fibrosis were observed. Notably, this report describes the first case of a congenital cystic nephropathy with overlapping features of heritable and acquired nephropathies in any species. Epithelio-mesenchymal-transition and altered cadherin expression seem to be crucial components of a suspected pathomechanism during cystogenesis.

Alopecia in Belgian Blue crossbred calves: a case series.

BACKGROUND: Alopecia is defined as the partial or complete absence of hair from areas of the body where it normally grows. Alopecia secondary to an infectious disease or parasitic infestation is commonly seen in cattle. It can also have metabolic causes, for example in newborn calves after a disease event such as diarrhoea. In the article, the investigation of a herd problem of acquired alopecia in Belgian Blue (BB) crossbred calves is described. CASE PRESENTATION: Several BB crossbred calves had presented with moderate to severe non-pruritic alopecia in a single small herd located in Southern Germany. The referring veterinarian had ruled out infectious causes, including parasitic infection and had supplemented calves with vitamins (vitamins A, B1, B2, B3, B5, B6, B7, B9, B12, C, and K3) orally. Results of the diagnostic workup at the Clinic for Ruminants are presented for three affected calves and findings from a farm visit are discussed. Because of these investigations, an additional four calves were brought to the referral clinic within the first week of life, and before onset of alopecia, in order to study the course of the condition; however, these calves never developed any signs of alopecia during their clinic stay. CONCLUSIONS: Because all other plausible differential diagnoses were ruled out during our investigation, we concluded that the documented alopecia was due to malabsorption of dietary fat and consecutive disruption of lipid metabolism leading to telogen or anagen effluvium. In this particular case, this was caused by a mixing error of milk replacer in conjunction with insufficiently tempered water. We conclude that nutritional, management or environmental factors alone can lead to moderate to severe alopecia in calves in the absence of a prior or concurrent disease event or infectious cause.

Recovering host cell-free Anaplasma phagocytophilum from HL-60 cells by using rock tumbler grit in comparison to the syringe lysis method.

Anaplasma phagocytophilum (Ap) is a tick-transmitted obligate intracellular bacterium and the causative agent of the granulocytic anaplasmosis in various species of domestic animals and in humans. During intracellular development Ap transforms from a dense-cored cell form into a reticulate cell form and vice versa. For isolation of intracellular bacteria, a range of different purification methods is used. However, unlike other Gram-negative bacteria Ap is considered to be sensitive to mechanical stress and osmolarity changes. An updated semi-purification method using rock tumbler grit is introduced here to increase the outcome of bacteria and to facilitate the procedure of host cell lysis. The objective of this study was to evaluate the structural integrity and infectivity of Ap after lysis of the host cells using rock tumbler grit and to compare the outcome to that of the frequently used method, syringe lysis. Human promyelocytic leukemia cell lines (HL-60) were infected with Ap and following host cell-free bacteria were assessed by transmission electron microscopy. The outcome of the different purification methods was compared using live/dead-staining based on immunofluorescence to count the number of viable bacteria and real-time PCR to compare the amount of DNA. Subsequently the isolated bacteria were tested to infect naive cell cultures. We observed that both Ap dense-cored cells and reticulate cells are preserved intact after the application of rock tumbler grit. The number of viable, host cell-free bacteria was higher by factor 1.7-2.4 compared to the syringe lysis protocol. Quantitative analysis based on real-time PCR showed an increase of bacterial DNA up to 1.6-2.9 times higher using the rock tumbler grit protocol. Bacteria released from the same number of infected host cells were used for new infections. Flow cytometric analysis of the cell cultures confirmed that the number of Ap organisms recovered by using the rock tumbler grit protocol resulted in higher infection rates than the number of Ap organisms recovered by using syringe lysis protocol. Our observations indicate that the rock tumbler grit protocol can be applied as a safe, robust and convenient method to recover Ap compared to syringe lysis.

Occurrence of dysentery-like diarrhoea associated with Brachyspira suanatina infection on a German fattening pig farm.

The anaerobic intestinal spirochaete Brachyspira (B.) suanatina was first described in 2007 but since then no further isolates have been reported from pigs. Accordingly, when the species was validly published in 2016, the overall occurrence and clinical relevance in pigs were unknown. In a fattening farm in southern Germany, mucohaemorrhagic diarrhoea was observed in 60 per cent (750 animals) of the finisher pigs. A diagnostic workup including Brachyspira culture, Salmonella culture, Lawsonia intracellularis-specific, B. hyodysenteriae-specific and B. pilosicoli-specific multiplex PCR and postmortem examination of severely affected pigs was performed. Tests for Salmonella species, Lawsonia intracellularis and B. hyodysenteriae were all negative. Gross and microscopic lesions were in agreement with dysentery and spirochaetes could be demonstrated by silver staining in tissue samples of the caecum at the ileal papilla. B. suanatina was cultured from faeces or colon of all (five) animals sampled and identified using nox-RFLP, partial nox-gene-sequencing and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). According to the initial report from Scandinavia, B. suanatina can be isolated from birds and cross-species infection could be demonstrated infecting pigs with an avian isolate. Thus outdoor production as in the case presented here and international trade may pose a risk for infection of naive herds.

A nonsense mutation in PLD4 is associated with a zinc deficiency-like syndrome in Fleckvieh cattle.

BACKGROUND: Bovine hereditary zinc deficiency (BHZD) is an autosomal recessive disorder of cattle, first described in Holstein-Friesian animals. Affected calves suffer from severe skin lesions and show a poor general health status. Recently, eight calves with the phenotypic appearance of BHZD have been reported in the Fleckvieh cattle population. RESULTS: In spite of the similar disease phenotypes, SLC39A4, the gene responsible for BHZD in Holstein-Friesian was excluded as underlying gene for the disorder in the affected Fleckvieh calves. In order to identify the disease-associated region, genotypes of eight affected calves obtained with the Illumina BovineHD BeadChip comprising 777,962 SNPs were contrasted with the genotypes of 1,339 unaffected animals. A strong association signal was observed on chromosome 21 (P = 5.87 × 10(-89)). Autozygosity mapping in the eight affected animals revealed a common segment of extended homozygosity encompassing 1,023 kb (BTA 21: 70,550,045 - 71,573,501). This region contains 17 genes/transcripts, among them two genes encoding gastro-intestinal zinc transporters (CRIP1, CRIP2). However, no mutation that was compatible with recessive inheritance could be detected in these candidate genes. One of the affected calves was re-sequenced together with 42 unaffected Fleckvieh animals. Analysis of the sequencing data revealed a nonsense mutation (p.W215X) in a phospholipase encoding gene (PLD4) as candidate causal polymorphism. To confirm the causality, genotypes of the p.W215X-mutation were obtained from 3,650 animals representing three different breeds. None of the unaffected animals was homozygous for the defect allele, while all eight affected calves were homozygous. The deleterious effect of the mutation is manifested in a significantly lower survival rate of descendants from risk matings when compared with the survival rate of descendants from non-risk matings. The deleterious allele has an estimated frequency of 1.1% in the Fleckvieh population. CONCLUSION: Our results provide strong evidence that a newly identified recessive disorder in the Fleckvieh population is caused by a nonsense mutation in PLD4, most likely resulting in an impaired function of the encoded protein. Although the phenotype of affected calves strongly resembles BHZD, a zinc deficiency resulting from malabsorption is unlikely to be responsible for the diseased Fleckvieh calves.