Nelson Marmiroli

Molecular genetics of heat tolerance and heat shock proteins in cereals.

Heat stress is common in most cereal-growing areas of the world. In this paper, we summarize the current knowledge on the molecular and genetic basis of thermotolerance in vegetative and reproductive tissues of cereals. Significance of heat stress response and expression of heat shock proteins (HSPs) in thermotolerance of cereal yield and quality is discussed. Major avenues for increasing thermotolerance in cereals via conventional breeding or genetic modification are outlined.

Tuberization in potato involves a switch from apoplastic to symplastic phloem unloading

Phloem unloading was studied in potato plants in real time during the early stages of tuberization using carboxyfluorescein (CF) as a phloem-mobile tracer, and the unloading pattern was compared with autoradiography of tubers that had transported 14C assimilates. In stolons undergoing extension growth, apoplastic phloem unloading predominated. However, during the first visible signs of tuberization, a transition occurred from apoplastic to symplastic transport, and both CF and 14C assimilates subsequently followed identical patterns of phloem unloading. It is suggested that the switch to symplastic sucrose unloading may be responsible for the upregulation of several genes involved in sucrose metabolism. A detailed analysis of sugar levels and 14C sugar partitioning in tuberizing stolons revealed a distinct difference between the apical region of the tuber and the subapical region. Analysis of invertase activity in nontuberizing and tuberizing stolons revealed a marked decline in soluble invertase in the subapical region of swelling stolons, consistent with the switch from apoplastic to symplastic unloading. However, cell wall–bound invertase activity remained high in the apical 1 to 2 mm of tuberizing stolons. Histochemical analysis of potato lines transformed with the promoter of an apoplastic invertase gene (invGE) linked to a reporter gene also revealed discrete gene expression in the apical bud region. Evidence is presented that the apical and lateral tuber buds function as isolated domains with respect to sucrose unloading and metabolism.

Accumulation of copper and zinc from liquid manure in agricultural soils and crop plants

In Northern Italy it is a common practice to utilise slurries and other manure from intensive animal farming to fertilise agricultural soils. Due to the relatively high copper and zinc contents of these materials, this practice could lead to contamination of soils and the crops grown on them. In this study we have evaluated the extent of copper and zinc contamination of soils subjected to different levels of fertilisation with liquid manure and the copper and zinc concentrations in the edible tissues of three crops (maize, sugar beet and lucerne) grown on the same soils. There was a direct correlation between soil copper and zinc contents and the levels of application of animal wastes and copper concentrations were high enough to represent a risk, according to current European legislation. The metal contents in the edible tissues of the three crops were relatively low and variable, but showed no clear correlations with the intensity of liquid manure applications. Overall, there was no risk of contamination of the food chain, as all concentrations were well below levels considered to be toxic to animals.

Application of a horizontal subsurface flow constructed wetland on treatment of dairy parlor wastewater

Two horizontal subsurface flow reed beds of 75 m(2) each, treating dairy parlor effluent and domestic sewage (about 6.5 m(3)/day), were set-up to determine the efficiency of this system in reducing the polluting load in an isolated mountain rural settlement.A total suspended solids value of about 0.70 g/l and chemical oxygen demand (COD) and biological oxygen demand (BOD(5)) values of about 1200 and 450 mg/l O(2), respectively, were characteristic of the influent waters. Removal of suspended solids and organic load constantly remained at levels above 90%, while those of the nutrients N and P were about 50% and 60%, respectively. The total number of coliform bacteria and Escherichia coli was reduced by more than 99%, and faecal streptococci by more than 98%. Nitrates, chlorides, sulfates, anionic and non-ionic surface-active agents and heavy metals were detected only in low concentrations. Concentration and localization of metals was also quantified in Phragmites tissues by microanalysis.Results demonstrated the use of reed beds as an appropriate treatment to reduce pollutants in wastewater from rural activities to values acceptable for discharge into surface waters.

Methods for detection of GMOs in food and feed

This paper reviews aspects relevant to detection and quantification of genetically modified (GM) material within the feed/food chain. The GM crop regulatory framework at the international level is evaluated with reference to traceability and labelling. Current analytical methods for the detection, identification, and quantification of transgenic DNA in food and feed are reviewed. These methods include quantitative real-time PCR, multiplex PCR, and multiplex real-time PCR. Particular attention is paid to methods able to identify multiple GM events in a single reaction and to the development of microdevices and microsensors, though they have not been fully validated for application.

Location of a gene regulating drought-induced abscisic acid production on the long arm of chromosome 5A of wheat.

The accumulation of abscisic acid (ABA) by detached and partially dehydrated wheat leaves is known to be inherited in a quantitative manner. The location of genes having a major effect on drought-induced ABA accumulation in wheat was determined using a set of single chromosome substitution lines and populations derived from a cross between a high-ABA- and a low-ABA-producing genotype. Examination of a series of chromosome substitution lines of the high-ABA genotype ‘Ciano 67’ into the low-ABA recipient ‘Chinese Spring’ showed that chromosome 5A carries gene(s) that have a major influence on ABA accumulation in a drought test with detached and partially dehydrated leaves (DLT). A similar DLT was used to examine ABA accumulation in a population of F2 plants and doubled haploid (DH) lines derived from the cross between ‘Chinese Spring’ (low-ABA) and ‘SQ1’ (high-ABA) in which the F2 population (139 plants) and DH lines (96 lines) were also mapped partially with molecular markers. Analysis of variance of ABA accumulation between and within marker allele classes in the F2 confirmed the location of a gene(s) regulating ABA accumulation on the long arm of chromosome 5A. MAPMAKERQTL showed the most likely position for the ABA quantitative trait locus (QTL) to be between the loci Xpsr575 and Xpsr426, about 8 cM from Xpsr426. A similar trend for high ABA accumulation was found in DH lines having the ‘SQ1’ allele at marker loci in the same region of chromosome 5AL, but the QTL effect was not significant. The function of the QTL is discussed.

Biochar addition to an arsenic contaminated soil increases arsenic concentrations in the pore water but reduces uptake to tomato plants (Solanum lycopersicum L.).

Arsenic (As) concentrations in soil, soil pore water and plant tissues were evaluated in a pot experiment following the transplantation of tomato (Solanum lycopersicum L.) plantlets to a heavily As contaminated mine soil (~6000 mg kg(-1) pseudo-total As) receiving an orchard prune residue biochar amendment, with and without NPK fertiliser. An in-vitro test was also performed to establish if tomato seeds were able to germinate in various proportions of biochar added to nutrient solution (MS). Biochar significantly increased arsenic concentrations in pore water (500 μg L(-1)-2000 μg L(-1)) whilst root and shoot concentrations were significantly reduced compared to the control without biochar. Fruit As concentrations were very low (<3 μg kg(-1)), indicating minimal toxicity and transfer risk. Fertilisation was required to significantly increase plant biomass above the control after biochar addition whilst plants transplanted to biochar only were heavily stunted and chlorotic. Given that increasing the amount of biochar added to nutrient solution in-vitro reduced seed germination by up to 40%, a lack of balanced nutrient provision from biochar could be concluded. In summary, solubility and mobility of As were increased by biochar addition to this soil, but uptake to plant was reduced, and toxicity-transfer risk was negligible. Therefore leaching rather than food chain transfer appears the most probable immediate consequence of biochar addition to As contaminated soils.

Growth, physiological and molecular traits in Salicaceae trees investigated for phytoremediation of heavy metals and organics

Worldwide, there are many large areas moderately contaminated with heavy metals and/or organics that have not been remediated due to the high cost and technical drawbacks of currently available technologies. Methods with a good potential for coping with these limitations are emerging from phytoremediation techniques, using, for example, specific amendments and/or plants selected from various candidates proven in several investigations to be reasonably efficient in extracting heavy metals from soil or water, or in co-metabolizing organics with bacteria flourishing or inoculated in their rhizospheres. Populus and Salix spp., two genera belonging to the Salicaceae family, include genotypes that can be considered among the candidates for this phytoremediation approach. This review shows the recent improvements in analytical tools based on the identification of useful genetic diversity associated with classical growth, physiological and biochemical traits, and the importance of plant genotype selection for enhancing phytoremediation efficiency. Particularly interesting are studies on the application of the phytoremediation of heavy metals and of chlorinated organics, in which microorganisms selected for their degradation capabilities were bioaugmented in the rhizosphere of Salicaceae planted at a high density for biomass and bioenergy production.

Stable changes in CD4+ T lymphocyte miRNA expression after exposure to HIV-1

MicroRNAs (miRNAs) inhibit HIV-1 expression by either modulating host innate immunity or by directly interfering with viral mRNAs. We evaluated the expression of 377 miRNAs in CD4(+) T cells from HIV-1 élite long-term nonprogressors (éLTNPs), naive patients, and multiply exposed uninfected (MEU) patients, and we observed that the éLTNP patients clustered with naive patients, whereas all MEU subjects grouped together. The discriminatory power of miRNAs showed that 21 miRNAs significantly differentiated éLTNP from MEU patients and 23 miRNAs distinguished naive from MEU patients, whereas only 1 miRNA (miR-155) discriminated éLTNP from naive patients. We proposed that miRNA expression may discriminate between HIV-1-infected and -exposed but negative patients. Analysis of miRNAs expression after exposure of healthy CD4(+) T cells to gp120 in vitro confirmed our hypothesis that a miRNA profile could be the result not only of a productive infection but also of the exposure to HIV-1 products that leave a signature in immune cells. The comparison of normalized Dicer and Drosha expression in ex vivo and in vitro condition revealed that these enzymes did not affect the change of miRNA profiles, supporting the existence of a Dicer-independent biogenesis pathway.

Multiplex real-time PCR using SYBR® GreenER™ for the detection of DNA allergens in food

AbstractWe describe the development of a six-target real-time multiplex PCR assay with the SYBR® GreenER™ fluorescent dye, targeted to genes encoding for allergenic proteins commonly present in many processed food products (patent application pending). The assay was successfully trialled on reconstructed food matrices and on a range of commercial foodstuffs, and is proposed as a ready-to-use analytical tool for food manufacturers to detect the presence or confirm the absence of sequences encoding for important allergenic proteins of plant origin. FigureMultiplex real-time PCR for the detection of DNA allergens in food using ABI Prism® 7000 Sequence detection system