Michele Zaccai

The water- and salt-stress-regulated Asr1 (abscisic acid stress ripening) gene encodes a zinc-dependent DNA-binding protein

Tomato (Lycopersicon esculantum) ASR1 (abscisic acid stress ripening protein), a small plant-specific protein whose cellular mode of action defies deduction based on its sequence or homology analyses, is one of numerous plant gene products with unknown biological roles that become over-expressed under water- and salt-stress conditions. Steady-state cellular levels of tomato ASR1 mRNA and protein are transiently increased following exposure of plants to poly(ethylene glycol), NaCl or abscisic acid. Western blot and indirect immunofluorescence analysis with anti-ASR1 antibodies demonstrated that ASR1 is present both in the cytoplasmic and nuclear subcellular compartments; approx. one-third of the total ASR1 protein could be detected in the nucleus. Nuclear ASR1 is a chromatin-bound protein, and can be extracted with 1 M NaCl, but not with 0.5% Triton X-100. ASR1, overexpressed in Escherichia coli and purified to homogeneity, possesses zinc-dependent DNA-binding activity. Competitive-binding experiments and SELEX (systematic evolution of ligands by exponential enrichment) analysis suggest that ASR1 binds at a preferred DNA sequence.

Potent antiviral flavone glycosides from Ficus benjamina leaves

Crude ethanol extracts from Ficus benjamina leaves strongly inhibit Herpes Simplex Virus 1 and 2 (HSV-1/2) as well as Varicella Zoster Virus (VZV) cell infection in vitro. Bioassay-guided fractionation of the crude extract demonstrated that the most efficient inhibition of HSV-1 and HSV-2 was obtained with the flavonoid fraction. The present study was aimed to further isolate, purify and identify substances with potent antiviral activity from the flavonoid fraction of F. benjamina extracts. Flavonoids were collected from the leaf ethanol extracts through repeated purification procedure and HPLC analysis. The antiviral activity of each substance was then evaluated in cell culture. Three known flavone glycosides, (1) quercetin 3-O-rutinoside, (2) kaempferol 3-O-rutinoside and (3) kaempferol 3-O-robinobioside, showing highest antiviral efficiency were selected and their structure was determined by spectroscopic analyses including NMR and mass spectrometry (MS). These three flavones were highly effective against HSV-1 reaching a selectivity index (SI) of 266, 100 and 666 for compound 1, 2 and 3, respectively, while the SI of their aglycons, quercetin and kaempferol amounted only in 7.1 and 3.2, respectively. Kaempferol 3-O-robinobioside showed similar SI to that of acyclovir (ACV), the standard anti-HSV drug. Although highly effective against HSV-1 and HSV-2, these flavone glycosides did not show any significant activity against VZV.

Antiviral activity of ethanol extracts of Ficus binjamina and Lilium candidum in vitro

The antiviral activity of plant ethanol extracts against Herpes Simplex Virus-1 and -2 (HSV-1 and HSV-2) and Varicella-Zoster Virus (VZV) was investigated in vitro. Ficus binjamina, resistant to plant viruses, and Lilium candidum, which has a high susceptibility to plant viruses were used. Leaf extracts of F. binjamina inhibited all studied viruses, while its fruit extracts inhibited only VZV. L. candidum leaf extracts had no effect on VZV but strongly inhibited HSV-1 and slightly HSV-2. None of the extracts showed significant cytotoxic effect on uninfected Vero cells even at a concentration of 250 microg/ml (CC(50)>400 microg/ml). The greatest antiviral effect was obtained when extracts were added to cells at the time of infection, whereas a partial inhibitory effect was observed when they were added post-infection. There was indirect evidence for strong interactions between the plant extracts and the viruses and weak interactions with the cell surface.

Potassium Chloride Enhances Fruit Appearance and Improves Quality of Fertigated Greenhouse Tomato as Compared to Potassium Nitrate

Abstract The effects of potassium chloride (KCl) as a potassium (K) source in fertigation solution on growth, yield and quality of tomato (cv. Durinta) in a controlled greenhouse were compared with potassium nitrate (KNO3)—the conventional K source for vegetable fertigation. The treatments consisted of four levels of KCl: (1) 0% KCl (100% KNO3), (2) 40% KCl (40% KCl and 60% KNO3), (3) 60% KCl (60% KCl and 40% KNO3), and (4) 100% KCl (0% KNO3) in fertigation solution in the season 1999–2000. In 2000–2001, early (12 days after planting) and late (47 days after planting) applications of 100% KCl and 0% KCl were tested. The concentrations of K and other major nutrients were similar in all the treatments. Ammonium nitrate (NH4NO3), calcium nitrate [Ca(NO3)2] and nitric acid (HNO3) were used as nitrogen (N) sources in KCl treatments. Electrical conductivity (EC) of all solutions ranged from 1.8 to 2.1 dS m−1; pH range was from 6.6 to 7.1. Perlite was used as a neutral growing medium. Plant height, time to anthesis, time to harvest, and leaf nutrient content were monitored. Total yield, average fruit weight and number, and fruit size were measured after harvest. The appearance and quality of the fruits were rated following cold storage simulation for export conditions. None of the plants showed chloride (Cl) toxicity symptoms. No significant differences in yield components and plant growth were recorded among the treatments. Fruit dry matter, total soluble solids (TSS), glucose, titrable acidity (TA), pH, and EC of juice after simulation storage were not affected by the K source. Interestingly, fruit firmness, and freshness of calyx were significantly improved, while the number of rotten and blotchy fruits was significantly decreased in KCl treatments. The fruit nitrate (NO3) content was decreased whereas iron (Fe) content was significantly increased in KCl treatments. The results show that KCl can be used as a substitute for KNO3 without detrimental effects on plant development and yield, while significantly improving some important quality parameters. It is concluded that KNO3 can be replaced fully or partially (depending on water quality) by KCl in tomato production while improving the quality of fruits.

Watermelon chlorotic stunt and Squash leaf curl begomoviruses—New threats to cucurbit crops in the Middle East

During the year 2002, two new diseases with unknown etiologies were detected in cucurbit crops in Israel. One disease was detected in squash fields throughout the country, while the second appeared in a single watermelon plot in the south, just outside the city of Elat. The infected watermelon plot was eradicated, but nonetheless the new disease spread throughout the country and today it is present in all the watermelon production areas. Both diseases were associated with elevated whitefly populations. Indeed, it was found that both are transmitted only by whiteflies, and are incited by two begomoviruses. Both viruses were cloned and sequenced, and were identified as Watermelon chlorotic stunt virus (WmCSV) and Squash leaf curl virus (SLCV). The host range of the Israeli WmCSV isolate (WmCSV-IL) was determined and resembles the host range of the three other known WmCSV isolates (from Yemen, Sudan, and Iran), but with a few differences. Although no genetic resistance to WmCSV was observed in cultivated wat...

Anti-Herpetic Activity of Callissia fragrans and Simmondsia chinensis Leaf Extracts In Vitro

The antiviral activity of Callissia fragrans and Simnondsia chinensis aquatic and ethanol leaf extracts, as well as purified fractions from these extracts was studied against herpetic viruses in vitro. Ethanol extract of C. fragrans effectively inhibited the infection of Vero cells by HSV-1, HSV-2 in vitro, while its aquatic extract inhibited only VZV. Although S. chinensis leaf extract strongly inhibited all studied viruses, the selectivity index of this extract was very low, due to its high toxicity. However, the majority of its fractions showed low toxicity and higher antiviral activity and therefore very high SI. Strong interactions between virus and extracts were found.

Characterization of expressed sequence tags from Lilium longiflorum in vernalized and non-vernalized bulbs.

In Lilium longiflorum, vernalization is both an obligatory requirement and the major factor affecting flowering time, however, little is known about the molecular regulation of this mechanism in Lilium and other flowering bulbs. Exposure of L. longiflorum bulbs to 9 weeks at 4°C greatly promoted stem elongation within the bulb, floral transition and flowering. Subtraction libraries of vernalized (V) and non-vernalized (NV) bulb meristems were constructed. 671 and 479 genes were sequenced, from which 72 and 82 proteins were inferred for the NV-V and the V-NV libraries, respectively. Much lower transcription levels and putative gene functions were recorded in the NV-V libraries compared the V-NV libraries. However, a large number of genes annotated to transposable elements (TEs), represented more than 20% of the sequenced cDNA were expressed in the NV-V libraries, as opposed to less than 2% in the V-NV libraries. The expression profile of several genes potentially involved in the vernalization pathway was assessed. Expression of LlSOC1, the lily homologue of SUPPRESSOR OF OVER-EXPRESSION OF CO1 (SOC1), an important flowering gene in several plant species, found in the V-NV library, was highly up-regulated during bulb meristem cold exposure. The subtraction libraries provided a fast tool for relevant gene isolation.

Analysis of phenotypic and genetic polymorphism among accessions of saltgrass (Distichlis spicata)

Morphological and genetic analyses were applied on 37 saltgrass (Distichlis spicata) accessions collected from distant locations throughout the American continent in order to: 1) develop and identify morphologic and genetic profiles for the saltgrass genotypes; 2) characterize the genetic distance among saltgrass accessions within the given germplasm collection; and 3) identify a possible linkage between patterns of genetic and eco-geographical parameters. Analysis based on 70 RAPD markers revealed broad polymorphism among the genotypes and enabled their individual characterization. A UPGMA dendrogram clustered the genotypes into groups according to a general pattern of their geographical origin. Yet, only the group of Californian accessions was significantly distinct from other groups, as determined by χ2 tests. Various statistical analyses indicated that only minor genetic differences existed between seashore and desert saltgrass genotypes, supporting that these two eco-geographical types belong to the same species, Distichlis spicata. Our results suggest that genetic interactions exist between geographically distant saltgrass populations, in spite of the dominance of clonal reproduction in this species. The contribution of dioecy, seed production, and epizoochory to the distribution patterns of saltgrass is discussed at the geographic and at the genetic levels.

Anti-Herpetic Activity of and Leaf Extracts

The antiviral activity of Callissia fragrans and Simnondsia chinensis aquatic and ethanol leaf extracts, as well as purified fractions from these extracts was studied against herpetic viruses in vitro. Ethanol extract of C. fragrans effectively inhibited the infection of Vero cells by HSV-1, HSV-2 in vitro, while its aquatic extract inhibited only VZV. Although S. chinensis leaf extract strongly inhibited all studied viruses, the selectivity index of this extract was very low, due to its high toxicity. However, the majority of its fractions showed low toxicity and higher antiviral activity and therefore very high SI. Strong interactions between virus and extracts were found.

Functional genomics to isolate genes involved in fragrance production for genetic engineering of scent in flowers.

Scents of flowers are usually made of mixtures of hundreds (or even thousands) of volatile compounds, normally emitted from flowers to attract pollinators. Different varieties of the same plant may emit a completely different array of compounds (Vainstein et al., 2001). Not only the presence or absence of an individual component might affect a particular flower scent, at times the same compound might have an agreeable or disagreeable scent depending on its concentration. Most of the research in flower scent has been aimed at elucidating the chemical structures of key scent components and in attempting their chemical synthesis for use in the perfumery and cosmetics industries. Despite the vast number of chemical structures involved, the large majority of scent compounds are biosynthesized by a surprisingly small number of metabolic pathways. These metabolic pathways are often ubiquitous, and specialization has developed through small but important modifications of ancestral genes and pathways (Pichersky and Gang, 2000).