The bovine acute phase protein α1-acid glycoprotein (AGP) can disrupt Staphylococcus aureus biofilm

Staphylococcus aureus biofilm-related infections are of clinical concern due to the capability of bacterial colonies to adapt to a hostile environment. The present study investigated the capability of the acute phase protein alpha 1-acid glycoprotein (AGP) to a) disrupt already established S. aureus biofilm and b) interfere with the biofilm de novo production by using Microtiter Plate assay (MtP) on field strains isolated from infected quarters by assessing. The present study also investigated whether AGP could interfere with the expression of bacterial genes related to biofilm formation (icaA, icaD, icaB, and icaC) and adhesive virulence determinants (fnbA, fnbB, clfA, clfB, fib, ebps, eno) by quantitative real-time PCR (qPCR). The results provided the evidence that AGP could disrupt the biofilm structure only when it was already developed, but could not prevent the de novo biofilm formation. Moreover, AGP could interfere with the expression levels of genes involved in biofilm formation in a dose- and strain-dependent way, by upregulating, or downregulating, icaABC genes and fnbB, respectively. The results presented in this study provide new insights about the direct antibacterial activity of AGP in bovine milk. It remains to be demonstrated the molecular bases of AGP mechanism of action, in particular for what concerns the scarce capability to interact with the de novo formation of biofilm

Effects of nucleotides administration on growth performance and immune response of post-weaning piglets

The aim of this study was to assess the effect of nucleotides administration on growth perform- ance and immune response in post-weaning piglets. Twenty-eight male weaned piglets, homo- geneous for age and weight were randomly allocated to two experimental treatments. Treated group (T) was daily orally administered 0.8g/head of a mixture of nucleotides suspended in 2.1 mL water solution; while control group (C) received 2.1 mL saline solution. Body weight (BW) and average daily gain (ADG) were individually recorded weekly, while feed intake (FI), and gain:feed (G:F) were recorded and calculated on pen basis. Faecal score was evaluated every seven days. On day 0, 9, 18 and 27 blood samples were collected to determine IgA, IgG and haptoglobin concentration. At day 28 all piglets were sacrificed, and tissue samples of ileal Peyer\u2019s patches were collected for the evaluation of IL1a, IL1b, IL6, IL10, TNFa, TLR2, TLR4 and PPARc gene expression. Nucleotides supplementation significantly increased BW (17.37 vs. 19.00kg/pig; p 1\u20444 <.01), ADG (.351 vs. .400kg/d; p < .01), and FI (3.96 vs. 4.39kg/d; p < .01), but not G:F (.61 vs. .64; p 1\u20444 .29). Faecal consistency was not different between the experimental groups and no occurrence of diarrhoea was reported. IgA and IgG content in blood was not influenced by the treatment, as well as gene expression of inflammatory cytokines in Peyer\u2019s patches. The present trial shows that nucleotide administration is able to improve growth per- formance of post-weaning piglets, with no effects on inflammatory response and the expression of immune-related genes

In-vitro effect of heat stress on bovine monocytes lifespan and polarization

Heat stress (HS) has a negative impact on dairy cows’ health, milk production, reproductive performance and immune defenses. Cellular and molecular responses to high temperatures in bovine polymorphonuclear cells and peripheral blood mononuclear cells (PBMCs) have been investigated so far. On the contrary, the effects of high temperatures on isolated monocytes remain almost undisclosed. The aim of this study was to unravel the in vitro effects of high temperatures, simulating a severe HS related body hyperthermia, on bovine lifespan and M1/M2 polarisation. The PBMCs were isolated from whole blood of 9 healthy dairy cattle. Monocytes were sorted by magnetic activated cell sorting and cultured over night at 39 °C (normothermia) or 41 °C (HS). Apoptotic rate and viability were assessed and mRNA abundance for heat shock proteins (HSPs), heat transcription factors (HSFs) and genes involved in monocyte/macrophage polarization (STAT1, STAT2, STAT3, STAT6, IL1β, TGF1β, IL-10, COX2) were quantified by qPCR. We found that apoptosis increased in monocytes exposed to 41 °C, as compared to control, while viability conversely decreased. HS increased the abundance of HSF1 and HSP70. The concomitant decrease of STAT1 and STAT2 and the increase of STAT6 genes abundance at 41 °C suggest, at transcriptional factors level, a polarization of monocytes from a classical activated M1 to a non-classically activated M2 monocytes. In conclusion, the exposure of bovine monocytes to high temperatures affects their lifespan as well as the abundance of genes involved in HS response and in monocyte/macrophages polarization phenotype, confirming that bovine immune response may be significantly affected by hyperthermia

The effects of superoxide dismutase-rich melon pulp concentrate on inflammation, antioxidant status and growth performance of challenged post-weaning piglets

Piglets can often suffer impaired antioxidant status and poor immune response during post-weaning, especially when chronic inflammation takes place, leading to lower growth rates than expected. Oral administration of dietary antioxidant compounds during this period could be a feasible way to balance oxidation processes and increase health and growth performance. The aim of the trial was to study the effects of an antioxidant feed supplement (melon pulp concentrate) that contains high concentration of the antioxidant superoxide dismutase (SOD) on inflammation, antioxidant status and growth performance of lipopolysaccharide (LPS) challenged weaned piglets. In total, 48 weaned piglets were individually allocated to four experimental groups in a 2 72 factorial design for 29 days. Two different dietary treatments were adopted: (a) control (CTR), fed a basal diet, (b) treatment (MPC), fed the basal diet plus 30 g/ton of melon pulp concentrate. On days 19, 21, 23 and 25 half of the animals within CTR and MPC groups were subjected to a challenge with intramuscular injections of an increasing dosage of LPS from Escherichia coli (serotype 0.55:B5) (+) or were injected with an equal amount of PBS solution ( 12). Blood samples were collected at the beginning of the trial and under the challenge period for interleukin 1\u3b2, interleukin 6, tumour necrosis factor \u3b1, haptoglobin, plasma SOD activity, total antioxidant capacity, reactive oxygen species, red blood cells and plasma resistance to haemolysis, and 8-oxo-7, 8-dihydro-2\u2019-deoxyguanosine. Growth performance was evaluated weekly. A positive effect of melon pulp concentrate was evidenced on total antioxidant capacity, half-haemolysis time of red blood cells, average daily gain (ADG) and feed intake, while LPS challenge increased pro-inflammatory cytokines and haptoglobin serum concentrations, with a reduced feed intake and gain : feed (G : F). The obtained results show that oral SOD supplementation with melon pulp concentrate ameliorates the total antioxidant capacity and the half-haemolysis time in red blood cell of post-weaning piglets, with positive results on growing performance

Short communication : Circulating extracellular miR-22, miR-155, and miR-365 as candidate biomarkers to assess transport-related stress in turkeys

MicroRNA (miRNA) have been identified in circulating blood and might have the potential to be used as biomarkers for several pathophysiological conditions. To identify miRNA that are altered following stress events, turkeys (Meleagris gallopavo) were subjected to 2 h of road transportation. The expression levels of five circulating miRNA, namely miR-22, miR-155-5p, miR-181a-3p, miR-204 and miR-365-3p, were detected and assessed by quantitative polymerase chain reaction using TaqMan\uae probes, as potential biomarkers of stress. The areas under the receiver operating characteristic curves were then used to evaluate the diagnostic performance of miRNA. A panel of three stress-responsive miRNA, miR-22, miR-155 and miR-365 were identified; their expression levels were significantly higher after road transportation and the area under the curve (AUC) were 0.763, 0.71 and 0.704, respectively. Combining the three miRNA a specificity similar to the one found for the three miRNA separately was found. The AUC of the weighted average of the three miRNA was 0.763. This preliminary study suggests that the expression levels of circulating miR-22, miR-155 and miR-365 are increased during transport-related stress and that they may have diagnostic value to discriminate between stressed- and unstressed animals

Characterization of circulating miRNA signature in water buffaloes (Bubalus bubalis) during Brucella abortus infection and evaluation as potential biomarkers for non-invasive diagnosis in vaginal fluid

Brucellosis is an infectious disease caused by bacteria from the Brucella genus that can be transmitted to humans through contact with infected animals or contaminated animal products. Brucellosis also causes financial losses in animal production. Ruminants are highly susceptible to brucellosis, and the causative agent water buffaloes (Bubalus bubalis) is Brucella abortus. Circulating microRNAs (miRNAs) are cropping up as promising biomarkers for several infectious diseases. The goals of this study were to characterize the serum miRNA signature associated with brucellosis in water buffaloes and investigate the miRNAs' potential use as biomarkers in vaginal fluids. Next Generation Sequencing was used to assess miRNA expression profiles in Brucella-positive and Brucella-negative blood sera; dysregulated miRNAs in blood serum and vaginal fluids were validated using RT-qPCR. ROC curves were generated to evaluate the diagnostic value of miRNAs for Brucella. GO and KEGG pathway enrichment analyses were exploited to investigate the biological functions of dysregulated miRNAs. The results showed that 20 miRNAs were modulated, of which, 12 were upregulated and 8 were downregulated. These findings were corroborated by RT-qPCR, and ROC curves indicated that the miRNAs can serve as potential biomarkers for Brucella. GO and KEGG pathway analyses pointed out that some of these miRNAs are related to immune response and apoptosis. These results provided an overview of miRNA expression profiles and highlighted potential biomarkers for Brucella infection in water buffaloes. We also demonstrated the potential of vaginal fluids in studies involving microRNA detection. Further functional and mechanistic studies of these miRNAs may improve our understanding of the biological processes involved in Brucella infection and host immune response

A three-gene signature marks the time to locoregional recurrence in luminal-like breast cancer

Background: Gene expression profiling (GEP)-based prognostic signatures are being rapidly integrated into clinical decision making for systemic management of breast cancer patients. However, GEP remains relatively underdeveloped for locoregional risk assessment. Yet, locoregional recurrence (LRR), especially early after surgery, is associated with poor survival. Patients and methods: GEP was carried out on two independent luminal-like breast cancer cohorts of patients developing early (≤5 years after surgery) or late (>5 years) LRR and used, by a training and testing approach, to build a gene signature able to intercept women at risk of developing early LRR. The GEP data of two in silico datasets and of a third independent cohort were used to explore its prognostic value. Results: Analysis of the first two cohorts led to the identification of three genes, CSTB, CCDC91 and ITGB1, whose expression, derived by principal component analysis, generated a three-gene signature significantly associated with early LRR in both cohorts (P value <0.001 and 0.005, respectively), overcoming the discriminatory capability of age, hormone receptor status and therapy. Remarkably, the integration of the signature with these clinical variables led to an area under the curve of 0.878 [95% confidence interval (CI) 0.810-0.945]. In in silico datasets we found that the three-gene signature retained its association, showing higher values in the early relapsed patients. Moreover, in the third additional cohort, the signature significantly associated with relapse-free survival (hazard ratio 1.56, 95% CI 1.04-2.35). Conclusions: Our three-gene signature represents a new exploitable tool to aid treatment choice in patients with luminal-like breast cancer at risk of developing early recurrence

Antiaggregatory activity in human platelets of potent antagonists of the P2Y1 receptor

Activation of the P2Y1 nucleotide receptor in platelets by ADP causes changes in shape and aggregation, mediated by activation of phospholipase C (PLC). Recently, MRS2500 (2-iodo-N6-methyl-(N)-methanocarba-2′-deoxyadenosine-3′,5′-bisphosphate) was introduced as a highly potent and selective antagonist for this receptor. We have studied the actions of MRS2500 in human platelets and compared these effects with the effects of two acyclic nucleotide analogues, a bisphosphate MRS2298 and a bisphosphonate derivative MRS2496, which act as P2Y1 receptor antagonists, although less potently than MRS2500. Improved synthetic methods for MRS2500 and MRS2496 were devised. The bisphosphonate is predicted to be more stable in general in biological systems than phosphate antagonists due to the non-hydrolyzable C–P bond. MRS2500 inhibited the ADP-induced aggregation of human platelets with an IC50 value of 0.95 nM. MRS2298 and MRS2496 also both inhibited the ADP-induced aggregation of human platelets with IC50 values of 62.8 nM and 1.5 μM, respectively. A similar order of potency was observed for the three antagonists in binding to the recombinant human P2Y1 receptor and in inhibition of ADP-induced shape change and ADP-induced rise in intracellular Ca2+. No substantial antagonism of the pathway linked to the inhibition of cyclic AMP was observed for the nucleotide derivatives, indicating no interaction of these three P2Y1 receptor antagonists with the proaggregatory P2Y12 receptor, which is also activated by ADP. Thus, all three of the bisphosphate derivatives are highly selective antagonists of the platelet P2Y1 receptor, and MRS2500 is the most potent such antagonist yet reported