Allestimento di strumenti per lo studio e il controllo delle infezioni sostenute da Micoplasmi

Mycoplasmas are the causal agent of arthritis in mammals, in fact lipoproteins and lipopeptides are Pathogen Associated Molecular Patterns (PAMPs) that interact with TLR causing release of pro-inflammatory cytokines and chemokines. Fibroblast-like synoviocytes (FLS) play a key role in pathogenesis of arthritis causing cytokines release that perpetuates inflammation. This study evaluate the ability of synthetic lipopeptide LipoP48 from M. agalactiae-derived lipoprotein P48 to trigger innate immunity using an in vitro approach, by qPCR for cytokines/chemokines expression (IL1β, TNF-α, IL6, IL8, GMCSF, Mip1β), and by immunofluorescence for MHCII, TLR2, CD80 and CD86 in ovine FLS. FLS stimulated with increasing concentrations of LipoP48 showed a dose-dependent expression of TNF-α, IL-6, IL-8 and GM-CSF. High levels of IL6, IL8 and GMCSF were found at different time point. Post stimulation, an increased expression of TLR2 and MHCII, CD80 and CD86 expression were induced. Neither stimulation with the peptide sequence nor with synthetic molecule pam2cys have been successful. Lack of MHCII expression induced by LipoP48 was observed when TLR2 have been blocked. Results show that LipoP48 acts as a potent immunomodulator, triggering the production of pro-inflammatory cytokines in ovine FLS. These data suggest a role of mycoplasmas PAMPs in the onset of the articular inflammatory process, and propose a useful model for the study of arthritis pathogenesis of bacterial etiology in mammals.</br

Peripheral nerve sheath myxoma. Clinicopathological and immunohistochemical study of a morphologically distinctive myxoid peripheral nerve sheath tumor in the forelimb of a cat

Peripheral nerve sheath tumors (PNST) are a class of nervous system tumors which arise in both schwann cells and perineural fibroblasts. Benign and malignant PNSTs are reported to occur in all domestic animals. In cats they represent 3% of all cutaneous and subcutaneous neoplasms. Only in dogs mixoid PNST has been observed generally localized in the fingers. In humans, PNSTs are rare neoplasms, and nerve sheath myxomas are a distinct neoplasia most commonly found in limb extremities

Evaluation of anatomical and histopathological changes in target organs of cattle slaughtered in Sardinia as a result of the illegal use of growth hormones. Preliminary results

Within the bovine specie, illegal use of anabolic agents can be grouped into four categories: beta-agonists, thyrostatics, glucocorticoids, sexual steroids. These substances, further their anabolic effect, cause morphological changes in target organs which can be evidenced by anatomical and histopathological testing. Such investigations are extremely important to screen and to detect in advance groups of animals in risk-breeding

Un Infrequente caso di "spindle cell tumor" a localizzazione splenica primaria in un cane: rilievi citologici ed istopatologici, osservazioni istochimiche ed immunoistochimiche

Tutti i tumori fusocellulari derivano verosimilmente da una unica cellula staminale mesenchimale pluripotente. Macroscopicamente e microscopicamente i tumori delle cellule muscolari lisce e i tumori del tessuto connettivo fibroso, si presentano morfologicamente simili. Abbiamo ritenuto interessante riferire su un cane meticcio, maschio, di quattro anni sottoposto ad eutanasia dopo aver mostrato letargia, anoressia e perdita di peso. Alla necroscopia si osservava una voluminosa neoformazione splenica di colore biancastro, dura e di consistenza fibrosa. Il fegato si presentava notevolmente aumentato di volume con numerose neoformazioni di piccole e medie dimensioni. Diversi campioni appartenenti agli organi sopramenzionati, venivano sottoposti ad una prima osservazione citologica. Parte di questi, dopo fissazione in formalina venivano inclusi in paraffina. Sezioni di 3-5 μ erano sottoposte a colorazioni ordinarie e speciali (H. E., tricromica di Mallory Azan e Van Gieson). Si eseguivano prove immunoistochimiche utilizzando il seguente algoritmo anticorpale: anti pan-citocheratine, anti vimentina, anti fattore VIII, anti-αactina L, anti desmina, anti CD 15, anti S-100. La neoplasia, altamente infiltrante, era costituita da ampi fasci intrecciati di cellule allungate fusiformi, fibroblastosimili, a moderato indice mitotico, disposte in maniera ordinata, a spina di pesce. Esse risultavano positive alla vimentina e alla αactina L e negative alla desmina. Dette caratteristiche ci consentivano di formulare diagnosi di fibrosarcoma, neoplasia a infrequente localizzazione splenica primaria, e di identificare come metastasi della stessa, le neoformazioni riscontrare a livello epatico

Histopathological, histochemical and immunoistochemical study on a rare case of granulocytic sarcoma in a dog

Granulocytic sarcoma is a malignant extramedullary solid tumor, composed of granulocytic precursor cells at various levels of differentiation. Three differentiation levels are considered: blastic, immature, and differentiated, and cases with unusual morphology. Nowadays the aid of more and more highly developed techniques allows to differentiate the various histotypes. We report a case of a 5 year-old female Schnautzer dog, died after a serious dyspnoea. During the autopsy the veterinary found several neoplasias in the lungs. It has been found a very large white-greyish mediastinal neoplasia, stuck at the trachea and several more in the parenchyma. Several samples were fixed in formalin, embedded in paraffin and exposed to histochemistry staining (H.E., Giemsa), cytochemistry (Naphthol-AS-D-Chloroacetate), and immunohistochemistry (anti-CD3, -CD79a, -CD45, -MPO, -CD45Ro, -CD34, -CD20, -CD68, -CD15, -CD30, -CD117, -CD235a, -Factor VIII, -elastase and anti-Pan-cytocheratine). Neoplasia, poorly circumscribed, was composed by a large number of neutrophil granulocytes with different degrees of differentiation, including elements of myeloid lineage. Cells were positive to MPO and focal to Naphtol-As-D-Chloroacetate and were negative to all the others antisera, allowing us to exclude lymphomas, small cells carcinomas, and tumors of monocytic and erythroid origin. By these characteristics we could diagnose a rare case of neutrophilic granulocytic sarcoma, of immature type progressing to mature form

Epidemiology and genetic characterization of <i>Border Disease Virus</i> circulating in Sardinia

Border Disease Virus (BDV), a pestivirus from the Flaviviridae family, is an important pathogen of sheep and goats responsible for significant losses in farms around the world. In spite of the relevance of this pathogen there are only a few epidemiological studies on BDV infection and, as a consequence, the economic impact on small ruminant productions is probably under-estimated. The aims of this study are i) to determine the distribution of BDV in small ruminant farms in Sardinia and genetically characterize circulating strains ii) analyze the relation between seroprevalence, Somatic Cells Count (SCC) an milk yeld. ELISA was performed using “BVDV/MD/BDV p80 Protein Antibody Test Kit” (IDEXX) on serum of bulk tank milk (BTM) samples collected from Sardinian sheep flocks and goat herds between spring 2014 and 2015. The number of sampled farms corresponded to 8.5% of all registered farms in Sardinia. RNA was isolated using Qiamp Viral RNA mini kit from the cellular fraction of each ELISA positive BTV sample and amplified by rt-PCR using complementary primers to a highly conserved region in the untranslated regions (UTRs) of the viral genome. The amplicons were sequenced for phylogenetic analysis. Geographic distribution of collected specimen, seroprevalence and virological positive samples were analyzed via GIS (ESRI ARCGIS 10.3). ELISA screening shows a seroprevalence of 8.3% among goat farms and 10.5% among ovine farms. Ten from the ELISA positive samples were found rt-PCR positive. The sequence analysis indicates that all the amplified samples match with BDV genomes and the phylogenetic analysis revealed that all the viruses clustered in the same group classified as BDV-7. BDV-7 is the only group isolated in Sardinia so far

Pathology of sea turtles <i>Caretta caretta</i> found on the coast of Apulia (south Italy)

On the basis of the most recent scientific literature, little is known on sea turtle renal pathology, as most published data mainly regard tortoises. The authors examined 49 carcasses of sea turtles belonging to the species Caretta caretta, stranded on the coasts of Apulia (South Italy). The subjects were classified by species and gender, weighed, morphometrical assessed, and submitted to pathological and bacteriological examinations

Development and In-House Validation of an Enzyme-Linked Immunosorbent Assay and a Lateral Flow Immunoassay for the Dosage of Tenofovir in Human Saliva

Highly active antiretroviral therapy (HAART) includes very potent drugs that are often characterized by high toxicity. Tenofovir (TFV) is a widely used drug prescribed mainly for pre-exposure prophylaxis (PreP) and the treatment of human immunodeficiency virus (HIV). The therapeutic range of TFV is narrow, and adverse effects occur with both underdose and overdose. The main factor contributing to therapeutic failure is the improper management of TFV, which may be caused by low compliance or patient variability. An important tool to prevent inappropriate administration is therapeutic drug monitoring (TDM) of compliance-relevant concentrations (ARCs) of TFV. TDM is performed routinely using time-consuming and expensive chromatographic methods coupled with mass spectrometry. Immunoassays, such as enzyme-linked immunosorbent assays (ELISAs) and lateral flow immunoassays (LFIAs), are based on antibody&ndash;antigen specific recognition and represent key tools for real-time quantitative and qualitative screening for point-of-care testing (POCT). Since saliva is a non-invasive and non-infectious biological sample, it is well-suited for TDM. However, saliva is expected to have a very low ARC for TFV, so tests with high sensitivity are required. Here, we have developed and validated a highly sensitive ELISA (IC50 1.2 ng/mL, dynamic range 0.4&ndash;10 ng/mL) that allows the quantification of TFV in saliva at ARCs and an extremely sensitive LFIA (visual LOD 0.5 ng/mL) that is able to distinguish between optimal and suboptimal ARCs of TFV in untreated saliva

Characterization of the interaction of African swine fever virus with monocytes and derived macrophage subsets.

Abstract African swine fever (ASF) is a devastating disease for which there is no vaccine available. The ASF virus (ASFV) primarily infects cells of the myeloid lineage and this tropism is thought to be crucial for disease pathogenesis. A detailed in vitro characterization of the interactions of a virulent Sardinian isolate (22653/14) and a tissue culture adapted avirulent strain (BA71V) of ASFV with porcine monocytes, un-activated (moMΦ), classically (moM1) and alternatively (moM2) activated monocyte-derived macrophages was conducted in an attempt to better understand this relationship. Using a multiplicity-of-infection (MOI) of 1, both viruses were able to infect monocytes and macrophage subsets, but BA71V presented a reduced ability to infect moM1 compared to 22653/14, with higher expression of early compared to late proteins. Using an MOI of 0.01, only 22653/14 was able to replicate in all the macrophage subsets, with initially lowest in moM1 and moM2. No differences were observed in the expression of CD163 between ASFV infected and uninfected bystander cells. ASFV down-regulated CD16 expression but did not modulate MHC class II levels in monocytes and macrophage subsets. BA71V-infected but not 22653/14-infected moMΦ and moM2 presented with a reduced expression of MHC class I compared to the mock-infected controls. Higher levels of IL-18, IL1-β and IL-1α were released from moM1 after infection with BA71V compared to 22653/14 or mock-infected control. These results revealed differences between these ASFV strains, suggesting that virulent isolates have evolved mechanisms to counteract activated macrophages responses, promoting their survival, dissemination in the host and so ASF pathogenesis

Circulating tumor cells as early predictors of metastatic spread in breast cancer patients with limited metastatic dissemination.

IntroductionTraditional factors currently used for prognostic stratification do not always predict adequately treatment response and disease evolution in advanced breast cancer patients. Therefore, the use of blood-based markers, such as circulating tumor cells (CTCs), represents a promising complementary strategy for disease monitoring. In this retrospective study, we explored the role of CTC counts as predictors of disease evolution in breast cancer patients with limited metastatic dissemination.Methods492 advanced breast cancer patients who had a CTC count assessed by CellSearch prior to starting a new line of systemic therapy were eligible for this analysis. Using the threshold of 5 cells/7.5 mL of blood, pretreatment CTC counts were correlated in the overall population with metastatic site distribution, evaluated at baseline and at the time of treatment failure, using the Fisher¿s Exact test. Time to visceral progression, as well as, time to the development of new metastatic lesions and sites were estimated in patients with non-visceral metastases and with single-site metastatic disease, respectively, by the Kaplan-Meier method. Survival times were compared among groups according to pretreatment CTC count by log-Rank test.ResultsIn the overall population, pretreatment CTCs¿¿¿5 were associated with increased baseline number of metastatic sites, compared with CTCs