Impeded Carbohydrate Metabolism in Rice Plants under Submergence Stress

The detrimental effects of submergence on physiological performances of some rice varieties with special references to carbohydrate metabolisms and their allied enzymes during post-flowering stages have been documented and clarified in the present investigation. It was found that photosynthetic rate and concomitant translocation of sugars into the panicles were both related to the yield. The detrimental effects of the complete submergence were recorded in generation of sucrose, starch, sucrose phosphate synthase and phosphorylase activity in the developing panicles of the plants as compared to those under normal or control （i.e. non-submerged） condition. The accumulation of starch was significantly lower in plants under submergence and that was correlated with ADP-glucose pyrophosphorylase activity. Photosynthetic rate was most affected under submergence in varying days of post-flowering and was also related to the down regulation of Ribulose bisphosphate carboxylase activity. However, under normal or control condition, there recorded a steady maintenance of photosynthetic rate at the post-flowering stages and significantly higher values of Ribulose bisphosphate carboxylase activity. Still, photosynthetic rate of the plants under both control and submerged conditions had hardly any significant correlation with sugar accumulation and other enzymes of carbohydrate metabolism like invertase with grain yield. Finally, plants under submergence suffered significant loss of yield by poor grain filling which was related to impeded carbohydrate metabolism in the tissues. It is evident that loss of yield under submergence is attributed both by lower sink size or sink capacity （number of panicles, in this case） as well as subdued carbohydrate metabolism in plants and its subsequent partitioning into the grains.

Physio-Biochemical and Genetic Exploration for Submergence Tolerance in Rice (<i>Oryza sativa</i>L.) Landraces with Special References to <i>Sub</i>1 Loci

In the present study a group of four indigenous and less popular rice genotypes (Meghi, Panibhasha, Jabra and Sholey) reported by growers as submergence tolerant lines from flood prone areas of south Bengal were explored through study of nodal anatomy, physio-biochemical screening under submergence and genotyping with submergence tolerance linked rice microsatellite loci (RM loci). To identify the different allelic forms of different Sub1 compnents (Sub1A, Sub1B and Sub1C) among the studied lines, the genomic DNA of individual genotypes was amplified with three ethylene response factor like genes from Sub1 loci, located on rice chromosome 9. From the different physio-biochemical experiments performed in this investigation, it has been shown that Meghi and Jabra are the two probable potent genotypes which share common properties of both submergence tolerant and deep water nature whereas rest two genotypes (Sholey and Panibhasha) behave like typical deep water rice. The submergence tolerance property of Meghi was also confirmed from submergence tolerance linked SSR based genotyping by sharing with FR13A for some common alleles as reflected in fingerprint derived dendrogram. The rest of the genotypes shared a number of alleles and were included in a separate cluster. The common behaviour of Meghi and FR13A under submergence was also confirmed from genetic study of Sub1 loci through sharing of some common alleles for three Sub1 components (Sub1A, Sub1B and Sub1C loci). One SSR loci (RM 285) was identified as a potent molecular marker for submergence tolerance breeding programme involving these two selected rice lines (Meghi and Jabra) as donor plant through marker assisted selection.

An Exploratory Study on Allelic Diversity for Five Genetic Loci Associated with Floral Organ Development in Rice

Allelic diversity for five genetic loci (DL, FON4, OsMADS24, OsMADS45 and Spw1) associated with floral organ development were investigated among a small heterogeneous rice population which included one wild species (O. rufipogon Griffiths), one indigenous less popular natural floral organ mutant (O. sativa var. indica cv. Jugal), one indigenous normal line (O. sativa var. indica cv. Bhutmoori) and one improved high yielding line (O. sativa var. indica cv. IR 36). Detailed spikelet morphology showed that var. Jugal had variable number (1 - 3) of carpels within a single spikelet which was unique and resulted in variable (1 - 3) number of kernels within a single matured spikelet (grain). The genomic DNA of each investigated line was amplified with primer sequences designed from the selected genetic loci and the derived polymorphism profiles were used for study of allelic diversity for the studied loci. The derived genetic distances among the rice lines were used for dendrogram construction. In constructed dendrogram, the mutant genotype (Jugal) showed highest similarity with the wild rice (O. rufipogon) instead of the rice lines. To verify this finding, the genomic DNA of each studied line was also amplified with four SSR loci, tightly linked to saltol QTL, mapped to rice chromosome 1. The amplified products were screened for polymorphism and another dendrogram was constructed to reveal the genetic distance among the lines for selected salt tolerance linked SSR loci. In SSR derived dendrogram, the wild rice (O. rufipogon) got totally separated from the all three rice genotypes though all the studied four lines showed equal sensitivity for salt sensitivity in a physiological screening experiment. From the combined experiment, it can be concluded that genetic architecture of floral organ development loci in var. Jugal may have some uniqueness which is not present in normal rice but common to O. rufipogon, a species which is regarded as immediate progenitor of present day modern rice (O. sativa). Though this uniqueness was not confirmed by second set genetic loci associated with salt tolerance in rice, the information resulted from this experiment was preliminary and based only on allelic size (molecular weight of amplicon), which should be confirmed through sequence analysis for further analysis.

Gene Expression Profiling during Wilting in Chickpea Caused by <i>Fusarium oxysporum</i>F. sp. <i>Ciceri</i>

Fusarium oxysporum f. sp. ciceri (Foc), one of the most important fungal pathogen of chickpea, is a constant threat to this crop plant. In the present study gene expression analysis of chickpea roots during Foc infection was performed using various approaches. cDNAs derived from total mRNA during infection process of susceptible (JG62)and resistant (Digvijay) cultivars, were amplified using random oligonucleotides. Sequence characterization of differentially expressed transcripts revealed their homology with many plant genes essential for various metabolic functions including defense. Further, expression patterns of specific candidate gene transcripts were analyzed in the Foc inoculated and uninoculated resistant and susceptible chickpea cultivars, on day 6 of infection. Semiquantitative RT-PCR analysis of defense related genes was performed using gene specific oligonucleotides in resistant and susceptible chickpea cultivars. The expression of fungal pathogenesis related genes and their race specific response was determined throughout the course of chickpea-Foc interaction. Temporal expression and race specific response of plant defense related and fungal virulence genes were studied in the resistant and susceptible cultivars of chickpea inoculated with three races of Foc highlighting the host-pathogen interactions. Few genes, involved in chickpea defense against Fusarium wilt which were not reported previously were unveiled in this study.

2,4-D Hyper Accumulation Induced Cellular Responses of Azolla pinnata R. Br. to Sustain Herbicidal Stress

In the present experiment with ongoing concentration(0µM,100µM,250µM,500µM and 1000µM)of 2,4-D,the responses of Azolla pinnata R.Br.was evaluated based on cellular functions.Initially,plants were significantly tolerated up to 1000µM of 2,4-D with its survival.This was accompanied by a steady decline of indole acetic acid(IAA)concentration in tissues with 78.8%over the control.Membrane bound H^(+)-ATPase activity was over expressed within a range of 1.14 to 1.25 folds with activator(KCl)and decreased within a range of 57.3 to 74.6%in response to inhibitor(Vanadate)application.With regards to IAA metabolism,plants recorded a linear increase with wall bound oxidase activity up to maximum concentration of 2,4-D.The variations were more moderated when wall bound IAA-oxidase recorded a linear increase proportionate to the 2,4-D concentrations.This was more extended with the presence of different isoforms of IAA-oxidase which was much more pronounced with distinct polymorphisms of expressed proteins,however,not independent to the 2,4-D concentrations.Polyamines like spermine,spermidine and putrescine(spm,spd and put)were not consistent in concentration with the dosages of 2,4-D.Besides these,plants were induced to apoplastic NAD(P)H oxidase activity maximally by 1.6 folds under 500µM 2,4-D over control.Still,putrescine responded more or less consistently and recorded maximally 11.9 folds at 500µM 2,4-D as compared to the control.NAD(P)H oxidase activity recorded maximally 1.6 folds against control and remain consistent throughout the concentrations of 2,4-D.GPX along with APX were more linear in responses through the concentration of 2,4-D except CAT as compared to control.On enzymatic antioxidative activity,peroxidases(GPX and APX)were overexpresed in a similar manner except for catalase with a non-significant rise.In stabilization of cellular redox,glutathione reductase attended maximum value by 2.45 folds at 1000µM evidenced with significant variations in protein polymorphism.The sensitivity of 2,4-D also appeared in Azolla with a maximum loss of nucleic acids as documented by the comet assay.Moreover,the Azolla might have some DNA damage protective activity as evident using frond extract with plasmid nick assay.Therefore,Azolla plants with its cellular responses is evident to sustain against the 2,4-D herbicidal stress and may be granted in bio remediation process for the contaminated soil.

Pb-Stress Induced Oxidative Stress Caused Alterations in Antioxidant Efficacy in Two Groundnut (<i>Arachis hypogaea</i>L.) Cultivars

Lead (Pb) is an important environmental pollutant extremely toxic to plants and other living organisms including humans. To assess Pb phytotoxicity, a pot culture experiment was carried out using two groundnut cultivars (Arachis hypogaea L. cultivar K6 and cultivar K9) on plant growth, ROS levels, lipid peroxidation, and antioxidant metabolism using biochemical, histochemical methods. Plants were grown in pots for 14 days, in the botanic garden, and subjected to Pb-stress (0, 100, 200, 400 and 800 ppm) by adding Pb (NO3)2 solution and further allowed to grow for 10 days. The results showed that cultivar K6 registered lower Pb accumulation than cultivar K9, however, localization of Pb was greater in roots than leaves in both groundnut cultivars. The Pb-stress results in an increase in free radicals (O2?- and H2O2) generation in both groundnut cultivars, but more significantly in cultivar K9 than K6. Pb-stress also caused significant changes in the rate of peroxidation as shown in the levels of malondialdehyde (MDA) content in roots and leaves of both groundnut cultivars. Free proline, ascorbic acid (AsA) and non-protein thiol (NP-SH) contents were increased in cultivar K6 due to Pb-stress, but less in cultivar K9. Pb treated plants showed increased levels of antioxidant enzymes such as superoxide dismutase (SOD), guaiacol peroxidase (GPX), ascorbate peroxidase (APX) glutathione reductase (GR) and glutathione S-transferase (GST). Isozyme band intensities of SOD, GPX and APX were more consistent with the respective changes in antioxidative enzyme activities. These results indicate that cultivar K6 possesses greater tolerance potential for Pb toxicity than cultivar K9.

Cadmium Accumulation in <i>Marsilea minuta</i>Linn. and Its Antioxidative Responses

The present study was carried out to investigate the extent of cadmium (Cd) accumulation with its possible impact on physiological and biochemical basis of heavy metal tolerance in Marsilea minuta Linn. Cd salt (0 μM, 50 μM and 100 μM) was allowed to absorb by the plants for prolong days in hydroponic culture and a significant deterioration of the plant biomass was recorded. However, roots absorbed more metals than the leaves. Plants recorded a significant rise of superoxide (O-2) and hydrogen peroxide (H2O2). A noticeable amount of protein oxidation and lipid peroxidation were in proportionate to Cd accumulation. Anthocyanin and flavonoid content were decreased as compared to control condition. Superoxide dismutase (SOD), guaiacol peroxidase (GPX) and glutathione reductase (GR) contributed their antioxidative functions according to the Cd doses. The expression of GR was also evident from its activity staining in gel. So, it may suggest that antioxidative enzymes are up regulated and likely to be responsible for tolerance to Cd induced oxidative stress in Marsilea minuta Linn.

Responses of <i>Marsilea minuta</i>L. to Cadmium Stress and Assessment of Some Oxidative Biomarkers

In a hydroponic based experiment, the Cd toxicity is monitored with some cellular responses of Marsilea plant. Initially, plants were grown under varying concentrations (0, 50, 100 and 200 μM of Cd) of cadmium (Cd) with supplementation of 2 mM spermidine (Spd). The oxidative stress developed by Cd overaccumulation was measured with fall in Relative Growth Rate (RGR) by 27.11% to 59.83% growth reduction over control under varying Cd treatments. The retrieval of RGR was recovered by 1.59 folds as compared to the highest concentration of Cd (200 μM) when plants were fed with Spd. A concomitant degradation of chlorophyll was recorded in dose-dependant manner, however, the retrieval was not much pronounced with Spd. On the contrary, the non-oxidant thiol had borne more clarity with ongoing Cd concentration and appeared to be 40.51% increase maximally for GSH: GSSG at the highest concentration of Cd. Spd has minimized the ratio by 27.4%. The recovery of osmotic turgidity was indexed with a sharp rise in glycine betaine by 3.86 folds maximum at the highest concentration of Cd over control which declined by 30.9% with Spd. Another cellular response of treated plants was more evident from their isozymic profiles with regard to superoxide dismutase (SOD), catalase (CAT), guaiacol peroxidase (GPX). The intensity of protein expression was significantly variable but not in band numbers as evident from Cd treated plants. In vitro enzyme assay of catalase showed as declining trend within the limit of 33.13% to 43.22% which was reported by 1.45 folds when Spd was applied. Therefore, from the present study, the cellular responses of Marsilea plant which showed compatibility for their expression with Cd toxicity could be hypothesized as a case of bioindication.

The expression of proteins involved in digestion and detoxification are regulated in Helicoverpa armigera to cope up with chlorpyrifos insecticide

Helicoverpa armigera is a key pest in many vital crops, which is mainly controlled by chemical strategies, To manage this pest is becoming challenging due to its ability and evolution of resistance against insecticides. Further, its subsequent spread on nonhost plant is remarkable in recent times. Hence, decoding resistance mechanism against phytochemicals and synthetic insecticides is a major challenge. The present work describes that the digestion, defense and immunity related enzymes are associated with chlorpyrifos resistance in 11. armigera. Proteomic analysis of/-/, armigera gut tissue upon feeding on chlorpyrifos containing diet （CH） and artificial diet （AD） using nano-liquid chromatography-mass spectrometry identified upregulated 23-proteins in CH fed larvae. Database searches combined with gene ontology analysis revealed that the identified gut proteins engrossed in digestion, proteins crucial for immunity, adaptive responses to stress, and detoxification. Biochemical and quantitative real-time polymerase chain reaction anal- ysis of candidate proteins indicated that insects were struggling to get nutrients and energy in presence of CH, while at the same time endeavoring to metabolize chlorpyrifos. More- over, we proposed a potential processing pathway of chlorpyrifos in H. armigera gut by examining the metabolites using gas chromatography-mass spectrometry. 1f. armigera exhibit a range of intriguing behavioral, morphological adaptations and resistance to in- secticides by regulating expression of proteins involved in digestion and detoxification mechanisms to cope up with chlorpyrifos. In these contexts, as gut is a rich repository of biological information; profound analysis of gut tissues can give clues of detoxification and resistance mechanism in insects.

Momordica charantia trypsin inhibitor Ⅱ inhibits growth and development of Helicoverpa armigera

Bitter gourd （Momordica charantia L.） seeds contain several squash-type serine proteinase inhibitors （PIs）, which inhibit the digestive proteinases of the polyphagous insect pest Helicoverpa armigera. In the present work isolation of a DNA sequence encoding the mature peptide of a trypsin inhibitor McTI-Ⅱ, its cloning and expression as a recombinant protein using Pichia pastoris have been reported. Recombinant McTI-Ⅱ inhibited bovine trypsin at 1 ： 1 molar ratio, as expected, but did not inhibit chymotrypsin or elastase. McTI-Ⅱ also strongly inhibited trypsin-like proteinases （81% inhibition） as well as the total proteolytic activity of digestive proteinases （70% inhibition） from the midgut of H. armigera larvae. The insect larvae fed with McTI-Ⅱ-incorporated artificial diet suffered over 70% reduction in the average larval weight after 12 days of feeding. Moreover, ingestion of McTI-Ⅱ resulted in 23% mortality in the larval population. The strong antimetabolic activity of McTI-Ⅱ toward H. armigera indicates its probable use in developing insect tolerance in susceptible plants.