Naila Shahid

Identification and application of biocontrol agents against Cotton leaf curl virus disease in Gossypium hirsutum under greenhouse conditions

ABSTRACT Biological control is a novel approach in crop protection. Bacteria, such as Bacillus spp. and Pseudomonas spp., are reported for this purpose and some of their products are already commercially available. In this study, the rhizosphere and phyllosphere of healthy cotton plants were used as a source of bacterial isolates with properties of potential biocontrol agents. The isolates were screened for phosphate solubilization activity, indole acetic acid (IAA) production and antifungal activity. Two isolates, S1HL3 and S1HL4, showed phosphate solubilization and IAA production simultaneously, while another two, JS2HR4 and JS3HR2, demonstrated potential to inhibit fungal pathogens. These bacteria were identified as Pseudomonas aeruginosa (S1HL3), Burkholderia sp. (S1HL4) and Bacillus sp. (JS2HR4 and JS3HR2) based on biochemical and molecular characteristics. The isolates were tested against Cotton leaf curl virus (CLCuV) in greenhouse conditions, both as individual bacterial isolates and consortia. Treated plants were healthy as compared to control plants, where up to 74% of the plants were symptomatic for CLCuV infection. Maximum inhibition of CLCuV was observed in the plants treated with a mixture of bacterial isolates: the viral load in the treated plants was only 0.4% vs. up to 74% in controls. This treatment consortium included P. aeruginosa S1HL3, Burkholderia sp. S1HL4 and Bacillus spp. isolates, JS2HR4 and JS3HR2. The principal-component biplot showed a highly significant correlation between the viral load percentage and the disease incidence.

Improved antifungal activity of barley derived chitinase I gene that overexpress a 32 kDa recombinant chitinase in Escherichia coli host

Agricultural crops suffer many diseases, including fungal and bacterial infections, causing significant yield losses. The identification and characterisation of pathogenesis-related protein genes, such as chitinases, can lead to reduction in pathogen growth, thereby increasing tolerance against fungal pathogens. In the present study, the chitinase I gene was isolated from the genomic DNA of Barley (Hordeum vulgare L.) cultivar, Haider-93. The isolated DNA was used as template for the amplification of the ∼935 bp full-length chitinase I gene. Based on the sequence of the amplified gene fragment, class I barley chitinase shares 93% amino acid sequence homology with class II wheat chitinase. Interestingly, barley class I chitinase and class II chitinase do not share sequence homology. Furthermore, the amplified fragment was expressed in Escherichia coli Rosetta strain under the control of T7 promoter in pET 30a vector. Recombinant chitinase protein of 35 kDa exhibited highest expression at 0.5 mM concentration of IPTG. Expressed recombinant protein of 35 kDa was purified to homogeneity with affinity chromatography. Following purification, a Western blot assay for recombinant chitinase protein measuring 35 kDa was developed with His-tag specific antibodies. The purified recombinant chitinase protein was demonstrated to inhibit significantly the important phytopathogenic fungi Alternaria solani, Fusarium spp, Rhizoctonia solani and Verticillium dahliae compared to the control at concentrations of 80 μg and 200 μg.

An overview of phytochrome: An important light switch and photo-sensory antenna for regulation of vital functioning of plants

Abstract Plants are the primary source of nutrition and essential to maintain life on earth. They have evolved very delicate and advanced photo-sensory antennae to sense their outer environment and transduce the received information for their growth and development accordingly. This “light switch” phenomenon of plants has slowly being unraveled and various plant photoreceptors, their role in downstream molecular signaling, mutual interaction, response to circadian cycle and light signals have been discovered. The photosensory antennae in plants; phytochromes, cryptochromes and phototropins play a very crucial role in sensing the ambient light intensities. By direct interaction with the environment through these photosensory antennae, plants shift their homeostasis to regulate their growth and development. The phytochrome light receptors of plants are responsive to R/FR light and by inducing signaling pathways, trigger the physiological responses such as germination and flowering. The phytochromes also directly contribute to plant development by affecting its photosynthetic rate. To elucidate the role of phytochromes in plant metabolism, this review will focus on the importance of phytochromes, their mechanism of action and their application as an emerging field in plant biology.

Multidimensional roles of flavonoids in background of Gossypium hirsutum

Abstract Anthocyanins are known as the central players in kingdom Planta, elucidating their major role in various abiotic and biotic stresses including physiological events such as plant growth, development and defense responses. Plants synthesize a wide range of chemically diverse secondary metabolites that are distinctively important industrial biochemical compounds. The elucidation of regulatory mechanisms by flavonoids could facilitate the generation of new transgenic cotton varieties along the rational to increase fiber characteristics and yield at the verge of auxin transport. The anthocyanin gene expression and its structural genes encoding enzymes have already been explored in many ornamental flowers. But its particular role in crop plant stress regulatory mechanism is yet to be explored. This brief review summarizes the role of flavonoids against various abiotic factors including drought, light/ultraviolet (UV) stress, water scarcity, temperature variations and so on in the background of cotton crop. These factors adversely affect the growth and productivity of the crops. Moreover, information of the flavonoids metabolic interactions in relation to cotton fiber characteristics still needs to be unrevealed.

A concise review of poultry vaccination and future implementation of plant-based vaccines

Every year the growth of the poultry industry is severely threatened by a number of infectious viral, bacterial and parasitic diseases. There are a number of vaccines to control these diseases including inactivated virus vaccines, attenuated virus vaccines, live virus vaccines, and subunit vaccines, but they are often relatively expensive and require cold storage and trained people to administer them, especially in developing countries. Plant-based vaccines provide a better option to control these diseases in low profit margin poultry industry. Still there are some challenges in the field of plant-based, so called ‘green’ vaccines. Injection-based oral priming is a big challenge for commercialisation of green vaccines so, new techniques are needed in the field of plant-based vaccine to pass these barriers for commercialisation. This discusses the potential for plant-based vaccines and whether they are good option to control poultry diseases.

Escherichia coli expression of NDV fusion protein gene and determination of its antigenic epitopes

Abstract Recurrent outbreaks of Newcastle disease have questioned the usage of existing vaccines that whether they are still adequate to protect clinical diseases and inhibit virus transmission in poultry. Advancement in molecular biology has led to the production of recombinant vaccines in recent years, which can be a more useful strategy to control infections of Newcastle disease virus (NDV). Studies indicate that the pathogenic nature of NDV is mediated by its membrane associated fusion (F) protein. Here we report the cloning of the full-length F gene-pET30a and its expression in Escherichia coli BL21 DE3 cells through isopropyl β-D-1-thiogalactopyranoside induction. Transferring the protein on nitrocellulose membrane in Western blotting confirmed its specificity with histidine-tagged antibody reaction at the proper size of 67 kDa. Protein purification with nickel charged sepharose column affinity chromatography resulted in a single band of 67 kDa purified His-tag F protein on SDS-PAGE. Analysis of its immunogenicity through bioinformatics tools revealed that more than 70% of its sequence is antigenically active comprising 24 linear immunogenic peptides predicted by the Linear epitope prediction tool and 9 immunogenic peptides predicted by ElliPro. This is a key achievement of the study, which may lead towards recombinant vaccine production in future. In conclusion, our findings suggest that rather than employing live viral vaccines, using a purified immunogenic recombinant F protein as a vaccine or cloning the same gene in a suitable plant vector for production of edible vaccine will provide better protection against the NDV into chicken.