Cinzia Montemurro

Genetic relationships and cultivar identification among 112 olive accessions using AFLP and SSR markers.

Summary A total of 111 accessions belonging to 60 olive cultivars, and one accession of oleaster, native to Italy, Spain, France and Greece, have been screened with three AFLP primer combinations and 27 microsatellite primer pairs in order to characterise their genotypes and reveal their genetic relationships. A total of 70 AFLP and 80 SSR polymorphic bands were scored. Comparisons were made between AFLP and SSR marker variability, efficiency and usefulness for genetic relationships and cultivar identification. The data obtained were analysed using the Jaccard genetic similarity coefficient, applying the SAHN clustering method. A dendrogram of genetic distances was produced. All genotypes studied could be distinguished unequivocally using a combination of SSR and AFLP markers. Cultivars were grouped into three clusters according to their type of use: oil, table, or dual purpose cultivars.

DNA Microsatellite Region for a Reliable Quantification of Soft Wheat Adulteration in Durum Wheat-Based Foodstuffs by Real-Time PCR

Italian industrial pasta and durum wheat typical breads must be prepared using exclusively durum wheat semolina. Previously, a microsatellite sequence specific of the wheat D-genome had been chosen for traceability of soft wheat in semolina and bread samples, using qualitative and quantitative Sybr green-based real-time experiments. In this work, we describe an improved method based on the same soft wheat genomic region by means of a quantitative real-time PCR using a dual-labeled probe. Standard curves based on dilutions of 100% soft wheat flour, pasta, or bread were constructed. Durum wheat semolina, pasta, and bread samples were prepared with increasing amounts of soft wheat to verify the accuracy of the method. Results show that reliable quantifications were obtained especially for the samples containing a lower amount of soft wheat DNA, fulfilling the need to verify labeling of pasta and typical durum wheat breads.

Genetic structure and natural variation associated with host of origin in Penicillium expansum strains causing blue mould.

Blue mould, caused by Penicillium expansum, is one of the most economically damaging postharvest diseases of pome fruits, although it may affect a wider host range, including sweet cherries and table grapes. Several reports on the role of mycotoxins in plant pathogenesis have been published, but few focussed on the influence of mycotoxins on the variation in host preference amongst producing fungi. In the present study the influence of the host on P. expansum pathogenicity/virulence was investigated, focussing mainly on the relationship with patulin production. Three P. expansum strain groups, originating from apples, sweet cherries, and table grapes (7 strains per host) were grown on their hosts of isolation and on artificial media derived from them. Strains within each P. expansum group proved to be more aggressive and produced more patulin than the other two groups under evaluation when grown on the host from which they originated. Table grape strains were the most aggressive (81% disease incidence) and strongest patulin producers (up to 554μg/g). The difference in aggressiveness amongst strains was appreciable only in the presence of a living host, suggesting that the complex pathogen-host interaction significantly influenced the ability of P. expansum to cause the disease. Incidence/severity of the disease and patulin production proved to be positively correlated, supporting the role of patulin as virulence/pathogenicity factor. The existence of genetic variation amongst isolates was confirmed by the High Resolution Melting method that was set up herein, which permitted discrimination of P. expansum from other species (P. chrysogenum and P. crustosum) and, within the same species, amongst the host of origin. Host effect on toxin production appeared to be exerted at a transcriptional level.

Polyphenol Oxidases in Crops: Biochemical, Physiological and Genetic Aspects

Enzymatic browning is a colour reaction occurring in plants, including cereals, fruit and horticultural crops, due to oxidation during postharvest processing and storage. This has a negative impact on the colour, flavour, nutritional properties and shelf life of food products. Browning is usually caused by polyphenol oxidases (PPOs), following cell damage caused by senescence, wounding and the attack of pests and pathogens. Several studies indicated that PPOs play a role in plant immunity, and emerging evidence suggested that PPOs might also be involved in other physiological processes. Genomic investigations ultimately led to the isolation of PPO homologs in several crops, which will be possibly characterized at the functional level in the near future. Here, focusing on the botanic families of Poaceae and Solanaceae, we provide an overview on available scientific literature on PPOs, resulting in useful information on biochemical, physiological and genetic aspects.

Traceability of PDO Olive Oil “Terra di Bari” Using High Resolution Melting

The aim of the research was to verify the applicability of microsatellite (SSR) markers in High Resolution Melting (HRM) analysis for the identification of the olive cultivars used in the “Terra di Bari” PDO extra virgin olive oil. A panel of nine cultivars, widespread in Apulia region, was tested with seventeen SSR primer pairs and the PCR products were at first analysed with a Genetic Analyzer automatic sequencer. An identification key was obtained for the nine cultivars, which showed an unambiguous discrimination among the varieties constituting the “Terra di Bari” PDO extra virgin olive oil: Cima di Bitonto, Coratina, and Ogliarola. Subsequently, an SSR based method was set up with the DCA18 marker, coupled with HRM analysis for the distinction of the Terra di Bari olive oil from non-Terra di Bari olive oil using different mixtures. Thus, this analysis enabled the distinction and identification of the PDO mixtures. Hence, this assay provided a flexible, cost-effective, and closed-tube microsatellite genotyping method, well suited to varietal identification and authentication analysis in olive oil.

Traceability of Italian Protected Designation of Origin (PDO) table olives by means of microsatellite molecular markers.

The aim of this work was to develop a DNA microsatellite-based method of analysis to allow traceability of the three Italian Protected Designation of Origin (PDO) table olives in comparison with fruits of another seven highly diffused table olive cultivars. The analyses were carried out by using 16 primer pairs, with a mean of five different alleles detected per primer set, and power of discrimination from 0.56 to 0.90. Allelic error rates in the range of 0-3.8% were observed. By combining data from the most reliable and highly informative microsatellites (DCA3, DCA16, DCA17, DCA18, UDO-043, and GAPU101), it was possible to identify the PDO fruits over the panel of 10 cultivars, with the probability of a chance match between different cultivars as low as 10(-9) and with 0.5% error rate. The amplification profile is independent of environmental and processing conditions and is helpful to verify the authenticity of PDO samples.

Cultivar classification of Apulian olive oils: Use of artificial neural networks for comparing NMR, NIR and merceological data

The development of an efficient and accurate method for extra-virgin olive oils cultivar and origin authentication is complicated by the broad range of variables (e.g., multiplicity of varieties, pedo-climatic aspects, production and storage conditions) influencing their properties. In this study, artificial neural networks (ANNs) were applied on several analytical datasets, namely standard merceological parameters, near-infra red data and 1H nuclear magnetic resonance (NMR) fingerprints, obtained on mono-cultivar olive oils of four representative Apulian varieties (Coratina, Ogliarola, Cima di Mola, Peranzana). We analyzed 888 samples produced at a laboratory-scale during two crop years from 444 plants, whose variety was genetically ascertained, and on 17 industrially produced samples. ANN models based on NMR data showed the highest capability to classify cultivars (in some cases, accuracy>99%), independently on the olive oil production process and year; hence, the NMR data resulted to be the most informative variables about the cultivars.

An evaluation of a new approach to the regeneration of Helichrysum italicum (Roth) G. Don, and the molecular characterization of the variation among sets of differently derived regenerants

A protocol for the induction of regeneration from leaves of Helichrysum italicum was established. Calli were found to form on the basal medium only when it was supplemented with thidiazuron (TDZ) alone or in combination with naphthalene acetic acid (NAA), with a percentage ranking of at least 80%. The hormone-free medium showed the highest percentage of shoot regeneration (62%) even though no callus formed. AFLP markers were employed to verify tissue culture-induced variation in the regenerated plantlets obtained by direct shoot regeneration or the indirect shoot regeneration process (callus formation). Seven out of the eleven AFLP primer pairs yielded polymorphic patterns. The average number of fragments per primer pair was 64.1. Singletons were represented by 12 (2.7%) fragments. Student’s T-test was performed both on the average number of shared fragments and on the nucleotide diversity, and no significant statistical difference was observed between the two regeneration treatments.

The coexistence of oleaster and traditional varieties affects genetic diversity and population structure in Algerian olive (Olea europaea) germplasm

The present work was aimed at assessing the genetic diversity of 42 local cultivars and oleaster genotypes from the area of Bejaia in Algeria. Fifteen highly polymorphic Simple Sequence Repeat markers were evaluated and proved to be very informative, producing a total number of 160 alleles with an average value of 10.7 per locus; the SSRs DCA09 and DCA16 were the most informative, distinguishing 17 and 19 genotypes, respectively. Phylogenetic and population structure analysis split the accessions in two main groups corresponding to most of oleasters and most of traditional varieties, respectively. Interestingly, ten traditional varieties resulted strictly related to the oleasters, indicating hybridization between the two botanical varieties. Genetic parameters and private alleles of groups confirmed this observation and indicated a wide genetic variability in Algerian olive germplasm. The results suggest the need to preserve and characterize this germplasm in order to limit the risk of losing potential important genetic traits present in the crop wild relatives.

A Distinct Genetic Cluster in Cultivated Chickpea as Revealed by Genome-wide Marker Discovery and Genotyping

Genotyping‐by‐sequencing analysis in cultivated chickpea generated 3187 high‐quality single nucleotide polymorphisms. Analysis of genetic diversity supports the identification of three subpopulations. Accessions traditionally grown in Italy form a clearly distinct genetic cluster. We identified genomic regions putatively resulting from directional selection. Our findings are of interest for chickpea conservation genetics and breeding.