Change in membrane fatty acid compositions and cold-induced responses in chickpea

Plant cells often increase cold tolerance by reprogramming their genes expression which results in adjusted metabolic alternations, a process enhanced under cold acclimation (CA) phase. In present study, we assessed the changes of membrane fatty acid compositions and defense machine (like antioxidative enzymes) along with damage indexes like electrolyte leakage index (ELI) and malondialdehyde (MDA) during CA, cold stress (CS) and recovery (R) phases in chickpea (Cicer arietinum L.). Results showed an increase in unsaturated fatty acids ratio compare to saturated ones which is a sign of cold tolerance especially after CA phase. Antioxidant enzymes had an important role during CA and R phases while CS affected their activity which can be a sign for associating other metabolites or enzymes activities to create cold tolerance in plants. To investigation of enzymes assay under experimental treatments, the expression pattern of some enzymes including superoxide dismutase (sod), catalase (cat) and lipoxygenase (lox) was studied using quantitative real time PCR. LOX activity has shown a bilateral behavior: a positive relation with membrane damage index in CA and an interesting link with double bond index (DBI) in CS indicating probably its role in secondary metabolites like jasmonic acid signaling pathway. It was suggested that increased DBI and low LOX activity under CS could be a reason for plant cold tolerance.     

Response of antioxidative enzymes and apoplastic bypass transport in Thlaspi caerulescens and Raphanus sativus to cadmium stress

A hydroponics experiment using hyperaccumulator Thlaspi caerulescens (alpine pennycress) and non-specific accumulator Raphanus sativus (common radish) was conducted to investigate the short-term effect of increasing Cd concentrations (0, 25, 50, 75, 100 microM) on metal uptake, chlorophyll content, antioxidative enzymes, and apoplastic bypass flow. As expected, T. caerulescens generally showed better resistance to metal stress, which was reflected by higher Cd accumulation within plant tissues with no signs of chlorosis, or wilt. Glutathione reductase (GR) and superoxide dismutase (SOD) activities in fresh leaves were monitored as the plant metal-detoxifying response. In general, both plant species exhibited an increase trend of GR activity before declining at 100 microM likely due to excessive levels of phytotoxic Cd. SOD activity exhibited almost a similar variation pattern to GR and decreased also at 100 microM Cd. For both plant species, fluorescent PTS uptake (8-hydroxy-1,3,6-pyrenetrisulphonic acid) increased significantly with metal level in exposure solutions indicating that Cd has a comparable effect to drought or salinity in terms of the gain of relative importance in apoplastic bypass transport under such stress conditions.     

Comparative expression in Escherichia coli of the native and hybrid genes for acyl-lipid delta(9) desaturase

Expression of the desC gene coding for acyl-lipid delta(9) desaturase of thermophilic cyanobacterium Synechocystis sp. PCC6803 was studied in Escherichia coli cells. In a hybrid gene constructed (desC-licBM3), a sequence of the native acyl-lipid delta(9) desaturase was fused in frame with the reporter gene coding for thermostable lichenase. Lichenase contained in the hybrid protein simplified selection and analysis of the expression of membrane desaturase in the heterologous host. Comparisons of the expression for the native and hybrid genes in bacterial cells showed that lichenase remained active and thermostable in the hybrid protein, while desaturase retains the capability of introducing a double bound in the corresponding position of fatty acids.     

Comparative expression in <Emphasis Type="Italic">Escherichia coli</Emphasis> of the native and hybrid genes for acyl-lipid Δ<Superscript>9</Superscript> desaturase

Expression of the desC gene coding for acyl-lipid Δ⁹ desaturase of thermophilic cyanobacterium Synechocystis sp. PCC6803 was studied in Escherichia coli cells. In a hybrid gene constructed (desC-licBM3), a sequence of the native acyl-lipid Δ⁹ desaturase was fused in frame with the reporter gene coding for thermostable lichenase. Lichenase contained in the hybrid protein simplified selection and analysis of the expression of membrane desaturase in the heterologous host. Comparisons of the expression for the native and hybrid genes in bacterial cells showed that lichenase remained active and thermostable in the hybrid protein, while desaturase retains the capability of introducing a double bound in the corresponding position of fatty acid residues.     

Molecular detection of nematicidal crystalliferous Bacillus thuringiensis strains of Iran and evaluation of their toxicity on free-living and plant-parasitic nematodes

The characterization of nematode-effective strains and cry genes in the Iranian Bacillus thuringiensis (Bt) collection (70 isolates) is presented. Characterization was based on PCR analysis using 12 specific primers for cry5, cry6, cry12, cry13, cry14, and cry21 genes encoding proteins active against nematodes, crystal morphology, and protein band patterns as well as their nematicidal activity on root-knot nematode (Meloidogyne incognita) and two free-living nematodes (Chiloplacus tenuis and Acrobeloides enoplus). PCR results with primers for these genes showed that 22 isolates (31.5%) contain a minimum of one nematode-active cry gene. Strains containing the cry6 gene were the most abundant and represent 22.8% of the isolates. Bt strains harboring cry14 genes were also abundant (14.2%). cry21 and cry5 genes were less abundant, found in 4.2% and 2.8% of the strains, respectively. In total, six different nematode-active cry gene profiles were detected in this collection. Four isolates did not show the expected PCR product size for cry5, cry6, and cry21 genes; they might contain potentially novel insecticidal crystal protein genes. Twenty-two Bt isolates containing nematode-active cry genes were selected for preliminary bioassays on M. incognita. Based on these bioassays, four isolates were selected for detailed bioassays. Isolates YD5 and KON4 at 2 x 10(8) CFU/mL concentrations showed 77% and 81% toxicity on M. incognita, respectively. The free-living nematodes C. tenuis and A. enoplus were more susceptible and the highest mortality was observed within 48 h of incubation at all of the concentrations tested. Maximum mortality was recorded for isolates SN1 and KON4 at 2 x 10(8) CFU/mL concentrations and resulted in 68% and 77% adults deaths of C. tenuis and 68% and 72% for A. enoplus, respectively. Our results showed that PCR is a useful technique for toxicity prediction of nematicidal Bt isolates.     

Epidemiological and clinical studies of ovine caseous lymphadenitis

A field survey was undertaken, to determine epidemiological, clinical and biological data on Ovine Caseous Lymphadenitis disease in 54 flocks originated from Sfax area in Tunisia. The mean morbidity rate of the cutaneous form was 5.1%. This form affected sheep over 3 months and under 2 years old. On the other hand, the mean morbidity rate of the visceral form, encountered in abattoirs, was 11.02%. The clinical aspect of the superficial (or cutaneous) form was often corresponding to one abscess, located particularly in the lymphatic nodes of the animal's head; while visceral (or internal) form of the disease was represented by the presence of a unique abscess found in the pulmonary lymphatic nodes. The macroscopic aspect of lesions showed that the size of abscess was comprised between 4 and 10 cm in diameter. At the cut, colour of the pus was white yellowish to yellow greyish with a fluid or a thick aspect like onion peels; pus was microscopic. Lesions were characterised by a fibrous shell, a pyogenic membrane and a necrotic center. Bacteriological research revealed that Corynebacterium pseudotuberculosis was the pathogen the most frequently isolated, followed by Staphylococcus aureus subsp anaerobius which was particularly found in sheep aged between 3 months and 2 years old.     

Experimental models for creation of transgenic plants resistant to stressors

Experimental models of primary potato transgenic plants that express the cry3aM-licBM2 hybrid gene were created. The molecular analysis and biotests of the experimental models allow a new system of cry genes expression in plants to be proposed. This system is based on the expression of hybrid genes containing the reporter lichenase gene sequence and the use of a light-induced promoter ensuring preferential expression of the regulated genes only in green plant tissues (leaves), the target tissues for pests, as a regulatory element. In is shown that the presence of lichenase in hybrid proteins facilitates selection and analysis of the level of expression of hybrid proteins in transgenic plants. Judging by the properties of the reporter protein lichenase in hybrid proteins, it seems possible to use this reporter system for transgene monitoring in agrocenosis, because this system is fairly simple and precise and does not need considerable material and time expenses.     

Experimental models for creation of transgenic plants resistant to stress factors

Experimental models of the potato primary transgenic plants which express the hybrid gene cry3aM-licBM2 have been created. Modecular analysis and the biotests of the experimental models allow proposing a new system of cry genes expression in plants. The system is based on the expression of hybrid genes possessing the sequence of reporter lichenase gene and the use as a regulator element of a light-induced promoter providing preferential expression of the controled genes only in green plant tissues (leaves)--the target tissues for pests. The lichanase presence in hybrid proteins facilitates selection and analysis of the expression level of the hybris proteins in transgenic organisms. Basing on the lichenase properties in hybrid proteins it seems possible to use this reporter system for transgene monitoring in agrocoenosis as this system is rather simple and precise and does not need large material and time expenses.