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Apple-related allergy is increasing among European population. Avoidance of apple fruit consumption is the only strategy for sensitized people, with a subsequent shortage of healthy nutrients in the diet. Four different protein families, Mal d 1 to 4, determine the allergy to apple causing syndromes known as oral allergy syndrome (OAS) or gastro-intestinal/generalized syndrome (GS) in sensitized patients. With the aim to highlight the dissimilarities in the immunological response inside the Mal d 1 family, recognized as main determinant for OAS, single purified allergens were tested against sera of paediatric and adult patients suffering for both syndromes. Here, we found that Mal d 1 isoallergens showed a remarkable different allergenicity potential, thus indicating an individual pattern in each patient and a diverse sensitization between age groups. Moreover, two different allergens, Mald 2.01 and Mal d 4.01, were analyzed with the above sera. In addition to patients and allergens variability, differences in amount and localization of the allergens, detected by patient’s sera, were revealed among hypo- and hyper allergenic apple varieties, confirming the complexity of this food allergy.

Sharka, a common disease among most stone fruit crops, is caused by the Plum Pox Virus (PPV). Resistant genotypes have been found in apricot (Prunus armeniaca L.), one of which--the cultivar 'Lito' heterozygous for the resistance--has been used to map a major quantitative trait locus (QTL) on linkage group 1, following a pseudo-test-cross mating design with 231 individuals. In addition, 19 SNP markers were selected from among the hundreds previously developed, which allowed the region to be limited to 236 kb on chromosome 1. A 'Lito' bacterial artificial chromosome (BAC) library was produced, screened with markers of the region, and positive BAC clones were sequenced. Resistant (R) and susceptible (S) haplotypes were assembled independently. To refine the assembly, the whole genome of 'Lito' was sequenced to high coverage (98×) using PacBio technology, enabling the development of a detailed assembly of the region that was able to predict and annotate the genes in the QTL region. The selected cultivar 'Lito' allowed not only to discriminate structural variants between the two haplotypic regions but also to distinguish specific allele expression, contributing towards mining the PPVres locus. In light of these findings, genes previously indicated (i.e., MATHd genes) to have a possible role in PPV resistance were further analyzed, and new candidates were discussed. Although the results are not conclusive, the accurate and independent assembly of R and S haplotypes of 'Lito' is a valuable resource to predict and test alternative transcription and regulation mechanisms underpinning PPV resistance.

In the last decades allergies are increasing in the western countries and different protein families were identified as determinants of allergenicity. In apple fruit, one of the most important fruit crop in the world, four allergen families (Mal d 1 to 4) have been identified so far, including pathogenesis-related proteins 10, thaumatin-like proteins, lipid-transfer proteins and profilins. Moreover, it has been demonstrated that patient’s sensitivity differs in relation to apple variety, fruit tissue, cultivation and conservation conditions, increasing the difficulties to associate genetic, molecular and biochemical data with the results of clinical tests. Mapping studies and the availability of the complete apple genome sequence confirmed that allergens are organized in large families with many different isoforms, which contribution to allergenicity is still largely unknown. The unravelling of allergy determinants is crucial for different reasons: i) to better understand environmental influences on allergenicity; ii) to produce genetically modified plants with a reduced allergen content through a targeted-gene silencing approach and iii) to drive future breeding programs toward hypoallergenic apple through the exploitation of natural variability. Finally, the above-mentioned knowledge will be essential to allow the consumption of this fruit by individuals who are allergic to apple.

Aims: To study the survival in the soil and the dispersion in the environment of Trichoderma atroviride SC1 after soil applications in a vineyard. Methods and Results: Trichoderma atroviride SC1 was introduced into soil in two consecutive years. The levels of T. atroviride populations at different spatial and temporal points following inoculation were assessed by counting the colony-forming units and by a specific quantitative real-time PCR. A high concentration of T. atroviride SC1 was still observed at the 18th week after inoculation. The vertical migration of the fungus to a soil depth of 0·4 m was already noticeable during the first week after inoculation. The fungus spread up to 4 m (horizontally) from the point of inoculation and its concentration decreased with the increasing distance (horizontal and vertical). It was able to colonize the rhizosphere and was also found on grapevine leaves. One year after soil inoculation, T. atroviride SC1 could still be recovered in the treated areas. Conclusions: Trichoderma atroviride SC1 survived and dispersed becoming an integrant part of the local microbial community under the tested conditions. Significance and Impact of the Study: The persistence and rapid spread of T. atroviride SC1 represent good qualities for its future use as biocontrol agent against soilborne pathogens. © 2009 The Society for Applied Microbiology.

Lactoferrin is a glycoprotein of about 80 kDa, belonging to the transferrin family. It is found in colostrum, milk and other biological fluids. This protein has antibacterial, antiviral and anticarcinogenic activity; it is involved in iron absorption and immunomodulation and it also promotes the growth of some strains of lactic acid bacteria (Brock, 2002; Kim et al., 2004). High LF concentrations in colostrum and milk have a positive role in the development of the newborn animal and in its protection against gastroenteric pathologies through its direct effect on pathogenic bacteria and in balancing newborn gut microflora toward probiotic bacteria (Schanbacher et al., 1997; Blum and Baumrucker, 2002). The administration of porcine LF in pig management could be a useful strategy to improve the growth of healthy piglets; during weaning they are subjected to several stressors that compromise their immunological status and can lead to gastroenteric diseases (Baldi et al., 1999). In view of the properties reviewed above, the aim of our work was the production of rpLF in heterologous expression system, the methylotrophic yeast Pichia pastoris, and evaluation of some of the biological activities of the recombinant protein expressed.

Quantitative Real-time PCR has proven to be a powerful tool to accurately 13 estimate transgene copy number and exogenous gene expression in plants. 14 Compared to Southern and Northern blot analysis, accuracy, rapidity and low-cost 15 are the main advantages. However, this technique requires important preliminary 16 work for standardizing and optimizing the many parameters involved in the 17 analysis. Grapevine data on this topic are not presently available in the literature. 18 We have developed a method for properly performing quantitative assays in grapes 19 where the genes for the b-glucoronidase (GUS) under the control of the 35S 20 promoter and for the neomicin phosphotransferase (NPTII) were transferred. 21 Validation of our method was verified with Southern blot analysis comparison. The 22 optimal endogenous genes to be applied as referee on grape were exploited and nine23 cis-epoxycarotenoid dioxygenase2 (NCED2) resulted the most amenable. For 24 transgene copy number detection, the approach based on synthetic hybrid amplicons 25 proved to be promising.