Characterization of Some Italian Common Bean (<Emphasis Type="Italic">Phaseolus Vulgaris</Emphasis> L.) Landraces by RAPD,Semi-random and ISSR Molecular Markers

Randomly amplified polymorphic DNA (RAPD), inter-simple sequence repeat (ISSR) and a semi-random PCR system were used to analyze the genetic diversity of 16 Italian common bean landraces and their relationship to four commercial cultivars. Of the primers tested, 8 ISSR, 6 RAPD and 7 semi-random primers produced polymorphic and reproducible DNA fragments. A higher proportion of polymorphic bands were observed using ISSR (85%) and semi-random (90%) primers than RAPD (69%) method. The combination of any two semi-random markers allowed the identification of all 20 bean genotypes. In contrast ISSR (except for primer (CAC)₃GC) and RAPD markers appeared to be less informative as more than two markers were necessary to achieve the same diagnostic level. Moreover, 7 ISSR, 2 RAPD and 8 semi-random exclusive bands were identified as putative population-specific markers. Semi-random and ISSR derived dendrograms showed similar tendencies in terms of genetic relatedness, whereas clustering of genotypes within groups was not similar when compared with the RAPD technique. Despite the different ability to resolve genetic variation among the investigated landraces, two major clusters with less than 60% (ISSR) and 40% (RAPD and semi-random) genetic similarity were formed with all three marker systems. The two groups were correlated with the phaseolin patterns and seed size of the landraces. The analysis showed that the cultivar ȁ8Lingua di Fuocoȁ9 and most of the landraces (13 out of 16) collected in Italy belong to the Andean gene pool, whereas only the three populations from Pratomagno belong to the Middle American gene pool.     

Effect of wine inhibitors on free pineapple stem bromelain activity in a model wine system

The influence of potential inhibitors, naturally present in wine, on the activity of stem bromelain was investigated in order to evaluate the applicability of this enzyme for protein stabilization in white wine. Bromelain proteolytic activity was tested against a synthetic substrate (Bz-Phe-Val-Arg-pNA) in a model wine system after adding ethanol, sulfur dioxide (SO(2)), skin, seed, and gallic and ellagic tannins at the average range of their concentration in wine. All the inhibitors of stem bromelain activity tested turned out to be reversible. Ethanol was a competitive inhibitor with a rather limited effect. Gallic and ellagic tannins have no inhibitory effect on stem bromelain activity, while both seed and skin tannins were uncompetitive inhibitors. The strongest inhibition effect was revealed for sulfur dioxide, which was a mixed-type inhibitor for the enzyme activity. This study provides useful information relative to a future biotechnological application of stem bromelain in winemaking.     

A multivariate approach for anomaly separation of potentially toxic trace elements in urban and peri-urban soils: an application in a southern Italy area

Purpose

Geogenic soil enrichment and anthropogenic pollution by potentially toxic trace elements (PTEs) are two processes acting together. Although it is often difficult, it is necessary to separate the two processes for risk assessment and understanding the environmental implications. The aim of this study was to analyse the soil concentrations of various PTEs in a southern Italy area in order to: (1) determine their different correlation structure to isolate sources of variation acting at different spatial scales and (2) to define potential anomalies based on the correlation structure.

Materials and methods

In the urban and peri-urban area of Cosenza-Rende, 149 topsoil samples were collected (0.10 m) and analysed for different elements by X-ray fluorescence spectrometry. Principal component analysis and factorial kriging analysis were used to map the spatial distribution of PTEs in topsoil and to identify the main factors influencing their spatial variability.

Results and discussion

Two groups of PTEs were identified: the first group included As, Pb and Zn; and the second one Al, Co, Cr, Fe, La, Nb, Ni, Ti and V. The first group was related to anthropogenic causes, while the second one was more related to parent rock composition. The regionalized factors at different scales of variability allowed to aggregate and summarize the joint variability in the PTEs and consider the probable causes of soil pollution.

Conclusions

The study allowed analysing and quantifying the sources (environmental or anthropogenic) of variation of PTEs acting at different spatial scale and defining the spatial anomalies based on the correlation structure associated at the different spatial scales.     

The contribution of stem cells to epidermal and hair follicle tumours in the dog

Background –  Although cutaneous stem cells have been implicated in skin tumourigenesis in humans, no studies have been conducted to elucidate the presence and the possible role of stem cells in hair follicle tumours in the dog. Hypothesis –  Stem cell markers are expressed in canine epidermal and follicular tumours and can be used to better understand the biology and origin of these tumours. Animals and Methods –  In the present study, normal skin sections and 44 follicular tumours were retrospectively investigated for the immunohistochemical expression of keratin 15 (K15) and nestin. In addition, 30 squamous cell carcinomas were evaluated for K15 expression. Results –  In normal skin, K15 and nestin were expressed in the outer root sheath cells of the isthmic portion of the hair follicle (bulge region), and K15 expression was also scattered in the basal cell layer of the epidermis. Infundibular keratinizing acanthomas, pilomatricomas and squamous cell carcinomas were mostly negative for K15, trichoblastomas were moderately to strongly positive, tricholemmomas were either negative or strongly positive, and trichoepitheliomas had heterogeneous staining. Nestin expression was generally faint in all follicular tumours. Conclusions and clinical importance –  Our results show that K15 can be a reliable marker for investigating the role of stem cells in hair follicle tumours of the dog, while nestin was judged to be a nonoptimal marker. Furthermore, our study suggests that hair follicle stem cells are present in the bulge region of hair follicles and could possibly play a role in tumourigenesis of canine tumours originating from this portion of the follicle, namely trichoblastomas, tricholemmomas and trichoepitheliomas. The loss of K15 expression in squamous cell carcinomas compared with normal skin suggests that this event could be important in the malignant transformation.     

Impact of the biocontrol agent Trichoderma atroviride SC1 on soil microbial communities of a vineyard in northern Italy

The fungus Trichoderma atroviride SC1 is an experimental biocontrol agent (BCA) that is active against the fungus Armillaria mellea. Following the application of Trichoderma to the surface soil of a vineyard, we used a highly specific real-time PCR, previously validated for the analysis of soil microcosms, to monitor the populations of this fungus at different soil depths over several months. The quantification obtained using this molecular method was highly correlated with laboratory assays of colony-forming units. The native communities of bacteria and fungi in the soil were analyzed using automated ribosomal intergenic spacer analysis (ARISA), and transient changes were observed following the application of T. atroviride SC1 conidia. A principal component of variance analysis demonstrated that the introduction of T. atroviride SC1 had an effect on the soil microflora during the first two weeks following inoculation. However, at later dates, environmental conditions had a higher influence on the surveyed communities than the BCA application, as confirmed through the use of the Shannon index of biodiversity. Soil depth had a strong influence on the composition and biodiversity of fungal communities.     

Characterization and antimicrobial activity of essential oils of industrial hemp varieties (Cannabis sativa L.)

The present study focused on inhibitory activity of freshly extracted essential oils from three legal (THC < 0.2% w/v) hemp varieties (Carmagnola, Fibranova and Futura) on microbial growth. The effect of different sowing times on oil composition and biological activity was also evaluated. Essential oils were distilled and then characterized through the gas chromatography and gas chromatography-mass spectrometry. Thereafter, the oils were compared to standard reagents on a broad range inhibition of microbial growth via minimum inhibitory concentration (MIC) assay. Microbial strains were divided into three groups: i) Gram (+) bacteria, which regard to food-borne pathogens or gastrointestinal bacteria, ii) Gram (?) bacteria and iii) yeasts, both being involved in plant interactions. The results showed that essential oils of industrial hemp can significantly inhibit the microbial growth, to an extent depending on variety and sowing time. It can be concluded that essential oils of industrial hemp, especially those of Futura, may have interesting applications to control spoilage and food-borne pathogens and phytopathogens microorganisms.     

Methods of assessing the incidence of Armillaria root rot across viticultural areas and the pathogen's genetic diversity and spatial–temporal pattern in northern Italy

Armillaria mellea (Vahl:Fr.) P. Kumm. is the causal agent of root rots of numerous perennial woody plants, including grapevine. In grapevine, A. mellea infections cause reduced plant vigour, poor grape quality and, in the last stage of the disease, plant death. No post-infection control agents are available for use against this disease. Disease management programmes rely solely on prophylactic measures. Therefore, knowledge of the distribution pattern of the disease over time, disease incidence and actual losses due to the disease across geographical areas is crucial. We evaluated the use of aerial photographs, surveys of growers’ experiences and field assessments to estimate the incidence of Armillaria root rot in a viticultural area (140 ha) situated in the Piana Rotaliana region (Italy). Results of the 4-year assessment were that 25% of the vineyards (36 ha) were infected by Armillaria, 2% of the cultivated vines exhibited disease symptoms and 0.25% of the vines died each autumn. The causal agent was almost exclusively A. mellea but, in two cases, Armillaria gallica was also isolated. The genetic diversity of the Italian population, according to IGS sequencing and somatic incompatibility testing, was very high, suggesting multiple pathogen introduction sites and the slow, vegetative underground spread of the fungus. Fungal clones were found within individual vineyards. Even if, within a particular area, the disease is spreading very slowly and crop losses are limited (one vine out of 400 dies each year), an effective agronomic plan for the prevention of infections should be implemented.     

A new technique to measure soil CO2 efflux at constant CO2 concentration

A new principle for measuring soil CO₂ efflux at constant ambient concentration is introduced. The measuring principle relies on the continuous absorption of CO₂ within the system to achieve a constant CO₂ concentration inside the soil chamber at ambient level, thus balancing the amount of CO₂ entering the soil chamber by diffusion from the soil. We report results that show reliable soil CO₂ efflux measurements with the new system. The novel measuring principle does not disturb the natural gradient of CO₂ within the soil, while allowing for continuous capture of the CO₂ released from the soil. It therefore holds great potential for application in simultaneous measurements of soil CO₂ efflux and its δ¹³C, since both variables show sensitivity to a distortion of the soil CO₂ profile commonly found in conventional chamber techniques.