The Introduction of a Gene Encoding a Novel Peptide Into Plants to Increase Plant Bacterial Resistance.

A 38 amino acid long peptide, Shiva-1, was designed and chemically synthesized in our laboratory. Shiva-1 has only 46% amino acid homology with the naturally occurring lytic peptide from Hyalophora cecropia, known as Cecropin B. However, hydrophobic properties and charge density of the natural molecule were conserved at 100% in the synthetic peptide. This novel peptide was shown to be capable of killing a number of different species of plant pathogenic bacteria at nanomolar concentrations. Comparative studies show that Shiva-1 is more effective in this bacteriolytic activity than Cecropin B. The gene for Shiva-1 was chemically synthesized and cloned into the binary vector Agrobacterium tumefaciens LBA4404/pBI121 under the control of a wound-inducible plant promoter. Tobacco leaf discs were transformed using this binary system containing the gene encoding Shiva-1 and transgenic plants were obtained. These plants were shown to be kanamycin-resistant and displayed $\beta$-glucoronidase activity indicating effective plant transformation. Southern blots confirmed the presence of a single copy Shiva-1 gene integrated into the genomes of individual transgenic plants. Southern analyses also showed stable inheritance of the Shiva-1 gene. Northern analyses verified that the expression of the gene could be triggered by mechanical and pathogen-induced wounding. Northern blots also showed that a mRNA with the expected size of Shiva-1 transcript hybridized to the Shiva-1 probe. Western blots indicated the presence of a distinct band hybridizing to antisera raised against Shiva-1. R1 plants from self-crossed mother transgenic plants demonstrated a 3:1 segregation pattern for kanamycin resistance gene. Plant bacterial challenge suggested that the R1 transgenic plants exhibit delayed symptoms, reduced disease severity and mortality after infection with Pseudomonas solenacearum, when compared to control plants

Genes, Proteins and Metabolites in the Interaction of Strawberry and Fungal Pathogen

Colletrotrichum acutatum and Botrytis cinerea are among the major fungal pathogens of Fragaria spp. Both pathogens could infect strawberry fruits during the fruit’s early developing stage and remain quiescent until ripening. In strawberry, a fruit ontogenic resistance to pathogen infection was described and correlated with fungal quiescence during the unripe stages of the fruit. Due to the period of fungal quiescence, the management of anthracnose and gray mould diseases becomes more complex as symptoms only manifest in ripe fruits. To identify the underlying component in the ontogenic resistance of strawberry fruits, transcriptomic and metabolomic approaches were used. White and red fruits of strawberry were artificially inoculated with C. acutatum and B. cinerea. Transcriptome profile of B. cinerea infected fruits exhibited a general up-regulation of defense-related genes in white fruits after 24 h of infection. Meanwhile, accumulation of phenolic compounds such as proanthocyanidins, catechins and the ellagitannin casuarictin was also observed in white fruits after 48 h of interacting with C. acutatum and B. cinerea. The acquisition of these findings could provide a benchmark to further investigate the interaction of strawberry against pathogens with latent infection. Hence, a strawberry transformation was performed to study the mechanism of a gene encoding for a mannose-binding lectin protein which was previously identified to be correlated with the resistance of white strawberry fruits to C. acutatum. The regeneration system utilized in the transformation is also discussed

Transcriptome Profiles of Strawberry (Fragaria vesca) Fruit Interacting With Botrytis cinerea at Different Ripening Stages

Gray mold caused by Botrytis cinerea is a major cause of economic losses in strawberry fruit production, limiting fruit shelf life and commercialization. When the fungus infects Fragaria x ananassa strawberry at flowering or unripe fruit stages, symptoms develop after an extended latent phase on ripe fruits before or after harvesting. To elucidate the growth kinetics of B. cinerea on flower/fruit and the molecular responses associated with low susceptibility of unripe fruit stages, woodland strawberry Fragaria vesca flowers and fruits, at unripe white and ripe red stages, were inoculated with B. cinerea. Quantification of fungal genomic DNA within 72 h postinoculation (hpi) showed limited fungal growth on open flower and white fruit, while on red fruit, the growth was exponential starting from 24 hpi and sporulation was observed within 48 hpi. RNA sequencing applied to white and red fruit at 24 hpi showed that a total of 2,141 genes (12.5% of the total expressed genes) were differentially expressed due to B. cinerea infection. A broad transcriptional reprogramming was observed in both unripe and ripe fruits, involving in particular receptor and signaling, secondary metabolites, and defense response pathways. Membrane-localized receptor-like kinases and nucleotide-binding site leucine-rich repeat genes were predominant in the surveillance system of the fruits, most of them being downregulated in white fruits and upregulated in red fruits. In general, unripe fruits exhibited a stronger defense response than red fruits. Genes encoding for pathogenesis-related proteins and flavonoid polyphenols as well as genes involved in cell-wall strengthening were upregulated, while cell-softening genes appeared to be switched off. As a result, B. cinerea remained quiescent in white fruits, while it was able to colonize ripe red fruits

Molecular analysis of the early interaction between the grapevine flower and Botrytis cinerea reveals that prompt activation of specific host pathways leads to fungus quiescence

Grape quality and yield can be impaired by bunch rot, caused by the necrotrophic fungus Botrytis cinerea. Infection often occurs at flowering, and the pathogen stays quiescent until fruit maturity. Here, we report a molecular analysis of the early interaction between B. cinerea and Vitis vinifera flowers, using a controlled infection system, confocal microscopy and integrated transcriptomic and metabolic analysis of the host and the pathogen. Flowers from fruiting cuttings of the cultivar Pinot Noir were infected with green fluorescent protein (GFP)-labelled B. cinerea and studied at 24 and 96 hours post-inoculation (h.p.i.). We observed that penetration of the epidermis by B. cinerea coincided with increased expression of genes encoding cell-wall-degrading enzymes, phytotoxins and proteases. Grapevine responded with a rapid defence reaction involving 1193 genes associated with the accumulation of antimicrobial proteins, polyphenols, reactive oxygen species and cell wall reinforcement. At 96 h.p.i., the reaction appears largely diminished both in the host and in the pathogen. Our data indicate that the defence responses of the grapevine flower collectively are able to restrict invasive fungal growth into the underlying tissues, thereby forcing the fungus to enter quiescence until the conditions become more favourable to resume pathogenic development

Transcriptome profiles of strawberry (Fragaria vesca) fruit interacting with Botrytis cinerea at different ripening stages

Gray mold caused by Botrytis cinerea is a major cause of economic losses in strawberry fruit production, limiting fruit shelf life and commercialization. When the fungus infects Fragaria × ananassa strawberry at flowering or unripe fruit stages, symptoms develop after an extended latent phase on ripe fruits before or after harvesting. To elucidate the growth kinetics of B. cinerea on flower/fruit and the molecular responses associated with low susceptibility of unripe fruit stages, woodland strawberry Fragaria vesca flowers and fruits, at unripe white and ripe red stages, were inoculated with B. cinerea. Quantification of fungal genomic DNA within 72 h postinoculation (hpi) showed limited fungal growth on open flower and white fruit, while on red fruit, the growth was exponential starting from 24 hpi and sporulation was observed within 48 hpi. RNA sequencing applied to white and red fruit at 24 hpi showed that a total of 2,141 genes (12.5% of the total expressed genes) were differentially expressed due to B. cinerea infection. A broad transcriptional reprogramming was observed in both unripe and ripe fruits, involving in particular receptor and signaling, secondary metabolites, and defense response pathways. Membrane-localized receptor-like kinases and nucleotide-binding site leucine-rich repeat genes were predominant in the surveillance system of the fruits, most of them being downregulated in white fruits and upregulated in red fruits. In general, unripe fruits exhibited a stronger defense response than red fruits. Genes encoding for pathogenesis-related proteins and flavonoid polyphenols as well as genes involved in cell-wall strengthening were upregulated, while cell-softening genes appeared to be switched off. As a result, B. cinerea remained quiescent in white fruits, while it was able to colonize ripe red fruit

Antioxidant and oxidative stress: a mutual interplay in age-related diseases

Aging is the progressive loss of organ and tissue function over time. Growing older is positively linked to cognitive and biological degeneration such as physical frailty, psychological impairment, and cognitive decline. Oxidative stress is considered as an imbalance between pro- and antioxidant species, which results in molecular and cellular damage. Oxidative stress plays a crucial role in the development of age-related diseases. Emerging research evidence has suggested that antioxidant can control the autoxidation by interrupting the propagation of free radicals or by inhibiting the formation of free radicals and subsequently reduce oxidative stress, improve immune function, and increase healthy longevity. Indeed, oxidation damage is highly dependent on the inherited or acquired defects in enzymes involved in the redox-mediated signaling pathways. Therefore, the role of molecules with antioxidant activity that promote healthy aging and counteract oxidative stress is worth to discuss further. Of particular interest in this article, we highlighted the molecular mechanisms of antioxidants involved in the prevention of age-related diseases. Taken together, a better understanding of the role of antioxidants involved in redox modulation of inflammation would provide a useful approach for potential interventions, and subsequently promoting healthy longevity

Development of ambient noise reduction android application for voice calls using equalization of audio frequency

Noise reduction now plays a major role in any form of communication since noise reduce the quality of information in data. Noise reduction technologies give the users ease in understanding the information and comfort since noise reduction also minimizes loud unwanted sound. The researchers maximized this opportunity to address the problems regarding mobile phone communication particularly on phone calls and contributes to the already existing body of knowledge that addresses such concerns.This thesis paper is about how the researchers designed an android application to capture live phone calls and minimize the noise frequencies. The researchers used equalization to reduce the gain of noise frequencies and amplify the voice frequency range so that the voice frequency will over power the noise frequency. With the use of MatLab and Audacity, the researchers was able to analyze the data.Through the researchers\u27 experiment and analysis it can be concluded that the scheme is indeed possible although it still needs major improvements over the speed and ram size of the phone\u27s hardware, the system is till functional and is capable of reducing noise in active phone calls

EXPRESSION OF A CECROPIN-B LYTIC PEPTIDE ANALOG IN TRANSGENIC TOBACCO CONFERS ENHANCED RESISTANCE TO BACTERIAL WILT CAUSED BY PSEUDOMONAS-SOLANACEARUM

WOS: A1993KT92300006Cecropin B is a naturally-occurring lytic peptide found in Hyalophora cecropia, the Giant Silk Moth. It is thought to comprise part of an inducible humoral defense system that combats infection in the insect. Two 38 amino acid peptides, SB-37 and Shiva-1, were produced as substitution analogs of Cecropin B. SB-37 is 95% homologous to Cecropin B while Shiva-1 retains only 46% homology to the natural molecule. However, hydrophobic properties and charge density of the native structure were conserved at 100% in the synthetic peptides. The genes for both peptides were chemically synthesized and cloned into the binary vector pBI121 under the control of a constitutive or wound-inducible plant promoter. Transgenic tobacco plants (RO) were subsequently obtained via Agrobacterium transformation. Bioassays to test disease resistance of R1 progeny indicate that, compared to transgenic control and SB-37 plants, Shiva-1 seedlings exhibited delayed wilt symptoms and reduced disease severity and mortality after infection with a highly virulent strain of Pseudomonas solanacearum

Expression of a Cecropin B lytic peptide analog in transgenic tobacco confers enhanced resistance to bacterial wilt caused by Pseudomonas solanacearum

Cecropin B is a naturally-occurring lytic peptide found in Hyalophora cecropia, the Giant Silk Moth. It is thought to comprise part of an inducible humoral defense system that combats infection in the insect. Two 38 amino acid peptides, SB-37 and Shiva-1, were produced as substitution analogs of Cecropin B. SB-37 is 95% homologous to Cecropin B while Shiva-1 retains only 46% homology to the natural molecule. However, hydrophobic properties and charge density of the native structure were conserved at 100% in the synthetic peptides. The genes for both peptides were chemically synthesized and cloned into the binary vector pBI121 under the control of a constitutive or wound-inducible plant promoter. Transgenic tobacco plants (RO) were subsequently obtained via Agrobacterium transformation. Bioassays to test disease resistance of R1 progeny indicate that, compared to transgenic control and SB-37 plants, Shiva-1 seedlings exhibited delayed wilt symptoms and reduced disease severity and mortality after infection with a highly virulent strain of Pseudomonas solanacearum. © 1993.Agricultural Experiment Station, New Mexico State UniversityThe authors wish to acknowledge the expert technical assistance in peptide synthesis and purification provided by Martha Juban. This research was supported, in part, by funds from the Louisiana State University Agricultural Experiment Station