Antibacterial activity of bovine milk lactoferrin on the emerging foodborne pathogen Cronobacter sakazakii: Effect of media and heat treatment

Cronobacter sakazakii is a pathogen transmitted by food, with high osmotic resistance and tolerance to desiccation, which affects mainly to newborns, infants and immunocompromised adults. C. sakazakii infection in infants has been associated with consumption of powdered milk. The purpose of this study was to evaluate the antibacterial activity of native and iron-saturated bovine lactoferrin (bLF) (from 0.5 to 5 mg/ml) on non-desiccated and desiccated C. sakazakii (104 CFU/ml) in different media (phosphate buffer, bovine skim milk and whey). In general, native bLF was the only effective form that inhibited growth of C. sakazakii in all media, its activity increasing with concentration and time of incubation. These results suggest that the antibacterial effect of bLF on C. sakazakii is mainly due to iron sequestration. However, iron-saturated bLF showed some effect by reducing the viability of C. sakazakii in whey. There has not been observed an increased sensitivity of desiccated bacteria to native bLF in phosphate buffer. However, although the antibacterial activity of native bLF against non-desiccated C. sakazakii was drastically reduced in milk or whey compared to phosphate buffer, there was a certain activity when it was assayed against desiccated cells in those media. The effect of some heat treatments on the antibacterial activity of native bLF was evaluated and only those of 72 °C for 15 s, 85 °C for 15 s, and 63 °C for 30 min maintained its whole activity

Detection of peanut (Arachis hypogaea) allergens in processed foods by immunoassay: Influence of selected target protein and ELISA format applied

Direct competitive and sandwich ELISA formats developed to determine Ara h1 and Ara h2 proteins were applied in the detection of peanut in model biscuits prepared with a commercial peanut butter as ingredient. The sandwich format for Ara h2 protein could detect the addition of 2.5% peanut butter, whereas the same format for Ara h1 could not detect 5% added peanut. Direct competitive formats for Ara h1 and Ara h2 proteins could detect the addition of 1% and 0.05% peanut butter, respectively. Therefore, competitive format for Ara h2 was selected to be evaluated by four laboratories, obtaining adequate results in term of repeatability and reproducibility. Results obtained indicate that processing decreased the level of extracted proteins and underestimated the amount of Ara h1 and Ara h2 proteins, the effect being more severe for Ara h1. The selection of the target protein and the ELISA format applied greatly influence the detection of peanut in processed foods

Ovine HSP90AA1 Expression Rate Is Affected by Several SNPs at the Promoter under Both Basal and Heat Stress Conditions

The aim of this work was to investigate the association between polymorphisms located at the HSP90AA1 ovine gene promoter and gene expression rate under different environmental conditions, using a mixed model approach. Blood samples from 120 unrelated rams of the Manchega sheep breed were collected at three time points differing in environmental conditions. Rams were selected on the basis of their genotype for the transversion G/C located 660 base pairs upstream the gene transcription initiation site. Animals were also genotyped for another set of 6 SNPs located at the gene promoter. Two SNPs, G/C-660 and A/G-444, were associated with gene overexpression resulting from heat stress. The composed genotype CC-660-AG-444 was the genotype having the highest expression rates with fold changes ranging from 2.2 to 3.0. The genotype AG-522 showed the highest expression levels under control conditions with a fold change of 1.4. Under these conditions, the composed genotype CC-601-TT-524-AG-522-TT-468 is expected to be correlated with higher basal expression of the gene according to genotype frequencies and linkage disequilibrium values. Some putative transcription factors were predicted for binding sites where the SNPs considered are located. Since the expression rate of the gene under alternative environmental conditions seems to depend on the composed genotype of several SNPs located at its promoter, a cooperative regulation of the transcription of the HSP90AA1 gene could be hypothesized. Nevertheless epigenetic regulation mechanisms cannot be discarded. © 2013 Salces-Ortiz et al

Evaluation of methods for DNA extraction from Clostridium tyrobutyricum spores and its detection by qPCR

Clostridium tyrobutyricum is the major agent that causes the blowing defect in cheese due to the germination of its dormant spores during the ripening stage. As a result, many of the affected cheeses show cavities and cracks, which cause the product loss in most cases. Nowadays, there is not a fast method capable of detecting milk contaminated with C. tyrobutyricum spores. The aim of this study has been to develop a fast and reliable method based on real time PCR (qPCR) to detect C. tyrobutyricum spores in raw milk. One of the main limitations has been to find a good procedure for the spore disruption to extract the DNA due to its high resistance. For this reason, different disruption methods have been tested, including chemical agents, bead beating, enzymatic and microwave treatment. Furthermore, an enzymatic treatment with subtilisin was applied for milk clarification and recovery of spores. The comparison of the assayed methods has been made using sterile milk spiked with C. tyrobutyricum spores, obtained in solid or liquid medium. The results showed that microwave treatment followed by a standard DNA purification step was found to be the best disruption method. The Ct values obtained for spores were higher than those found for vegetative cells by qPCR, for the same quantity of DNA. This difference could be due to the action of the Small Acid Soluble Proteins (SASP) in the DNA packaging of spores. Moreover, spores obtained in agar plate were found more resistant to disruption than those obtained in liquid medium. Subtilisin and microwave treatments were found to be successful for DNA extraction from C. tyrobutyricum spores in milk and subsequent identification by qPCR. However, the differences observed between the amplification of DNA from spores obtained in different media and from vegetative cells have to be taken into account to optimize a method for C. tyrobutyricum detection

Detection of butyric spores by different approaches in raw milks from cow, ewe and goat

Clostridium tyrobutyricum is described as the main causative agent of late blowing defect in cheese. Currently, there are no fast methods to detect this microorganism in raw milk, which would allow determining the use of milk for fresh or cured cheese. The technique commonly used is the Most Probable Number, which is laborious and non-specific. In this work, we present the optimization of a real-time PCR-based detection method for C. tyrobutyricum spores in raw milk samples. This novel approach extracts DNA in a semi-automatic system with magnetic beads. The applicability of the developed procedure has been tested in field milk samples from cow, ewe and goat (n = 202), allowing detection of low levels of butyric spores. Raw milk samples were also analyzed by microbiological culture in a selective medium for butyric bacteria, and positive colonies were identified by multiplex PCR and 16S rDNA sequencing. Apart from C. tyrobutyricum, other Clostridium spp. were identified, which should be considered for further development of detection methods

Exploring Health Science Students’ Notions on Organ Donation and Transplantation: A Multicenter Study

The knowledge acquired during university education about organ donation and transplantation (ODT) decisively influences the information future health professionals transmit. This is important in ODT where the participation of the general public is essential to obtain organs. Objective: To determine notions of Spanish medicine and nursing students on ODT and its relationship with attitude toward ODT. Methods and Design: and design. We conducted a sociologic, multicenter, and observational study. The population for our study consisted of medical and nursing students in Spanish universities. Our database was the Collaborative International Donor Project, stratified by geographic area and academic course. A validated questionnaire (PCID-DTO-RIOS) was self-administered and completed anonymously. Our sample consisted of 9598 medical and 10, 566 nursing students (99% confidence interval; precision of ±1%), stratified by geographic area and year of study. Results: The completion rate for our study was 90%. Only 20% (n=3640) of students thought their notions on ODT were good; 41% (n=7531) thought their notions were normal; 36% (n=6550) thought their notions were scarce. Comparing groups, there were differences between those who believed that their notions on ODT were good (44% nursing vs 56% medical students; P < .000), and those who believed it scarce (54% nursing vs 46% medical students; P < .000). Notions on ODT were related with attitude toward the donation of one''s own organs: those who considered their notions were good were more in favor then those who considered it scarce (88% vs 72%; P < .000). Conclusion: Only 20% of Spanish medical and nursing students thought their notions on ODT were good. Having good knowledge is related to a favorable attitude towards ODT. Receiving specific information on the subject could improve their knowledge about ODT during their training

Model-independent search for CP violation in D0→K−K+π−π+ and D0→π−π+π+π− decays

A search for CP violation in the phase-space structures of D0 and View the MathML source decays to the final states K−K+π−π+ and π−π+π+π− is presented. The search is carried out with a data set corresponding to an integrated luminosity of 1.0 fb−1 collected in 2011 by the LHCb experiment in pp collisions at a centre-of-mass energy of 7 TeV. For the K−K+π−π+ final state, the four-body phase space is divided into 32 bins, each bin with approximately 1800 decays. The p-value under the hypothesis of no CP violation is 9.1%, and in no bin is a CP asymmetry greater than 6.5% observed. The phase space of the π−π+π+π− final state is partitioned into 128 bins, each bin with approximately 2500 decays. The p-value under the hypothesis of no CP violation is 41%, and in no bin is a CP asymmetry greater than 5.5% observed. All results are consistent with the hypothesis of no CP violation at the current sensitivity

Search for the lepton-flavor-violating decays Bs0→e±μ∓ and B0→e±μ∓

A search for the lepton-flavor-violating decays Bs0→e±μ∓ and B0→e±μ∓ is performed with a data sample, corresponding to an integrated luminosity of 1.0  fb-1 of pp collisions at √s=7  TeV, collected by the LHCb experiment. The observed number of Bs0→e±μ∓ and B0→e±μ∓ candidates is consistent with background expectations. Upper limits on the branching fractions of both decays are determined to be B(Bs0→e±μ∓)101  TeV/c2 and MLQ(B0→e±μ∓)>126  TeV/c2 at 95% C.L., and are a factor of 2 higher than the previous bounds

Branching fraction and CP asymmetry of the decays B+→K0Sπ+ and B+→K0SK+

An analysis of B+ → K0 Sπ+ and B+ → K0 S K+ decays is performed with the LHCb experiment. The pp collision data used correspond to integrated luminosities of 1 fb−1 and 2 fb−1 collected at centre-ofmass energies of √ s = 7 TeV and √ s = 8 TeV, respectively. The ratio of branching fractions and the direct CP asymmetries are measured to be B(B+ → K0 S K+ )/B(B+ → K0 Sπ+ ) = 0.064 ± 0.009 (stat.) ± 0.004 (syst.), ACP(B+ → K0 Sπ+ ) = −0.022 ± 0.025 (stat.) ± 0.010 (syst.) and ACP(B+ → K0 S K+ ) = −0.21 ± 0.14 (stat.) ± 0.01 (syst.). The data sample taken at √ s = 7 TeV is used to search for B+ c → K0 S K+ decays and results in the upper limit ( fc · B(B+ c → K0 S K+ ))/( fu · B(B+ → K0 Sπ+ )) < 5.8 × 10−2 at 90% confidence level, where fc and fu denote the hadronisation fractions of a ¯b quark into a B+ c or a B+ meson, respectively