Procalcitonin gene expression after LPS stimulation in the porcine animal model

Procalcitonin (PCT), recognised as a marker of sepsis, was investigated in a porcine model of endotoxic shock. The results showed that continuous IV infusion (1-4 h) of LPS (40 μg/kg) in pigs was able to induce a generalised increase of PCT expression in lung, heart, kidney and liver. The increase in PCT was significant only in kidney and was accompanied by an increase in IL-6 gene expression. In vitro results demonstrated that peripheral blood mononuclear cells (PBMCs), as well as endothelial cells, were potentially capable of contributing to in vivo extrathyroidal PCT production. These findings support previous data from pigs concerning the occurrence of widespread activation of PCT extrathyroidal gene expression during endotoxic shock in pigs. Nevertheless, the levels of PCT detected were very low, suggesting the need for additional studies to validate the pig as a reliable animal model for investigating the role of PCT in sepsis.     

Attenuation of virulence of Lawsonia intracellularis after in vitro passages and its effects on the experimental reproduction of porcine proliferative enteropathy

Non-pathogenic Lawsonia intracellularis variants have been obtained through multiple passages in cell culture but there is no information regarding the number of passages necessary to attenuate a pathogenic isolate. The present study evaluated the susceptibility of pigs to L. intracellularis after 10, 20 and 40 passages in vitro. Three groups (six animals/group) were inoculated with pure culture of L. intracellularis on passage 10, 20 or 40 and one group with placebo. The animals were monitored for clinical signs, fecal shedding and serological IgG response during 28 days post-inoculation. Gross and histologic lesions and the level of infection based on the amount of L. intracellularis-specific antigen in the intestinal mucosa identified by immunohistochemistry were evaluated in two animals from each group on days 14, 21 and 28. Animals inoculated with passages 10 and 20 demonstrated proliferative lesions typical of porcine proliferative enteropathy associated with the presence of Lawsonia-specific antigen in the intestinal mucosa. Passage 40-inoculated pigs did not show proliferative lesions or presence of Lawsonia antigen at any time point throughout the study. Similar patterns of the fecal shedding were observed in passage 10 and 20-infected pigs but those infected with passage 40 shed for a short period. Serological IgG responses in passage 10 and 20-inoculated pigs were detected from day 14 post-infection but not at all in passage 40-inoculated animals. These results demonstrate attenuation of the virulence properties of L. intracellularis between 20 and 40 cell passages in vitro. This information will be valuable for design of future experimental models and for studying the mechanisms involved in the attenuation of L. intracellularis virulence.     

Seeding strategies of bahiagrass and pintoi peanut affect pasture establishment under weed competition

Pintoi peanut (Arachis pintoi Krapov. & W.C. Greg.) is a warm‐season perennial legume with potential for use in grass–legume mixtures in Florida; however, limited information exists about its establishment in mixtures with bahiagrass (Paspalum notatum Flügge). The objective of this experiment was to evaluate the establishment of bahiagrass cv. “Argentine” and pintoi peanut cv. “Amarillo” as monocultures or mixture. The experiment was conducted in Ona, FL, from June to October of 2014 and 2015. Treatments were a split‐plot design of seeding strategies (bahiagrass monoculture, pintoi peanut monoculture or bahiagrass‐pintoi peanut mixtures; main plots) and two N fertilization strategies (30 or 80 kg/ha N; 30N and 80N; subplots), with four replicates. Measurements of plant density and frequency were taken every 4 weeks after seeding. Ground cover and herbage mass (HM) measurements were taken 112 days after seeding. Pintoi peanut ground cover was affected by seeding strategy × N level interaction. Ground cover was greater with 80N than 30N when pintoi was seeded in monoculture (3.6% vs 1.5% respectively) but not when it was seeded with bahiagrass (2.1%). There was no effect of seeding or N strategy on pintoi peanut proportion in HM (1.4%). Bahiagrass ground cover was not affected by seeding or N strategy (15.9%); however, its proportion in the HM was greater in 80N than 30N (12.1% vs 9.4% respectively). Mixed seeding did not negatively affect the establishment of bahiagrass and pintoi peanut and greater N fertilization levels improved some establishment parameters, with no negative effect for pintoi peanut.     

Assay of the set of all Sudan azodye (I, II, III, IV, and Para-Red) contaminating agents by liquid chromatography-tandem mass spectrometry and isotope dilution methodology

A high-throughput mass spectrometric method is presented for the simultaneous detection of Sudan I, II, III, IV, and Para-Red azodyes in foodstuff. The method is based on the use of deuterium-labeled internal standards and atmospheric pressure chemical ionization (APCI) in a triple-quadrupole instrument. The gas-phase breakdown pattern of each labeled and unlabeled analogue displays the naphthoic moiety as a common fragment. The search for the parents of this common species (parent ion scans) allows, by flow injection and in a single run, the evaluation of the presence of each polluting species spiked in typical foodstuffs. A detailed assay of each azodye was performed by LC-APCI and isotope dilution method, through the multiple reaction monitoring approach, using deuterium-labeled internal standards. Sudan dyes can be quantified above the threshold of 10 ppb except for Sudan Para-Red, for which the limit of quantification was 20 ppb, likely due to the different ionization efficiency.     

Applications of high-resolution solid-state NMR spectroscopy in food science

The principal applications of high-resolution solid-state NMR spectroscopy, in the field of food science, are reviewed, after a short general introduction, mainly focusing on the potential of these investigations, which are, today, routine tools for resolving technological problems. Selected examples of the applications in the field of food science of high-resolution solid-state NMR spectroscopy both in (13)C and in (1)H NMR particularly illustrative of the results obtainable are reported in some detail.     

Detection and quantification of genetically modified organisms using very short, locked nucleic acid TaqMan probes

Many countries have introduced mandatory labeling requirements on foods derived from genetically modified organisms (GMOs). Real-time quantitative polymerase chain reaction (PCR) based upon the TaqMan probe chemistry has become the method mostly used to support these regulations; moreover, event-specific PCR is the preferred method in GMO detection because of its high specificity based on the flanking sequence of the exogenous integrant. The aim of this study was to evaluate the use of very short (eight-nucleotide long), locked nucleic acid (LNA) TaqMan probes in 5'-nuclease PCR assays for the detection and quantification of GMOs. Classic TaqMan and LNA TaqMan probes were compared for the analysis of the maize MON810 transgene. The performance of the two types of probes was tested on the maize endogenous reference gene hmga, the CaMV 35S promoter, and the hsp70/cryIA(b) construct as well as for the event-specific 5'-integration junction of MON810, using plasmids as standard reference molecules. The results of our study demonstrate that the LNA 5'-nuclease PCR assays represent a valid and reliable analytical system for the detection and quantification of transgenes. Application of very short LNA TaqMan probes to GMO quantification can simplify the design of 5'-nuclease assays.