Abstract. The whole genome of Mangalica animals has been screened on the Illumina porcine chip giving the possibility (1) to replace the previously applied ten microsatellite markers by nine SNP loci to classify the Blond, Swallow-Belly and Red Mangalica individuals into three different breed groups (P>0.95); (2) to propose 54 SNP loci for parentage testing in Mangalica pigs where the exclusion probability is 0.999115 if one parent is known and the probability of identity is 1.54×10-23.

Background: With the increasing number of GMOs on the global market the maintenance of European GMO regulations is becoming more complex. For the analysis of a single food or feed sample it is necessary to assess the sample for the presence of many GMO-targets simultaneously at a sensitive level. Several methods have been published regarding DNA-based multidetection. Multiplex ligation detection methods have been described that use the same basic approach: i) hybridisation and ligation of specific probes, ii) amplification of the ligated probes and iii) detection and identification of the amplified products. Despite they all have this same basis, the published ligation methods differ radically. The present study investigated with real-time PCR whether these different ligation methods have any influence on the performance of the probes. Sensitivity and the specificity of the padlock probes (PLPs) with the ligation protocol with the best performance were also tested and the selected method was initially validated in a laboratory exchange study.Results: Of the ligation protocols tested in this study, the best results were obtained with the PPLMD I and PPLMD II protocols and no consistent differences between these two protocols were observed. Both protocols are based on padlock probe ligation combined with microarray detection. Twenty PLPs were tested for specificity and the best probes were subjected to further evaluation. Up to 13 targets were detected specifically and simultaneously. During the interlaboratory exchange study similar results were achieved by the two participating institutes (NIB, Slovenia, and RIKILT, the Netherlands).Conclusions: From the comparison of ligation protocols it can be concluded that two protocols perform equally well on the basis of the selected set of PLPs. Using the most ideal parameters the multiplicity of one of the methods was tested and 13 targets were successfully and specifically detected. In the interlaboratory exchange study it was shown that the selected method meets the 0.1% sensitivity criterion. The present study thus shows that specific and sensitive multidetection of GMO targets is now feasible.

BackgroundWe aimed to determine the effect of feeding transgenic maize to sows during gestation and lactation on maternal and offspring immunity and to assess the fate of transgenic material.Methodology/principal findingsOn the day of insemination, sows were assigned to one of two treatments (n = 12/treatment); 1) non-Bt control maize diet or 2) Bt-MON810 maize diet, which were fed for ~143 days throughout gestation and lactation. Immune function was assessed by leukocyte phenotyping, haematology and Cry1Ab-specific antibody presence in blood on days 0, 28 and 110 of gestation and at the end of lactation. Peripheral-blood mononuclear cell cytokine production was investigated on days 28 and 110 of gestation. Haematological analysis was performed on offspring at birth (n = 12/treatment). Presence of the cry1Ab transgene was assessed in sows' blood and faeces on day 110 of gestation and in blood and tissues of offspring at birth. Cry1Ab protein presence was assessed in sows' blood during gestation and lactation and in tissues of offspring at birth. Blood monocyte count and percentage were higher (P<0.05), while granulocyte percentage was lower (P<0.05) in Bt maize-fed sows on day 110 of gestation. Leukocyte count and granulocyte count and percentage were lower (P<0.05), while lymphocyte percentage was higher (P<0.05) in offspring of Bt maize-fed sows. Bt maize-fed sows had a lower percentage of monocytes on day 28 of lactation and of CD4(+)CD8(+) lymphocytes on day 110 of gestation, day 28 of lactation and overall (P<0.05). Cytokine production was similar between treatments. Transgenic material or Cry1Ab-specific antibodies were not detected in sows or offspring.Conclusions/significanceTreatment differences observed following feeding of Bt maize to sows did not indicate inflammation or allergy and are unlikely to be of major importance. These results provide additional data for Bt maize safety assessment.

Background The objective of this study was to evaluate potential long-term (110 days) and age-specific effects of feeding genetically modified Bt maize on peripheral immune response in pigs and to determine the digestive fate of the cry1Ab gene and truncated Bt toxin. Methodology/Principal Findings Forty day old pigs (n = 40) were fed one of the following treatments: 1) isogenic maize-based diet for 110 days (isogenic); 2) Bt maize-based diet (MON810) for 110 days (Bt); 3) Isogenic maize-based diet for 30 days followed by Bt maize-based diet for 80 days (isogenic/Bt); and 4) Bt maize-based diet (MON810) for 30 days followed by isogenic maize-based diet for 80 days (Bt/isogenic). Blood samples were collected during the study for haematological analysis, measurement of cytokine and Cry1Ab-specific antibody production, immune cell phenotyping and cry1Ab gene and truncated Bt toxin detection. Pigs were sacrificed on day 110 and digesta and organ samples were taken for detection of the cry1Ab gene and the truncated Bt toxin. On day 100, lymphocyte counts were higher (P<0.05) in pigs fed Bt/isogenic than pigs fed Bt or isogenic. Erythrocyte counts on day 100 were lower in pigs fed Bt or isogenic/Bt than pigs fed Bt/isogenic (P<0.05). Neither the truncated Bt toxin nor the cry1Ab gene were detected in the organs or blood of pigs fed Bt maize. The cry1Ab gene was detected in stomach digesta and at low frequency in the ileum but not in the distal gastrointestinal tract (GIT), while the Bt toxin fragments were detected at all sites in the GIT. Conclusions/Significance Perturbations in peripheral immune response were thought not to be age-specific and were not indicative of Th 2 type allergenic or Th 1 type inflammatory responses. There was no evidence of cry1Ab gene or Bt toxin translocation to organs or blood following long-term feeding.

The main goal of our work was to develop a rapid, simple, and economical DNA extraction method for food (especially for meat products) analysis. This extraction and purification procedure was based on the three-phase partitioning (TPP) method. The developed new DNA-TPP method and Wizard DNA Clean-Up System (Promega, USA) have been compared concerning extraction efficiency, purity and DNA suitability for amplification. The quality and quantity of the purified DNA solutions were controlled by spectrophotometer and the amplification efficiency by simple qualitative PCR. All of prepared DNA solutions were pure enough for the PCR and contained appropriate quantity of DNA. Thus, 118 bp length amplicons could have been obtained by the specific lectin-gene PCR in all cases. This method proved to be an alternative one to isolate DNA from meat samples simply and economically.

The main goal of our research was to establish to what extent Hungarian consumers are exposed to consuming genetically modified foods. To achieve this goal, 251 food samples were bought from the Hungarian food market and examined by the Central Food Research Institute (91 samples) and by the National Institute for Food Safety and Nutrition (160 samples) with PCR. The GMO screening was based on the detection of 35S promoter and nos terminator sequences. The GM positive samples were subjected to event specific detection of RR soy with quantitative real-time PCR. According to the results 38% of the examined samples indicated presence of RR soy in soy-containing foods and 6% of these samples contained RR soy above the 0.9% threshold. These results demonstrate for the first time the assessment of the presence of GMO in the Hungarian food market for large sample numbers.

Polymerase chain reaction-restriction fragment length polymorphism was applied to species-specific detection of poultry meat in commercial sausage products. The sequence selected for amplification was a 359 base pair fragment of the mitochondrial cytochrome b gene as part of the template DNA. DNA was extracted from raw pork, beef, chicken, turkey, heat-treated model meat mixtures and from commercial sausage products using a DNA-binding silica resin, and subjected to polymerase chain reaction-restriction fragment length polymorphism analysis. Polymerase chain reaction analysis demonstrated a 359 base pair fragment for all meat samples examined. Rsa1 enzyme restriction profiles showed that poultry produced two different fragment patterns: two fragments (210 and 149 base pairs) for chicken and three fragments (149, 109, 101 base pairs) for turkey. The detection limit for chicken in meat model mixtures was 0.5%. Three positive commercial sausage samples were found to contain non-declared poultry.

The aim of the study was to assay by PCR screening method whether the processing and the thermal stress have any influence on the feasibility of the detection of genetically modified DNA in different kinds of processed meat products such as sausages, liver cans, ready-to-eat hamburgers. The model meat products have been prepared with soybean meal spiked with RR (Roundup Ready) soybean meal in 0.5%, 1%, 1.5% and 2%. The samples were prepared under industrial circumstances. The assay was based on the detection of the specific part of the 35S promoter and the NOS terminator sequences. The modified PCR method was shown to be suitable for screening of GMOs in raw and also in moderately and highly processed meat samples when extreme heat treatment and pressure were used for the preparation of meat products. Half a percent RR soy contamination could be detected even if the food products underwent high temperature treatment.

It is well established that the ingestion of cereal prolamins, such as gluten, causes the characteristic symptoms of celiac disease (CD) in people predisposed to it. DNA-based PCR method provides new ways to detect gluten in processed foodstuffs, such as bread. The aim of this work was to adapt a new primer pair combination and to initiate a carefully elaborated PCR methodology to experiment with DNA-based analysis. At first, the purity of cleaned DNA was verified using B49317 and A49855 chloroplast DNA primer pair. Then TR01/2 wheat specific PCR primer pair was used for checking the origin of the DNA, and P1/2 microsatellite (SSR) adapted primer pair for detecting allergen (gluten) specific residues. Method optimisation was achieved with cereal flour samples, then bread and dry pasta products front wheat were used, which were analysed as heat-treated samples with three primer pairs. The gluten specific primer pair was tested on cross-reactive cereals such as rye, barley, triticale and on some questionable cereals, such as oat, and pseudo-cereals, e.g. buckwheat and amaranth.