Single nucleotide polymorphisms detected and in silico analysis of the 5' flanking sequence and exon 1 in the Bubalus bubalis leptin gene

In this study, we have sequenced the 5' flanking region and exon 1 of the leptin gene in buffalo, and have detected eight single nucleotide polymorphisms; we have made evidence, through in silico analysis, that many of them fall within putative binding sites for transcription factors. Starting from the bovine whole genome shotgun sequence, that encodes the complete sequence of the leptin gene, we had designed primers to amplify two amplicons, so to cover the 5' flanking and exon 1 of the leptin gene of 41 non related buffalos. The newly sequenced buffalo fragment was submitted to profile search for transcription factor binding sites, using the MATCH program, focusing on the areas where the single nucleotide polymorphisms had been detected. Our analysis shows that the majority of the identified single nucleotide polymorphisms fall into the core sequence of transcription factor binding sites that regulate the expression of target genes in many physiological processes within mammalian tissues. Because the leptin gene plays an important role in influencing economic traits in cattle, the novel detected single nucleotide polymorphisms might be used in association studies to assess their potential of being genetic markers for selection

Assessment of Poly(ADP-ribose) Polymerase1 (PARP1) expression and activity in cells purified from blood and milk of dairy cattle

Poly(ADP-ribosyl)ation (PAR) is a post-translational protein modification catalysed by enzyme member of the poly(ADP-ribose) polymerases (PARPs) family. The activation of several PARPs is triggered by DNA strand breakage and the main PARP enzyme involved in this process is PARP1. Besides its involvement in DNA repair, PARP1 is involved in several cellular processes including transcription, epigenetics, chromatin re-modelling as well as in the maintenance of genomic stability. Moreover, several studies in human and animal models showed PARP1 activation in various inflammatory disorders. The aims of the study were (1) to characterize PARP1 expression in bovine peripheral blood mononuclear cells (PBMC) and (2) to evaluate PAR levels as a potential inflammatory marker in cells isolated from blood and milk samples following different types of infection, including mastitis. Our results show that (i) bovine PBMC express PARP1; (ii) lymphocytes exhibit higher expression of PARP1 than monocytes; (iii) PARP1 and PAR levels were higher in circulating PBMCs of infected cows; (iv) PAR levels were higher in cells isolated from milk with higher Somatic Cell Counts (SCC > 100,000 cells/mL) than in cells from milk with low SCCs. In conclusion, these findings suggest that PARP1 is activated during mastitis, which may prove to be a useful biomarker of mastitis

New insights into the significance of PARP-1 activation: flow cytometric detection of poly(ADP-ribose) as marker of bovine intramammary infection

Bovine intramammary infections are common diseases affecting dairy cattle worldwide and represent a major focus of veterinary research due to financial losses and food safety concerns. The identification of new biomarkers of intramammary infection, useful for monitoring dairy cows’ health and wellness verification, represents a key advancement having potential beneficial effects on public health. In vitro experiments, using bovine peripheral blood mononuclear cells (PBMC) stimulated with the bacterial endotoxin lipopolysaccharide (LPS) allowed to perform a flow cytometric assay to evaluate in vivo poly-ADP-ribose (PAR) levels. Results showed a significant increase of PAR after 1h of treatment, which is consistent with the involvement of PARP activity in the inflammatory response. This study investigated PARP-1 activation in leukocyte subpopulations from bovine milk samples during udder infection. A flow cytometric assay was therefore performed to evaluate the PAR content on milk leukocytes subsets of cows with and without intramammary infection (IMI). Results showed that milk lymphocytes and macrophages isolated from cows with IMI had a significant increase of PAR content compared to uninfected samples. These results suggest mastitis as a new model for the study of the role of PARP in zoonotic inflammatory diseases, thereby opening new horizons for the "One Health" perspective connecting animal and human health

Intradermal Tuberculin Test in Water Buffalo (Bubalus bubalis): Experimental use of Mycobacterial Antigens for the Diagnosis of Bovine Tuberculosis

The study aims to evaluate the potential use of mycobacterial ESAT6 and CFP10 antigens, Early Secretory Proteins (ESP) in the Skin Test used for bovine tuberculosis (TB) diagnosis in Water Buffalo. A pilot study was performed on 21 buffaloes from a TB outbreak and 11 buffaloes from a TB-free herd. Three concentrations of ESAT6-CFP10 (10, 20, and 30 mg) and two of ESP (50 and 100 µg) were inoculated in the Skin Test, along with PPDB, PPDA, and PBS as a negative control. Skin thickness was measured with calipers before the test and every 24 hours for 4 days. Then, to evaluate the specificity of the antigens, a field study was conducted, and 100 buffaloes from a TB-free herd were inoculated using the best antigens concentration derived from the pilot study. In the positive buffaloes, the strongest skin response was to PPDB at 24h, with some subjects becoming inconclusive at 72 and 96 h. A peak response to PPDA at 48 hours was detected, followed by a slight decrease. The response to ESP-100 µg remained high at 24 and 48 h, then decreased, remaining positive at 72 h. In the 100 TB-free buffaloes, the best specificity was observed using ESAT6-CFP10 and ESP. ESP yielded the best results, showing higher reactivity in infected animals and no reactivity in the healthy ones at 72 h. Therefore, ESP could be an excellent candidate for further extensive studies in the buffalo species to improve Skin Test performance

Comparison of metabolic, oxidative and inflammatory status of Simmental × Holstein crossbred with parental breeds during the peripartal and early lactation periods.

AbstractThe aim of the research reported in this paper was to evaluate plasma concentrations of energy, oxidative and inflammatory biomarkers of Simmental (sire) × Holstein (dam) crossbred cows, in comparison with the two parental breeds during the peripartal and early lactation periods and to estimate the effects of heterosis for these traits. Thirty-three animals, managed under the same conditions, 8 Simmental (SI), 9 Holstein (HO) and 16 crossbred (CR) cows were enrolled in this study. Glucose, non-esterified fatty acids (NEFA), β-hydroxybutyrate (BHB), total bilirubin, aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), creatine kinase (CK), total protein, albumin, creatinine and urea were determined in blood sampled at six different time points (30 ± 3 and 15 ± 3 d before the expected calving date, at calving and 15, 30 and 60 d after calving). Furthermore, derived reactive oxygen metabolites (d-ROMs), biological antioxidant potential (BAP), interleukin-6 (IL-6), haptoglobin (Hp) and serum amyloid A protein (SAA) were determined to evaluate inflammatory and oxidative status. Results showed that the CR group had significantly lower average values of glucose and NEFA when compared to HO group; signifcantly lower values of urea than SI group and significantly higher values of creatinine than HO. Furthermore, CR cows showed the lowest average value of d-ROMs with respect to SI and HO parental breeds. Finally, the average value of haptoglobin was significantly lower in CR and HO groups, when compared to SI group. As for the heterosis we found the highest (positive) percentage for CK (98%) and BAP (47%) and the lowest (negative) percentage for OSi (−75%) and d-ROMs (−39%). A negative percentage was also found for the glucose (−11%) and NEFA (−20%) toward the Simmental parental breed. Our results suggest a different response among the three genetic groups during the peripartal and early lactation periods. In particular, CR and SI cows seem more adaptable regarding energy metabolism and oxidative status. Heterosis led to a positive effect on those parameters in Simmental (sire) × Holstein (dam) crossbred cows F1 population (50% Simmental and 50% Holstein)

A Guideline for Implementing a Robust Optimization of a Complex Multi-Stage Manufacturing Process

In the industrial production scenario, the goal of engineering is focused on the continuous improvement of the process performance by maximizing the effectiveness of the manufacturing and the quality of the products. In order to address these aims, the advanced robust process optimization techniques have been designed, implemented, and applied to the manufacturing process of ultrasound (US) probes for medical imaging. The suggested guideline plays a key role for improving a complex multi-stage manufacturing process; it consists of statistical methods applied for improving the product quality, and for achieving a higher productivity, jointly with engineering techniques oriented to problem solving. Starting from the Six Sigma approach, the high definition of the production process was analyzed through a risk analysis, and thus providing a successful implementation of the PDCA (plan-do-check-act) methodology. Therefore, the multidisciplinary analysis is carried out by applying statistical models and by detecting the latent failures by means of NDT (non-destructive testing), i.e., scanning acoustic microscopy (SAM). The presented approach, driven by the statistical analysis, allows the engineers to distinguish the potential weak points of the complex manufacturing, in order to implement the corrective actions. Furthermore, in this paper we illustrate this approach by considering a pilot study, e.g., a process of US probes for medical imaging, by detailing all the guideline steps

Il supporto satellitare dell'Europa per le emergenze

Satellite plays an imported role in emergency management and is the basis of e-GEOS activities under the emergency support services. In Europe this instrument was inserted as the Copernicus EMS operating assets, which supports the different countries in the first phase of mapping the territory after a disaster and monitors the phases immediately after the engagement