Medical management and positive outcome after prolonged recumbency in a case of equine herpesvirus myeloencephalopathy.

A 17-year-old mare presenting with acute fever, weakness and bladder dysfunction was diagnosed with equine herpesvirus myeloencephalopathy (EHM). The mare become transiently recumbent, underwent parenteral fluid therapy, plasma infusion, steroidal/nonsteroidal anti-inflammatory drugs (SAID/NSAIDs) and bladder catheterization. After 10 days the mare was hospitalized. Neurological evaluation revealed ataxia and proprioceptive deficits mainly in the hind limbs. The mare was able to stand but unable to rise from recumbency or walk. Secondary complications included Escherichia coli cystitis, corneal ulcers and pressure sores. A full-body support sling was used for 21 days. Medical treatment included systemic antimicrobials, NSAIDs, gradual discontinuation of SAIDs, parenteral fluid therapy and bladder lavage. The mare tested positive for Varicellovirus equidalpha 1 (EHV-1) DNA in nasal swab and blood samples on day 13 and in urine samples on days 13 and 25 after the onset of fever. Neurological signs improved over a period of 34 days and the mare was discharged with mild hind limb weakness/ataxia. Secondary complications resolved within 2 weeks. At the eight-month follow-up, marked improvement in locomotory function had been achieved.

Spread of Toxoplasma gondii among animals and humans in Northern Italy: A retrospective analysis in a One-Health framework.

Toxoplasmosis occurs worldwide and is considered one of the most important food-borne parasitic zoonoses. The consumption of undercooked meat containing viable tissue cysts and ingestion of environmental oocyst are the most important sources of infection. The aim of this retrospective study was to evaluate the spread of Toxoplasma gondii in the province of Bologna (Emilia-Romagna region) in northern Italy, with a One Health approach, comparing seropositivity rates in different animal species and in humans over the last 19 and 4 years respectively. Analyses were performed on serological data collected over different periods at three separate locations: Istituto Zooprofilattico Sperimentale della Lombardia e della Emilia-Romagna (IZSLER); Veterinary University Hospital Clinical Pathology Service, Department of Veterinary Medical Sciences, University of Bologna; and Unit of Microbiology, St. Orsola Hospital, Bologna. Most relevant seropositivity rates observed in animals were 15.5% (wild boar), 25% (roe deer), 18.7% (goat), 29.9% (sheep), 9.7% (pigs), 42.9% and 21.8% in cat and dog, respectively. A comprehensive screening was conducted on a population of 36,814 individuals, revealing a prevalence of 20.4%. Among pregnant women, a frequence of 0.39% for active toxoplasmosis was observed. Despite certain limitations, this study provided valuable insights into the extensive distribution of this parasitic infection among diverse animal species and human populations in the province of Bologna. These findings underscore the importance of implementing consistent and proactive toxoplasmosis screening protocols during pregnancy, while emphasizing the critical need for adopting a One Health approach for effective control of this parasitic disease.

Comparison of clinicopathological patterns of renal tubular damage in dogs with acute kidney injury caused by leptospirosis and other aetiologies.

In humans, leptospiral acute kidney injury (AKI) is characterised by tubulointerstitial involvement and renal electrolyte losses, impacting clinical presentation and case management. The aim of this study was to evaluate urine chemistry findings in dogs with leptospirosis in order to identify characteristic patterns of tubular damage associated with this disease. Dogs with intrinsic AKI caused by leptospirosis and by other aetiologies were prospectively enrolled. Clinical and clinicopathological variables, including serum and urine chemistry, fractional excretion (FE%) of electrolytes, and urinary neutrophil gelatinase-associated lipocalin (NGAL), were evaluated in both groups and compared statistically. Dogs with leptospirosis (n = 38) had significantly higher serum creatinine concentration than dogs with AKI caused by other aetiologies (n = 37). Serum potassium and glucose concentrations were comparable between groups. Dogs with leptospiral AKI had significantly higher FE of potassium (median 100%, range 20-480 vs. median 68%, range 5-300; P = 0.048), as well as higher magnitude of glucosuria (urine glucose to creatinine ratio, median 0.64, range 0-26 vs. median 0.22, range 0-13; P = 0.023) and frequency of positive glucose dipstick reaction (59% vs. 18%; P = 0.002), than dogs with AKI of other aetiologies. Additional markers of tubular damage considered in this study, including FE of other electrolytes and urinary NGAL, did not differ between groups. In conclusion, when compared to other aetiologies of intrinsic AKI, canine leptospirosis was characterised by increased glucosuria and kaliuresis.

A novel pancreatic tumour and stellate cell 3D co-culture spheroid model.

BACKGROUND: Pancreatic ductal adenocarcinoma is a devastating disease with poor outcome, generally characterized by an excessive stroma component. The purpose of this study was to develop a simple and reproducible in vitro 3D-assay employing the main constituents of pancreatic ductal adenocarcinoma, namely pancreatic stellate and cancer cells. METHOD: A spheroid assay, directly co-culturing human pancreatic stellate cells with human pancreatic tumour cells in 3D was established and characterized by electron microscopy, immunohistochemistry and real-time RT-PCR. In order to facilitate the cell type-specific crosstalk analysis by real-time RT-PCR, we developed a novel in vitro 3D co-culture model, where the participating cell types were from different species, human and mouse, respectively. Using species-specific PCR primers, we were able to investigate the crosstalk between stromal and cancer cells without previous cell separation and sorting. RESULTS: We found clear evidence for mutual influence, such as increased proliferation and a shift towards a more mesenchymal phenotype in cancer cells and an activation of pancreatic stellate cells towards the myofibroblast phenotype. Using a heterospecies approach, which we coined virtual sorting, confirmed the findings we made initially in the human-human spheroids. CONCLUSIONS: We developed and characterized different easy to set up 3D models to investigate the crosstalk between cancer and stroma cells for pancreatic cancer.

Prospective evaluation of rapid point-of-care tests for the diagnosis of acute leptospirosis in dogs.

The early diagnosis of acute leptospirosis is still a major challenge in dogs. The aim of this prospective study was to evaluate the suitability of two in-clinic tests detecting anti-leptospiral IgM and IgG antibodies in diagnosing canine leptospirosis. The performances of the two rapid tests were compared to the microscopic agglutination test (MAT) carried out on acute sera and to diagnostic criteria adopted in this study to confirm leptospirosis infection (MAT upon admission, convalescent MAT and quantitative real-time PCR on blood and/or urine). The dogs were enrolled on the basis of reported exposure to known risk factors and clinical presentation (acute kidney injury and/or systemic inflammatory response syndrome with multi-organ damage). Eighty-nine dogs included in the study were sub-grouped on the basis of the results of the diagnostic criteria adopted: (1) confirmed leptospirosis cases (42/89 dogs); (2) negative leptospirosis cases (36/89 dogs); and (3) unconfirmed leptospirosis cases (11/89 dogs). The results supported the usefulness of the two rapid diagnostic tests as a first in-clinic screening tool for suspected leptospirosis; positive results in the in-clinic tests in dogs with suggestive clinical and laboratory signs strongly indicated acute leptospirosis, while negative results required additional diagnostic investigation to exclude the infection. Confirmatory tests recommended for canine leptospirosis are still necessary in addition to the use of rapid in-clinic tests.

Novel sequence variants of viral hexon and fibre genes in two dogs with canine adenovirus type 1-associated disease.

There is little information on sequence variation of canine adenovirus type 1 (CAdV-1), the aetiological agent of infectious canine hepatitis (ICH). This study reports hexon and fibre gene sequence variants of CAdV-1 in a dog with systemic ICH and a dog with the ocular form of the disease ('blue eye') in Northern Italy in 2013. One of the sequence variants matched a CAdV-1 fox sequence previously detected in Italy.

Investigation of the presence of canine adenovirus (CAdV) in owned dogs in Northern Italy.

The use of a modified live canine adenovirus (CAdV) vaccine has greatly reduced the incidence of infectious canine hepatitis (ICH) in dogs. Nevertheless, cases of CAdV type 1 and 2 (CAdV-1 and CAdV-2) infection have been recently reported posing questions about the epidemiological situation of CAdV in dogs. In order to assess the presence of CAdV, samples from 51 dogs presented at a Veterinary Teaching Hospital in Bologna, Italy, for reasons unrelated with CAdV infection, were tested with a polymerase chain reaction (PCR) assay for CAdV. Thirty dogs (58.8%) were PCR positive for CAdV-2 infection and four of them (7.8%) were positive for CAdV-1. Sequence analysis performed on the obtained PCR products suggests that a genetically stable CAdV-1 strain and different CAdV-2 strains circulate in the canine population examined and that coinfections are relatively frequent.

MiR203 mediates subversion of stem cell properties during mammary epithelial differentiation via repression of ΔNP63α and promotes mesenchymal-to-epithelial transition.

During reproductive life, the mammary epithelium undergoes consecutive cycles of proliferation, differentiation and apoptosis. Doing so relies on the retained proliferative capacity, prolonged lifespan and developmental potency of mammary stem cells (MaSCs). ΔNp63α, the predominant TP63 isoform in mammary epithelia, is robustly expressed in MaSCs and is required for preservation of self-renewing capacity in diverse epithelial structures. However, the mechanism(s) underlying subversion of this activity during forfeiture of self-renewing capacity are poorly understood. MicroRNAs (miRNAs) govern critical cellular functions including stem cell maintenance, development, cell cycle regulation and differentiation by disrupting translation of target mRNAs. Data presented here indicate that expression of miR203, a miRNA that targets ΔNp63α and ΔNp63ß is activated during luminal epithelial differentiation and that this pattern is observed in the murine mammary hierarchy. In addition, we present evidence that the transcription factor Zeb1 represses miR203 expression, thus enhancing ΔNp63α protein levels. Furthermore, ectopic miR203 suppresses ΔNp63α expression, proliferation and colony formation. The anti-clonogenic effects mediated by miR203 require suppression of ΔNp63α. In addition, ectopic miR203 promotes mesenchymal-to-epithelial transition and disrupts activities associated with epithelial stem cells. These studies support a model in which induction of miR203 mediates forfeiture of self-renewing capacity via suppression of ΔNp63α and may also have anti-tumorigenic activity through its reduction of EMT and cancer stem cell populations.

AGR2 is a SMAD4-suppressible gene that modulates MUC1 levels and promotes the initiation and progression of pancreatic intraepithelial neoplasia.

The mechanisms controlling expression of the putative oncogene Anterior gradient 2 (AGR2) in pancreatic ductal adenocarcinoma (PDAC) are not well understood. We now show that AGR2 is a transforming growth factor-ß (TGF-ß)-responsive gene in human pancreatic cancer cells, whose downregulation is SMAD4 dependent. We also provide evidence supporting a role for AGR2 as an ER-chaperone for the cancer-associated mucin, MUC1. AGR2 is both sufficient and required for MUC1 expression in pancreatic cancer cells. Furthermore, AGR2 is coexpressed with MUC1 in mouse pancreatic intraepithelial neoplasia (mPanIN)-like lesions and in the cancer cells of four distinct genetically engineered mouse models of PDAC. We also show that Pdx1-Cre/LSL-Kras(G12D)/Smad4(lox/lox) mice heterozygous for Agr2 exhibit a delay in mPanIN initiation and progression to PDAC. It is proposed that loss of Smad4 may convert TGF-ß from a tumor suppressor to a tumor promoter by causing the upregulation of AGR2, which then leads to increased MUC1 expression, at which point both AGR2 and MUC1 facilitate mPanIN initiation and progression to PDAC.

Detection of a virus related to betacoronaviruses in Italian greater horseshoe bats.

The association between coronaviruses and bats is a worldwide phenomenon and bats belonging to genus Rhinolophus are the reservoir host for several coronaviruses, including a large number of viruses closely related genetically to severe acute respiratory syndrome-coronavirus (SARS-CoV). We carried out a survey in colonies of Italian bats (Rhinolophus ferrumequinum) for the presence of coronaviruses. Two of 52 R. ferrumequinum captured from different Italian areas tested positive by reverse transcription-PCR for a fragment of RNA-dependent RNA polymerase (RdRp) gene of viruses related to Coronavirus. Phylogenetic analysis revealed close correlations between one of the positive samples and SARS-related CoV belonging to the genus Betacoronavirus.

Association between HLA genes and susceptibility to toluene diisocyanate-induced asthma.

BACKGROUND: Only a small proportion of subjects exposed to isocyanates develop occupational asthma, suggesting individual predisposition. The human leucocyte antigen (HLA) class II molecules may play a crucial role in the development of the immune response to isocyanates. OBJECTIVES: To investigate the role of HLA class II molecules in the development of toluene diisocyanate (TDI)-induced asthma. SUBJECTS: Sixty-seven asthmatic subjects and 27 asymptomatic exposed subjects (controls) were typed at the HLA class II DQA1, DQB1 and DRB1 loci by polymerase chain reaction (PCR)-based techniques. RESULTS: The frequencies of DQA1*0104 and DQB1*0503 were significantly increased in asthmatics compared with asymptomatic exposed subjects, while DQA1*0101 and DQB1*0501 were significantly increased in asymptomatic exposed subjects. No significant difference was found in the distribution of DRB1 alleles between asthmatics and controls. CONCLUSIONS: The results of this study indicate that HLA-regulated immune mechanisms are involved in TDI-induced asthma and that, in exposed subjects, specific factors may increase or decrease the risk of developing disease.

Human leukocyte antigen associations in occupational asthma induced by isocyanates.

Exposure to diisocyanates is recognized as a leading cause of occupational asthma. Occupational asthma induced by isocyanates shares many characteristics with immunoglobulin E (IgE)-mediated asthma: in both, the responsible agent is known, and the clinical presentation, response to inhalation challenge in the laboratory, and response to antiasthma drugs are similar. Although asthma mediated by an IgE mechanism occurs in atopic subjects, occupational asthma induced by isocyanates occurs mostly in nonatopic asthmatics, and an IgE-mediated mechanism has not been consistently demonstrated. However, activated T lymphocytes, methacromatic cells, and eosinophils are increased in the bronchial mucosa of allergic and nonallergic asthmatics and subjects with occupational asthma induced by isocyanates, suggesting similar, probably immunologically mediated mechanisms for both nonoccupational and occupational asthma. Occupational asthma occurs in up to 5-10% of the exposed subjects. Evaluation of major histocompatibility complex (MHC) class II genes in exposed subjects who develop toluene diisocyanate (TDI) asthma has shown a negative association with HLA-DQB1*0501 and a positive association with HLA-DQB1*0503 alleles. In addition, a high proportion of TDI asthmatics express the HLA-DQB1*0503-associated aspartic acid at residue 57, suggesting that HLA-DQ may have a key role in conferring susceptibility. Thus, asthma induced by the low-molecular-weight agent TDI may result from an immunologic reaction due to the interaction of genetic susceptibility with exposure in the workplace.

Association between toluene diisocyanate-induced asthma and DQB1 markers: a possible role for aspartic acid at position 57.

Toluene diisocyanate (TDI) is the most common cause of occupational asthma in western countries. The aim of this study was to investigate whether genetic factors are involved in toluene diisocyanate-induced asthma. We studied the frequency of human leucocyte antigen (HLA) class II genetic markers in three groups of subjects: 1) subjects with TDI-induced asthma (n = 30); 2) exposed subjects with no history of TDI-induced asthma (n = 12); and 3) normal subjects not exposed to TDI (n = 126). Venous blood samples were collected from the three groups and the polymorphic second exon of DQA and DQB genes was amplified by the polymerase chain reaction (PCR) method. Evaluation of HLA class II gene products in TDI-induced asthma cases showed a positive association with HLA-DQB1 * 0503 and a negative association with HLA-DQB1 * 0501 alleles, which differed at residue 57 for a single amino acid, i.e. aspartic acid in DQB1 * 0503 and valine in DQB1 * 0501. No significant difference was found in the distribution of DQA1 alleles between asthmatics and controls. Our results confirm the hypothesis that HLA-DQB1 * 0503 has a role in conferring susceptibility to TDI-induced asthma and that residue 57 of HLA-DQB1 is a potentially critical location.

Identification of a novel protein kinase C inhibitor in microsomes from phytohaemagglutinin activated human peripheral blood mononuclear cells.

A peptide inhibiting either corpuscolate or purified PKC has been identified from microsomes of PHA-activated human PBMC but it is not detectable in microsomes of resting PBMC. The peptide was obtained from a microsomal preparation in an oligomeric form that could be transformed into a monomeric form by beta-MSH. The active peptide (IN) was retained on a PC-11 chromatographic column and could be eluted with NaCl. IN is ineffective on PKC-dependent protamine phosphorylation of protamine and on Ca2+ and phospholipid-independent activity generated by mild hydrolysis with trypsin of PKC. Ca2+ binding is permissive for IN activity. IN inhibits particulate PKC in PHA-activated PBMC, but is ineffective after TPA activation. All these data indicate that IN acts at the regulatory domain of PKC.

Anchored anti-HLA class I monoclonal antibody fails to induce inhibition of PHA-activated lymphocytes proliferation.

It is known that anti-HLA Class I antibodies inhibit the proliferative response of PHA-activated T-lymphocytes. We found that plastic- or sepharose-linked anti-HLA Class I monoclonal antibody 01.65 does not inhibit either [3H]Thymidine incorporation or recruitment in the cell cycle, nor does it reduce the expression of c-myc mRNA and the membrane expression of Interleukin-2 Receptor and Transferrin Receptor. Furthermore, particulate Protein Kinase C is not affected by anchored anti-HLA Class I monoclonal antibody 01.65. We suggest that anti-HLA Class I monoclonal antibody may act through crosslinking or internalization of HLA Class I antigens.

Altered proliferative kinetics in PHA-activated human T-lymphocytes treated with the anti-HLA class I monoclonal antibody 01.65.

Anti-HLA class I monoclonal antibody (mAb) 01.65 inhibited phytohaemagglutinin (PHA)-induced human lymphocyte proliferation. The inhibitory effect was inversely correlated to the strength of the proliferative response. It was increased when lymphocytes were stimulated with suboptimal doses of PHA but it disappeared with supraoptimal doses. Proliferation inhibition was achieved by prolonging the cell cycle time and by slowing down its recruitment rate. The former effect was not restricted to the G1-phase but also included the S phase. These results support the idea that HLA class I molecules are important in the PHA-induced proliferation of human T-lymphocytes.