A New ELISA Using the ANANAS Technology Showing High Sensitivity to diagnose the Bovine Rhinotracheitis from Individual Sera to Pooled Milk

Diagnostic tests for veterinary surveillance programs should be efficient, easy to use and, possibly, economical. In this context, classic Enzyme linked ImmunoSorbent Assay (ELISA) remains the most common analytical platform employed for serological analyses. The analysis of pooled samples instead of individual ones is a common procedure that permits to certify, with one single test, entire herds as "disease-free". However, diagnostic tests for pooled samples need to be particularly sensitive, especially when the levels of disease markers are low, as in the case of anti-BoHV1 antibodies in milk as markers of Infectious Bovine Rhinotracheitis (IBR) disease. The avidin-nucleic-acid-nanoassembly (ANANAS) is a novel kind of signal amplification platform for immunodiagnostics based on colloidal poly-avidin nanoparticles that, using model analytes, was shown to strongly increase ELISA test performance as compared to monomeric avidin. Here, for the first time, we applied the ANANAS reagent integration in a real diagnostic context. The monoclonal 1G10 anti-bovine IgG1 antibody was biotinylated and integrated with the ANANAS reagents for indirect IBR diagnosis from pooled milk mimicking tank samples from herds with IBR prevalence between 1 to 8%. The sensitivity and specificity of the ANANAS integrated method was compared to that of a classic test based on the same 1G10 antibody directly linked to horseradish peroxidase, and a commercial IDEXX kit recently introduced in the market. ANANAS integration increased by 5-fold the sensitivity of the 1G10 mAb-based conventional ELISA without loosing specificity. When compared to the commercial kit, the 1G10-ANANAS integrated method was capable to detect the presence of anti-BHV1 antibodies from bulk milk of gE antibody positive animals with 2-fold higher sensitivity and similar specificity. The results demonstrate the potentials of this new amplification technology, which permits improving current classic ELISA sensitivity limits without the need for new hardware investments

H2O2 production at gas-diffusion cathodes made from agarose-derived carbons with different textural properties for acebutolol degradation in chloride media

The excessive cost, unsustainability or complex production of new highly selective electrocatalysts for H2O2 production, especially noble-metal-based ones, is prohibitive in the water treatment sector. To solve this conundrum, biomass-derived carbons with adequate textural properties were synthesized via agarose doublestep pyrolysis followed by steam activation. A longer steam treatment enhanced the graphitization and porosity, even surpassing commercial carbon black. Steam treatment for 20 min yielded the greatest surface area (1248 m2 g-1), enhanced the mesopore/micropore volume distribution and increased the activity (E1/2 = 0.609 V) and yield of H2O2 (40%) as determined by RRDE. The upgraded textural properties had very positive impact on the ability of the corresponding gas-diffusion electrodes (GDEs) to accumulate H2O2, reaching Faradaic current efficiencies of ~95% at 30 min. Acidic solutions of β-blocker acebutolol were treated by photoelectro- Fenton (PEF) process in synthetic media with and without chloride. In urban wastewater, total drug disappearance was reached at 60 min with almost 50% mineralization after 360 min at only 10 mA cm-2. Up to 14 degradation products were identified in the Cl--containing medium

Detection of a fluorescent-labeled avidin-nucleic acid nanoassembly by confocal laser endomicroscopy in the microvasculature of chronically inflamed intestinal mucosa

Inflammatory bowel diseases are chronic gastrointestinal pathologies causing great discomfort in both children and adults. The pathogenesis of inflammatory bowel diseases is not yet fully understood and their diagnosis and treatment are often challenging. Nanoparticle-based strategies have been tested in local drug delivery to the inflamed colon. Here, we have investigated the use of the novel avidin-nucleic acid nanoassembly (ANANAS) platform as a potential diagnostic carrier in an experimental model of inflammatory bowel diseases. Fluorescent- labeled ANANAS nanoparticles were administered to mice with chemically induced chronic inflammation of the large intestine. Localization of mucosal nanoparticles was assessed in vivo by dual-band confocal laser endomicroscopy. This technique enables characterization of the mucosal microvasculature and crypt architecture at subcellular resolution. Intravascular nanoparticle distribution was observed in the inflamed mucosa but not in healthy controls, demonstrating the utility of the combination of ANANAS and confocal laser endomicroscopy for highlighting intestinal inflammatory conditions. The specific localization of ANANAS in inflamed tissues supports the potential of this platform as a targeted carrier for bioactive moieties in the treatment of inflammatory bowel disease

Antimicrobial treatment with the fixed-dose antibiotic combination RHB-104 for Mycobacterium avium subspecies paratuberculosis in Crohn\u2019s disease: pharmacological and clinical implications

Introduction: Crohn\u2019s disease (CD) is an inflammatory bowel disease of unknown etiology. However, increasing evidence suggests Mycobacterium avium subspecies paratuberculosis (MAP) as a putative causative agent: 1) MAP is the etiological agent of Johne\u2019s disease, a granulomatous enteritis affecting ruminants, which shares clinical and pathological features with CD; 2) MAP has been detected in tissues and blood samples from CD patients; 3) case reports have documented a favorable therapeutic response to anti-MAP antibiotics. Area covered: This review provides an appraisal of current information on MAP characteristics, diagnostic methodologies and emerging drug treatments. The authors focus on RHB-104, a novel oral formulation containing a fixed-dose combination of clarithromycin, clofazimine and rifabutin, endowed with synergistic inhibitory activity on MAP strains isolated from CD patients. Expert opinion: Based on encouraging in vitro data, RHB-104 has entered recently the clinical phase of its development, and is being investigated in a randomized, placebo-controlled phase III trial aimed at evaluating its efficacy and safety in CD. Provided that the overall clinical development will support the suitability of RHB-104 for inducing disease remission in CD patients with documented MAP infection, this novel antibiotic combination will likely take a relevant position in the therapeutic armamentarium for CD management

Analisi funzionale di piD261/Bud32, una protein chinasi atipica di lievito conservata evolutivamente.

Analisi funzionale di piD261/Bud32, una protein chinasi atipica di lievito conservata evolutivamente

Acidophilic character of yeast piD261/Bud32, a putative ancestor of eukaryotic protein kinases

Yeast piD261/Bud32 and its homologues are present in eukaryotes and in archaea but not in bacteria and are believed to make up a primordial branch of the eukaryotic protein kinase superfamily. Here, we show that, at variance with the majority of Ser/Thr protein kinases which recognize phosphoacceptor sites specified by basic and/or proline residues, piD261 phosphorylates in vitro a number of acidic proteins and peptides, and it recognizes seryl residues specified by carboxylic side chains. These data suggest that recognition of acidic sites might have been a primordial trait of protein kinases, which was modified during evolution to cope with the increasing complexity of protein phosphorylation in eukaryotes

Editorial: non‐invasive testing for EoE —does microbiome testing hold the key? Authors' reply

Authors' reply to article "non-invasive testing for EoE—does microbiome testing hold the key