Development and Evaluation of qPCR Detection Method and Zn-MgO/Alginate Active Packaging for Controlling Listeria monocytogenes Contamination in Cold-Smoked Salmon

To answer to food industry requests to monitor the presence of L. monocytogenes in cold-smoked salmon samples and to extend their shelf-life, a qPCR protocol for the detection of L. monocytogenes, and an antibacterial active packaging reinforced with zinc magnesium oxide nanoparticles (Zn-MgO NPs) were developed. The qPCR allowed the sensitive and easy detection of L. monocytogenes in naturally contaminated samples, with specificity in full agreement with the standard methods. The halo diusion study indicated a high antibacterial eciency of 1 mg/mL Zn-MgO NPs against L. monocytogenes, while the flow cytometry showed only moderate cytotoxicity of the nanoparticles towards mammalian cells at a concentration above 1 mg/mL. Thus, the novel active packaging was developed by using 1 mg/mL of Zn-MgO NPs to reinforce the alginate film. Cold-smoked salmon samples inoculated with L. monocytogenes and air-packed with the Zn-MgO NPs-alginate nanobiocomposite film showed no bacterial proliferation at 4 C during 4 days. In the same condition, L. monocytogenes growth in control contaminated samples packed with alginate film alone. Our results suggest that Zn-MgO nanoparticles can extend the shelf-life of cold-smoked salmon samples

Isolation and expression of a novel MBL-like collectin cDNA enhanced by LPS injection in the body wall of the ascidian Ciona intestinalis

Collectins are a family of calcium-dependent lectins that are characterized by their collagen-like domains. Considerable interest has been focused on this class of proteins because of their ability to interact with components of the complement system activating a cascade of events responsible for the activation of the innate immune system. A differential screening between LPS-challenged and na\uefve Ciona intestinalis has been performed allowing the isolation of a full length cDNA encoding for a 221 AA protein. In silico analysis has shown that this polypeptide displays protein domains with similarities to mannose-binding lectins. A phylogenetic analysis suggested that C. intestinalis MBL has evolved early as a prototype of vertebrate MBL. Real-time PCR assay demonstrated that this gene is strongly activated after LPS injection in the tunica. In situ hybridization performed in LPS-induced animals has shown that this gene is expressed in granular amoebocytes and large granules hemocytes in the inflamed body wall tissue. Finally, an antimicrobial activity of the C. intestinalis MBL has been demonstrate

LPS Challenge Regulates Gene Expression and Tissue Localization of a Ciona intestinalis Gene through an Alternative Polyadenylation Mechanism

subtractive hybridization strategy for the identification of differentially expressed genes was performed between LPSchallenged and naive Ciona intestinalis. This strategy allowed the characterization of two transcripts (Ci8short and Ci8long) generated by the use of two Alternative Polyadenylation sites. The Ci8long transcript contains a protein domain with relevant homology to several components of the Receptor Transporting Protein (RTP) family not present in the Ci8short mRNA. By means of Real Time PCR and Northern Blot, the Ci8short and Ci8long transcripts showed a different pattern of gene expression with the Ci8short mRNA being strongly activated after LPS injection in the pharynx. In situ hybridization analysis demonstrated that the activation of the APA site also influenced the tissue localization of the Ci8short transcript. This analysis showed that the Ci8long mRNA was expressed in hemocytes meanwhile the Ci8short mRNA was highly transcribed also in vessel endothelial cells and in the epithelium of pharynx. These findings demonstrated that regulation of gene expression based on different polyadenylation sites is an ancestral powerful strategy influencing both the level of expression and tissue distribution of alternative transcripts

Temperature modulates the response of the thermophilous sea urchin Arbacia lixula early life stages to CO2-driven acidification

The increasing abundances of the thermophilous black sea urchin Arbacia lixula in the Mediterranean Sea are attributed to the Western Mediterranean warming. However, few data are available on the potential impact of this warming on A. lixula in combination with other global stressors such as ocean acidification. The aim of this study is to investigate the interactive effects of increased temperature and of decreased pH on fertilization and early development of A. lixula. This was tested using a fully crossed design with four temperatures (20, 24, 26 and 27 C) and two pH levels (pHNBS 8.2 and 7.9). Temperature and pH had no significant effect on fertilization and larval survival (2d) for temperature <27 C. At 27 C, the fertilization success was very low (<1%) and all larvae died within 2d. Both temperature and pH had effects on the developmental dynamics. Temperature appeared to modulate the impact of decreasing pH on the % of larvae reaching the pluteus stage leading to a positive effect (faster growth compared to pH 8.2) of low pH at 20 C, a neutral effect at 24 C and a negative effect (slower growth) at 26 C. These results highlight the importance of considering a range of temperatures covering today and the future environmental variability in any experiment aiming at studying the impact of ocean acidificatio

Fish assemblages cope with ocean acidification in a shallow volcanic CO2 vent benefiting from an adjacent recovery area

Shallow CO2 vents are used to test ecological hypotheses about the effects of ocean acidification (OA). Here, we studied fish assemblages associated with Cymodocea nodosa meadows exposed to high pCO2/low pH conditions at a natural CO2 vent in the Mediterranean Sea. Using underwater visual census, we assessed fish community structure and biodiversity in a low pH site (close to the CO2 vent), a close control site and a far control site, hypothesising a decline in biodiversity and a homogenization of fish assemblages under OA conditions. Our findings revealed that fish diversity did not show a unique spatial pattern, or even significant relationships with pH, but correlated with seagrass leaf canopy. Among-site similarity was found in the abundance of juveniles, contrary to the expected impacts of OA on early life stages. However, pH seems an important driver in structuring fish assemblage in the low pH site, despite its high similarity with the close control site. This unexpected pattern may represent a combined response of fish mobility, enhanced food resources in the acidified site, and a ‘recovery area’ effect of the adjacent control site

Isolation of a novel LPS-induced component of the ML superfamily in Ciona intestinalis

ML superfamily represents a group of proteins playing important roles in lipid metabolism and innate immune response. In this study, we report the identification of the first component of the ML superfamily in the invertebrate Ciona intestinalis by means of a subtractive hybridization strategy. Sequence homology and phylogenetic analysis showed that this protein forms a specific clade with vertebrate components of the Niemann-Pick type C2 protein and, for this reason, it has been named Ci-NPC2. The putative Ci-NPC2 is a 150 amino acids long protein with a short signal peptide, seven cysteine residues, three putative lipid binding site and a three-dimensional model showing a characteristic b-strand structure. Gene expression analysis demonstrated that the Ci-NPC2 protein is positively upregulated after LPS inoculum with a peak of expression 1 h after challenge. Finally, in-situ hybridization demonstrated that the Ci-NPC2 protein is preferentially expressed in hemocytes inside the vessel lumen

Detection of Listeria monocytogenes in foods with a textile organic electrochemical transistor biosensor

Abstract: Foods contaminated by pathogens are responsible for foodborne diseases which have socioeconomic impacts. Many approaches have been extensively investigated to obtain specific and sensitive methods to detect pathogens in food, but they are often not easy to perform and require trained personnel. This work aims to propose a textile organic electrochemical transistor-based (OECT) biosensor to detect L. monocytogenes in food samples. The analyses were performed with culture-based methods, Listeria Precis™ method, PCR, and our textile OECT biosensor which used poly(3,4-ethylenedioxythiophene) (PEDOT):polystyrene sulfonate (PSS) (PEDOT:PSS) for doping the organic channel. Atomic force microscopy (AFM) was used to obtain topographic maps of the gold gate. The electrochemical activity on gate electrodes was measured and related to the concentration of DNA extracted from samples and hybridized to the specific capture probe immobilized onto the gold surface of the gate. This assay reached a limit of detection of 1.05 ng/μL, corresponding to 0.56 pM of L. monocytogenes ATCC 7644, and allowed the specific and rapid detection of L. monocytogenes in the analyzed samples. Keypoints: • Textile organic electrochemical transistors functionalized with a specific DNA probe • AFM topographic and surface potential maps of a functionalized gold gate surface • Comparison between the Listeria monocytogenes Precis™ method and an OECT biosenso