Showing papers by "Imran Amin published in 2020"

The Cotton Wall-Associated Kinase GhWAK7A Mediates Responses to Fungal Wilt Pathogens by Complexing with the Chitin Sensory Receptors.

Abstract: Plant receptor-like kinases (RLKs) are important players in response to pathogen infections. Verticillium and Fusarium wilts, caused by Verticillium dahliae (Vd) and Fusarium oxysporum f. sp vasinfectum (Fov), respectively, are among the most devastating diseases in cotton (Gossypium spp). To understand the cotton response to these soil-borne fungal pathogens, we performed a genome-wide in silico characterization and functional screen of diverse RLKs for their involvement in cotton wilt diseases. We identified Gossypium hirsutum GhWAK7A, a wall-associated kinase, that positively regulates cotton response to both Vd and Fov infections. Chitin, the major constituent of the fungal cell wall, is perceived by lysin-motif-containing RLKs (LYKs/CERK1), leading to the activation of plant defense against fungal pathogens. A conserved chitin sensing and signaling system is present in cotton, including chitin-induced GhLYK5-GhCERK1 dimerization and phosphorylation, and contributes to cotton defense against Vd and Fov Importantly, GhWAK7A directly interacts with both GhLYK5 and GhCERK1 and promotes chitin-induced GhLYK5-GhCERK1 dimerization. GhWAK7A phosphorylates GhLYK5, which itself does not have kinase activity, but requires phosphorylation for its function. Consequently, GhWAK7A plays a crucial role in chitin-induced responses. Thus, our data reveal GhWAK7A as an important component in cotton response to fungal wilt pathogens by complexing with the chitin receptors.

Precise CRISPR-Cas9 Mediated Genome Editing in Super Basmati Rice for Resistance Against Bacterial Blight by Targeting the Major Susceptibility Gene.

Abstract: Basmati rice is famous around the world for its flavor, aroma, and long grain. Its demand is increasing worldwide, especially in Asia. However, its production is threatened by various problems faced in the fields, resulting in major crop losses. One of the major problems is bacterial blight caused by Xanthomonas oryzae pv. oryzae (Xoo). Xoo hijacks the host machinery by activating the susceptibility genes (OsSWEET family genes), using its endogenous transcription activator like effectors (TALEs). TALEs have effector binding elements (EBEs) in the promoter region of the OsSWEET genes. Out of six well-known TALEs found to have EBEs in Clade III SWEET genes, four are present in OsSWEET14 gene's promoter region. Thus, targeting the promoter of OsSWEET14 is very important for creating broad-spectrum resistance. To engineer resistance against bacterial blight, we established CRISPR-Cas9 mediated genome editing in Super Basmati rice by targeting 4 EBEs present in the promoter of OsSWEET14. We were able to obtain four different Super Basmati lines (SB-E1, SB-E2, SB-E3, and SB-E4) having edited EBEs of three TALEs (AvrXa7, PthXo3, and TalF). The edited lines were then evaluated in triplicate for resistance against bacterial blight by choosing one of the locally isolated virulent Xoo strains with AvrXa7 and infecting Super Basmati. The lines with deletions in EBE of AvrXa7 showed resistance against the Xoo strain. Thus, it was confirmed that edited EBEs provide resistance against their respective TALEs present in Xoo strains. In this study up to 9% editing efficiency was obtained. Our findings showed that CRISPR-Cas9 can be harnessed to generate resistance against bacterial blight in indigenous varieties, against locally prevalent Xoo strains.

Molecular insight into cotton leaf curl geminivirus disease resistance in cultivated cotton (Gossypium hirsutum).

Abstract: Cultivated cotton (Gossypium hirsutum) is the most important fibre crop in the world. Cotton leaf curl disease (CLCuD) is the major limiting factor and a threat to textile industry in India and Pakistan. All the local cotton cultivars exhibit moderate to no resistance against CLCuD. In this study, we evaluated an exotic cotton accession Mac7 as a resistance source to CLCuD by challenging it with viruliferous whiteflies and performing qPCR to evaluate the presence/absence and relative titre of CLCuD-associated geminiviruses/betasatellites. The results indicated that replication of pathogenicity determinant betasatellite is significantly attenuated in Mac7 and probably responsible for resistance phenotype. Afterwards, to decipher the genetic basis of CLCuD resistance in Mac7, we performed RNA sequencing on CLCuD-infested Mac7 and validated RNA-Seq data with qPCR on 24 independent genes. We performed co-expression network and pathway analysis for regulation of geminivirus/betasatellite-interacting genes. We identified nine novel modules with 52 hubs of highly connected genes in network topology within the co-expression network. Analysis of these hubs indicated the differential regulation of auxin stimulus and cellular localization pathways in response to CLCuD. We also analysed the differential regulation of geminivirus/betasatellite-interacting genes in Mac7. We further performed the functional validation of selected candidate genes via virus-induced gene silencing (VIGS). Finally, we evaluated the genomic context of resistance responsive genes and found that these genes are not specific to A or D sub-genomes of G. hirsutum. These results have important implications in understanding CLCuD resistance mechanism and developing a durable resistance in cultivated cotton.

Genome Editing Technologies for Rice Improvement: Progress, Prospects, and Safety Concerns.

Abstract: Rice (Oryza sativa) is an important staple food crop worldwide; to meet the growing nutritional requirements of the increasing population in the face of climate change, qualitative and quantitative traits of rice need to be improved. Stress-tolerant crop varieties must be developed with higher yields and increased adaptability to the changing climate. Genome editing and speed breeding have improved the accuracy and pace of rice breeding. New breeding technologies including genome editing have been established in rice, expanding the potential for crop improvement. Recently, other genome editing techniques such as CRISPR-directed evolution, CRISPR-Cpf1, and base editors have also been used for efficient genome editing in rice. Since rice is an excellent model system for functional studies due to its small genome and close syntenic relationships with other cereal crops, new genome-editing technologies continue to be developed for use in rice. In this review, we focus on genome-editing tools for rice improvement to address current challenges and provide examples of genome editing in rice. We also shed light on expanding the scope of genome editing and systems for delivering homology-directed repair templates. Finally, we discuss safety concerns and methods for obtaining transgene-free crops.

Association of cotton leaf curl Multan betasatellite and Ageratum conyzoides symptomless alphasatellite with tomato leaf curl New Delhi virus in Luffa cylindrica in Pakistan

Abstract: Luffa plants showing symptoms of begomovirus infection were collected from Vehari, Punjab in 2016. RCA and PCR amplifications of begomovirus genomic components showed the presence of a typical Old World bipartite begomovirus Tomato leaf curl New Delhi virus (ToLCNDV). Interestingly, Ageratum conyzoides symptomless alphasatellite and CLCuMB were also found associated with bipartite begomoviruses in the same host, however, no monopartite begomovirus was detected. To the best of our knowledge, this is the first report of ToLCNDV with associated satellites infecting L. cylindrica in Pakistan. The importance of begomovirus extended host range with different combinations of associated satellites has been discussed.

Improving COVID-19 Testing Efficiency using Guided Agglomerative Sampling

Abstract: One of the challenges in the current COVID-19 crisis is the time and cost of performing tests especially for large-scale population surveillance. Since, the probability of testing positive in large population studies is expected to be small (

Effects of the transient expression of heterologous RNA virus-encoded silencing suppressors on the infectivity and systemic movement of tomato leaf curl New Delhi virus

Abstract: RNA silencing or RNA interference (RNAi) is an endogenous mechanism of gene regulation and adaptive defence against invading microbes of higher organisms, triggered by double-stranded RNA. Viruses circumvent this host defence by encoding specific proteins, RNA silencing suppressors (RSSs). Evidence has been mounting that, in common with RNA viruses, the movement of geminiviruses in plants is countered by an RNAi-based resistance. To investigate this idea, the impact of the transient expression of suppressor proteins encoded by three RNA viruses on the infectivity and movement of tomato leaf curl New Delhi virus (ToLCNDV; a bipartite begomovirus family Geminiviridae) was investigated. Both genomic components (designated as DNA A [TA] and DNA B [TB]) are essential for symptomatic infection of plants, although TA alone may inefficiently spread to plants without inducing symptoms. Each of the three heterologous RSSs was shown to enhance the pathogenicity of ToLCNDV when expressed transiently at the site of inoculation. The effects ranged from an increase in the numbers of plants ultimately containing a virus or DNA A component (for inoculations with only TA or only with TA harbouring a mutation of the coat protein gene [TAΔCP]), through to the induction of symptoms (for TA and TAΔCP inoculations of tomato), to a reduction in latent period (for all inoculations involving TB). These results show that heterologous RSSs may enhance ToLCNDV pathogenicity and are consistent with the idea that geminivirus movement in plants is constrained by an RNAi-based resistance.

Cotton leaf curl Kokhran virus in association with Chili leaf curl betasatellite infecting mungbean (Vigna radiata.) and black gram (Vigna mungo.) in Pakistan

Abstract: In 2016, begomovirus-like symptoms were observed in mungbean (Vigna radiata) and black gram (Vigna mungo) in Pakistan. Symptomatic and asymptomatic leaf samples were collected and total genomic DNA was extracted. Full length begomovirus and betasatellite genomes were PCR amplified and cloned in plasmid pTZ/57RT cloning vector. The cloned molecules were subjected to Sanger sequencing. Based on Sequence Demarcation Tool, the begomovirus clone SR-3 identified in mungbean showed 94.50% highest identity with cotton leaf curl Kokhran virus-Kokhran (CLCuKoV-Ko), while the clone SR-5 isolated from black gram showed 99.7% highest identity with CLCuKoV-Ko. In phylogenetic tree analysis the identified isolates made a lower branch tree with CLCuKoV-Ko. The betasatellite clone SR-79 isolated from black gram showed 97% similarity with chili leaf curl betasatellite (ChLCB) and the betasatellite clone SR-93 isolated from mungbean showed 97.7% similarity with ChLCB. Southern blot analysis also confirmed the presence of CLCuKoV-Ko. This is the first report of CLCuKoV-Ko associated with ChLCB infecting both mungbean and black gram in Pakistan. This presents the new combination of disease complex and increase in host range of this virus species, which shows a real threat to other crops especially legume crops in Pakistan.

Exogenous Application of Elicitors Induces Biochemical Alterations and Develops Tolerance against Yellow Mosaic Disease in Mungbean

Abstract: Mungbean ( Vigna radiata ) is seriously affected by mungbean yellow mosaic disease (MYMD) caused by Mungbean yellow mosaic India virus (MYMIV) in the Indian subcontinent. The present study was conducted to evaluate the effectiveness of salicylic acid (SA) and benzothiadiazole (BTH) for the management of MYMD and to find their role in inducing alteration in different biochemical parameters in susceptible mungbean genotype VC3061A. Exogenous application of the elicitors resulted in delayed symptom development and reduced disease severity (DS). The severity index (SI) was higher on positive control plants while the minimum was observed in post combined application of BTH+SA, followed by BTH and SA application. Total phenolic contents (TPC) and malondialdehyde (MDA) increased significantly in virus inoculated plants of all treatments as compared to their healthy controls. SOD activity was increased significantly in BTH+SA treated plants but decreased in disease control and BTH treated virus inoculated plants. A significant decrease in catalase (CAT) activity, while an increase in peroxidase (POD) was observed in BTH+SA, treated virus inoculated plants. Protease and esterase activity were significantly increased in SA treated virus inoculated plants. Plant pigments exhibited decreased concentration in virus inoculated plants compared to non-inoculated plants under all the treatments except SA treated plants. Enhanced or suppressed levels of antioxidants suggest an association between constitutive induced levels of these enzymes. In this study, we also report for the first time the protein profiling of mungbean genotype VC3061A after the exogenous application of different combinations of elicitors. Proteomic analyses revealed the expression of two proteins phosphatase 2C 16-like isoform and capsid protein after MYMIV inoculation in SA and BTH+SA treated plants which may trigger signal transduction pathway and consequently induces resistance against MYMIV in V. radiata by activating PR protein.