Chondroitin sulfate proteoglycan-4: a biomarker and a potential immunotherapeutic target for canine malignant melanoma

Chondroitin sulfate proteoglycan-4 (CSPG4), also known as high molecular weight-melanoma associated antigen (HMW-MAA), is a membrane-bound chondroitin sulfate proteoglycan highly expressed by human melanoma cells. This phylogenetically conserved tumour antigen plays an important biological role in human melanoma, where it is used as a marker to diagnose forms with unusual characteristics, such as desmoplastic melanoma, and to detect melanoma cells in lymph nodes and peripheral blood, and as a target for immunotherapy because of its restricted distribution in normal tissues. To identify suitable targets to develop novel approaches of treating canine melanoma, CSPG4 was studies to see whether it is expressed in canine malignant melanomas. Immunohistochemical staining of 65 canine malignant melanomas with an anti-human CSPG4-specific antibody detected CSPG4 in 37 cases (56.9%). Positive staining was more frequent, albeit not significantly, in amelanotic compared to melanotic tumours and was statistically associated with tumours having both melanin and the epithelioid histotype. The frequency of CSPG4 expression was similar to that of other melanoma antigens used as diagnostic markers for canine malignant melanoma, such as Melan A and the protein recognized by the PNL2 monoclonal antibody. The results suggest that CSPG4 constitutes a new potential immunohistochemical marker of canine malignant melanoma and may represent an immunotherapeutic target as in humans.     

Tetrodotoxins (TTXs) and Vibrio alginolyticus in Mussels from Central Adriatic Sea (Italy): Are They Closely Related?

Tetrodotoxins (TTXs), potent neurotoxins, have become an increasing concern in Europe in recent decades, especially because of their presence in mollusks. The European Food Safety Authority published a Scientific Opinion setting a recommended threshold for TTX in mollusks of 44 µg equivalent kg⁻¹ and calling all member states to contribute to an effort to gather data in order to produce a more exhaustive risk assessment. The objective of this work was to assess TTX levels in wild and farmed mussels (Mytilus galloprovincialis) harvested in 2018–2019 along the coastal area of the Marche region in the Central Adriatic Sea (Italy). The presence of Vibrio spp. carrying the non-ribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) genes, which are suspected to be involved in TTX biosynthesis, was also investigated. Out of 158 mussel samples analyzed by hydrophilic interaction liquid chromatography coupled with tandem mass spectrometry (HILIC-MS/MS), 11 (7%) contained the toxins at detectable levels (8–26 µg kg⁻¹) and 3 (2%) contained levels above the EFSA safety threshold (61–76 µg kg⁻¹). Contaminated mussels were all harvested from natural beds in spring or summer. Of the 2019 samples, 70% of them contained V. alginolyticus strains with the NRPS and/or PKS genes. None of the strains containing NRPS and/or PKS genes showed detectable levels of TTXs. TTXs in mussels are not yet a threat in the Marche region nor in Europe, but further investigations are surely needed.     

Right auricle aneurysm in dogs can be detected with multidetector-row CT and presents with varying morphology

Right auricle aneurysm (RAA) has been reported to be a rare congenital or acquired condition in dogs, however published CT characteristics are limited to a small number of cases. The aim of this 13-year, single-center, retrospective, cross-sectional study was to describe the prevalence and appearance of RAA in a larger group of dogs. Reports of 10,886 dogs that underwent thoracic CT examination for various reasons were reviewed. Computed tomographic data of dogs with presumed RAA (based on published imaging characteristics) were retrieved from the archive. Oblique multiplanar reformatted images were used to record RAA site, shape, and dimensions (neck, maximum height and width). Patients’ sex, age, and body weight were evaluated for any association with the presence of RAA. Presumed RAA was detected in 23/10,886 dogs having thoracic CT in the selected period of time (prevalence 0.21%). Three RAA phenotypes were described: diffuse saccular dilation in 19 of 23 dogs, focal saccular dilation in three of 23 dogs, and fusiform in one of 23 dogs. The bodyweights of dogs with presumed RAA were significantly lower than those of the remaining 10836 dogs without RAA (median 8 kg [IQR 5; range 37.6] and median 16 kg [IQR 23; range 116.9] respectively; P < 0.003. Female sex was also significantly associated with RAA (P = 0.03). Findings indicated that RAA has a low prevalence in dogs, can be detected with CT, can be present in asymptomatic dogs, and can have varying appearances. In this sample of dogs, female sex and low body weight were significantly associated with RAA.     

Results of magnetic resonance imaging in 14 cats with meningoencephalitis

Medical records and magnetic resonance (MR) images of 14 cats with inflammatory diseases affecting the central nervous system (CNS) were reviewed retrospectively. Cases included eight cats with feline infectious peritonitis and two cats with toxoplasmosis. Abnormalities affecting the CNS were observed in MR images in 10 (71%) cats. Intracranial lesions appeared as slightly hypointense foci in T1-weighted images in two (14%) cats, as hyperintense foci in T2-weighted images in seven (50%) cats and as hyperintense foci after intravenous administration of a gadolinium-based contrast medium in 10 (71%) cats. In six cats with lesions in T1- and/or T2-weighted images, additional lesions were visible in T1-weighted images obtained after gadolinium-based contrast medium administration. In three cats, lesions were visible only after contrast medium administration. In our study, MR imaging (MRI) did not appear to detect all cases of CNS inflammation in the population of cats with inflammatory cerebrospinal fluid (CSF); however, MRI adds information about the sites and morphology of intracranial lesions that should help to distinguish between neoplasia and inflammatory conditions and, possibly, between different inflammatory conditions.     

Acute basophilic leukaemia in a three-month-old calf

A three-month-old female Holstein–Friesian calf was presented with acute tetraparesis. After neurological examination a multifocal lesion in the central nervous system was suspected with the most pronounced lesions between the third thoracic and the third lumbar vertebrae. Haematological examination revealed moderate anaemia as well as severe thrombocytopenia, neutropenia and leucocytosis. A blood smear and bone marrow aspirate exhibited predominantly blasts with basophilic granulation leading to a diagnosis of acute (myeloid) leukaemia with involvement of the basophilic lineage or an acute basophilic leukaemia. Magnetic resonance imaging revealed spinal cord compression; at necropsy, extensive localised haemorrhages extending into the thoracic vertebral canal were found. Histopathology revealed a large population of blast cells in several tissues including the meninges. Due to multifocal detection of neoplastic cells in the vascular system, neoplasia of the haematopoietic system was assumed in agreement with haematological findings. Signs of paresis could be explained by intramedullary spinal cord haemorrhage and myeloid infiltrations of meningeal vessels. In conclusion, despite its rarity, acute myeloid leukaemia with involvement of the basophilic lineage may be considered in diagnosing calves with progressive deteriorating general condition, paresis, leucocytosis with moderate basophilic differentiation or haemorrhagic disorders.     

Comparison of equine bone marrow-, umbilical cord matrix and amniotic fluid-derived progenitor cells

The aim of the study was to compare in vitro the stemness features of horse progenitor cells derived from bone marrow (BM-MSCs), amniotic fluid (AF-MSCs) and umbilical cord matrix (EUC-MSCs). It has been suggested that there may be a stem cell population within both umbilical cord matrix and amniotic fluid. However, little knowledge exists about the characteristics of these progenitor cells within these sources in the equine species. This study wanted to investigate an alternative and non-invasive stem cell source for the equine tissue engineering and to learn more about the properties of these cells for future cell banking. Bone marrow, umbilical cord and amniotic fluid samples were harvested from different horses. Cells were analyzed for proliferation, immunocytochemical, stem cell gene expression and multilineage plasticity. BM- and AF-MSCs took similar time to reach confluence and showed comparable plating efficiency. All cell lines expressed identical stem cell markers and capability to differentiate towards osteogenic lineage. Almost all cell lines differentiated into the adipogenic lineage as demonstrated by cytochemical staining, even if no adipose gene expression was detectable for AF-MSCs. AF- and EUC-MSCs showed a limited chondrogenic differentiation compared with BM-MSCs as demonstrated by histological and biochemical analyses. These findings suggest that AF-MSCs appeared to be a readily obtainable and highly proliferative cell line from an uninvasive source that may represent a good model system for stem cell biology. More studies are needed to investigate their multilineage potential. EUC-MSCs need to be further investigated regarding their particular behavior in vitro represented by spheroid formation.     

Multivariate analysis of soils: microbial biomass,metabolic activity,and bacterial‐community structure and their relationships with soil depth and type

A multivariate statistical approach based on a large data set of abiotic and biotic variables was used to classify four contrasting‐land‐use soils. Soil samples were collected at increasing depth from a calcareous agricultural soil, a temperate upland grassland soil, a moderately acidic agricultural soil, and an acidic pine forest soil. Analytical investigations were carried out by using a combination of conventional physical, chemical, and biochemical methods coupled with denaturing gradient gel electrophoresis (DGGE) community fingerprinting of PCR‐amplified 16S rRNA gene‐coding fragments from soil‐extracted total‐community DNA. The data set of soil physical, chemical, and biochemical variables was reduced in dimensionality by means of a principal‐component‐analysis (PCA) procedure. Compositional shifts in soil bacterial‐community structure were analyzed through a clustering algorithm that allowed identifying six main bacterial‐community clusters. DGGE fingerprinting clusters were further analyzed by discriminant analysis (DA) using extracted PCA components as explanatory variables. Soil organic matter–related pools (TOC, TN) and functionally related active pools (microbial biomass C and N, K₂SO₄‐extractable C) significantly decreased with soil depth, and resulted statistically linked to one other and positively related to enzymatic activities (acid phosphatase, arylsulfatase, β‐glucosidase, dehydrogenase, hydrolysis of fluorescein diacetate) and silt content. Besides organic‐C gradients, pedogenetic‐driven physico‐chemical properties, and possibly soil thermal and moisture regimes seemed to play a key role in regulating size and energetic ecophysiological status of soil microbial communities. DGGE analysis showed that contrasting horizons were conducive to the dominance of particular bacterial ribotypes. DA revealed that the bacterial‐community structure was mainly influenced by organic matter–related variables (TOC, TN, CEC, C_flush, N_flush, Extr‐C), chemical properties such as pH, CaCO₃, and EC, together with textural properties. Results indicate that, beyond land use or plant cover, pedogenetic‐driven physico‐chemical conditions changing with soil type and depth are the key factors regulating microbial size and activity, and determining the genetic structure of bacterial community.