Browsing by Author "Swain, Durga Madhab"

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Bacteria-fungal confrontation and fungal growth prevention assay
(Bio-protocol LLC., 2018) Kumar, Rahul; Swain, Durga Madhab; Yadav, Sunil Kumar; Tyagi, Isha; Kumar, Rajeev; Das, Joyati; Ghosh, Srayan; Jha, Gopaljee
There are some bacteria which can grow and multiply at the cost of living fungal biomass. They can potentially utilize fungi as a source of nutrients to forage over them. Such phenomenon is known as bacterial mycophagy, however, its mechanistic insights need to be explored to identify the molecules involved in mycophagy for potential utilization in controlling various fungal diseases. Recently we have demonstrated that a rice-associated bacteria Burkholderia gladioli strain NGJ1 exhibits mycophagous ability on several fungi, including Rhizoctonia solani, the necrotrophic fungal pathogen causing sheath blight disease in rice. We hereby describe our validated and efficient methods used to study B. gladioli strain NGJ1-R. solani interactions. These methodologies would be useful for designing assays to study the confrontation between bacteria and fungi which in turn enable discovery of novel antifungal molecules from such bacteria.
Burkholderia gladioli strain NGJ1 deploys a prophage tail-like protein for mycophagy
(Shared Science Publishers OG, 2018) Kumar, Rahul; Yadav, Sunil Kumar; Swain, Durga Madhab; Jha, Gopaljee
Fungal pathogens are responsible for approximately two third of the infectious plant diseases. Historically they have been associated with several devastating famines, causing death and disabilities in humans. Mostly fungal diseases are being controlled by using fungicides which otherwise have adverse side effects on the health of consumers as well as environment. Due to extensive usages, pathogens have evolved resistance against most of the commonly used fungicides and rendered them ineffective. Controlling fungal disease in a sustainable and eco-friendly fashion remains a challenge. The antifungal biocontrol agents are being considered as potent, alternative and ecofriendly approach to manage fungal diseases. In our recent work, we have identified a rice associated bacterium; Burkholderia gladioli strain NGJ1 which demonstrates broad spectrum fungal eating (mycophagous) property. We determined that the bacterium utilizes its type III secretion system (Injectisome) machinery to deploy a prophage tail-like protein (Bg_9562) into fungal cells to devour them. The purified Bg_9562 protein from overexpressing recombinant E. coli strain demonstrates broad spectrum antifungal activity. Overall our study opens up a new opportunity to exploit prophage taillike protein as potent antifungal compound to control plant as well as animal fungal diseases.
Concurrent overexpression of rice G-protein β and γ subunits provide enhanced tolerance to sheath blight disease and abiotic stress in rice
(Springer Nature Publishing AG, 2019) Swain, Durga Madhab; Sahoo, Ranjan Kumar; Chandan, Ravindra Kumar; Ghosh, Srayan; Kumar, Rahul; Jha, Gopaljee; Tuteja, Narendra
The heterotrimeric G-proteins act as signalling molecules and modulate various cellular responses including stress tolerance in eukaryotes. The gamma (γ) subunit of rice G-protein (RGG1) was earlier reported to promote salinity stress tolerance in rice. In the present study, we report that a rice gene-encoding beta (β) subunit of G-protein (RGB1) gets upregulated during both biotic (upon a necrotrophic fungal pathogen, Rhizoctonia solani infection) and drought stresses. Markerfree transgenic IR64 rice lines that simultaneously overexpress both RGB1 and RGG1 genes under CaMV35S promoter were raised. The overexpressing (OE) lines showed enhanced tolerance to R. solani infection and salinity/drought stresses. Several defense marker genes including OsMPK3 were signifcantly upregulated in the R. solani-infected OE lines. We also found the antioxidant machineries to be upregulated during salinity as well as drought stress in the OE lines. Overall, the present study provides evidence that concurrent overexpression of G-protein subunits (RGG1 and RGB1) impart multiple (both biotic and abiotic) stress tolerance in rice which could be due to the enhanced expression of stress-marker genes and better management of reactive oxygen species (ROS)-scavenging/photosynthetic machinery. The current study suggests an improved approach for simultaneous improvement of biotic and abiotic stress tolerance in rice which remains a major challenge for its sustainable cultivation.
Heterologous overexpression of PDH45 gene of pea provides tolerance against sheath blight disease and drought stress in rice
(Elsevier B.V., 2022) Sahoo, Ranjan Kumar; Chandan, Ravindra Kumar; Swain, Durga Madhab; Tuteja, Narendra; Jha, Gopaljee
Biotic and abiotic stress tolerant crops are required for sustainable agriculture as well as ensuring global food security. In a previous study, we have reported that heterologous overexpression of pea DNA helicase (PDH45), a DEAD-box family member protein, provides salinity stress tolerance in rice. The improved management of photosynthetic machinery and scavenging of reactive oxygen species (ROS) are associated with PDH45 mediated salinity stress tolerance. However, the role of PDH45 in biotic and other abiotic stress (drought) tolerance remains unexplored. In the present study, we have generated marker-free transgenic IR64 rice lines that overexpress PDH45 under the CaMV35S promoter. The transgenic rice lines exhibited a significant level of tolerance against sheath blight disease, caused by Rhizoctonia solani, a polyphagous necrotrophic fungal pathogen. The defense as well as antioxidant responsive marker genes were significantly upregulated in the PDH45 overexpressing (OE) rice lines, upon pathogen infection. Moreover, the OE lines exhibited tolerance to drought stress and various antioxidant as well as drought responsive marker genes were significantly upregulated in them, upon drought stress. Overall, the current study emphasizes that heterologous overexpression of PDH45 provides abiotic as well as biotic stress tolerance in rice. Tolerance against drought as well as sheath blight disease by overexpression of a single gene (PDH45) signifies the practical implication of the present study. Moreover, considering the conserved nature of the gene in different plant species, we anticipate that PDH45 can be gainfully deployed to impart tolerance against multiple stresses in agriculturally important crops.
A prophage tail-like protein facilitates the endophytic growth of Burkholderia gladioli and mounting immunity in tomato
(John Wiley & Sons, 2023) Chandan, Ravindra Kumar; Kumar, Rahul; Kabyashree, Kristi; Yadav, Sunil Kumar; Roy, Mandira; Swain, Durga Madhab; Jha, Gopaljee
A prophage tail-like protein (Bg_9562) of Burkholderia gladioli strain NGJ1 possesses broad-spectrum antifungal activity, and it is required for the bacterial ability to forage over fungi. Here, we analyzed whether heterologous overexpression of Bg_9562 or exogenous treatment with purified protein can impart disease tolerance in tomato. The physiological relevance of Bg_9562 during endophytic growth of NGJ1 was also investigated. Bg_9562 overexpressing lines demonstrate fungal and bacterial disease tolerance. They exhibit enhanced expression of defense genes and activation of mitogen-activated protein kinases. Treatment with Bg_9562 protein induces defense responses and imparts immunity in wild-type tomato. The defense-inducing ability lies within 18-51 aa region of Bg_9562 and is due to sequence homology with the bacterial flagellin epitope. Interaction studies suggest that Bg_9562 is perceived by FLAGELLIN-SENSING 2 homologs in tomato. The silencing of SlSERK3s (BAK1 homologs) prevents Bg_9562-triggered immunity. Moreover, type III secretion system-dependent translocation of Bg_9562 into host apoplast is important for elicitation of immune responses during colonization of NGJ1. Our study emphasizes that Bg_9562 is important for the endophytic growth of B. gladioli, while the plant perceives it as an indirect indicator of the presence of bacteria to mount immune responses. The findings have practical implications for controlling plant diseases.
A prophage tail-like protein is deployed by Burkholderia bacteria to feed on fungi
(Nature Publishing Group, 2017) Swain, Durga Madhab; Yadav, Sunil Kumar; Tyagi, Isha; Kumar, Rahul; Kumar, Rajeev; Ghosh, Srayan; Das, Joyati; Jha, Gopaljee
Some bacteria can feed on fungi, a phenomenon known as mycophagy. Here we show that a prophage tail-like protein (Bg_9562) is essential for mycophagy in Burkholderia gladioli strain NGJ1. The purified protein causes hyphal disintegration and inhibits growth of several fungal species. Disruption of the Bg_9562 gene abolishes mycophagy. Bg_9562 is a potential effector secreted by a type III secretion system (T3SS) and is translocated into fungal mycelia during confrontation. Heterologous expression of Bg_9562 in another bacterial species, Ralstonia solanacearum, confers mycophagous ability in a T3SS-dependent manner. We propose that the ability to feed on fungi conferred by Bg_9562 may help the bacteria to survive in certain ecological niches. Furthermore, considering its broad-spectrum antifungal activity, the protein may be potentially useful in biotechnological applications to control fungal diseases.
RAV1 family members function as transcriptional regulators and play a positive role in plant disease resistance
(John Wiley & Sons, 2023) Chandan, Ravindra Kumar; Kumar, Rahul; Swain, Durga Madhab; Ghosh, Srayan; Bhagat, Prakash Kumar; Patel, Sunita; Bagler, Ganesh; Sinha, Alok Krishna; Jha, Gopaljee
Phytopathogens pose a severe threat to agriculture and strengthening plant defense response is an important strategy for disease control. Here, we report that AtRAV1, an AP2 and B3 domain-containing transcription factor is required for basal plant defense in Arabidopsis thaliana. The atrav1 mutant lines demonstrate hyper-susceptibility against fungal pathogens (Rhizoctonia solani and Botrytis cinerea) while AtRAV1 overexpressing (OE) lines exhibit disease resistance against them. Enhanced expression of various defense genes and activation of MAP kinases (AtMPK3 and AtMPK6) are observed in the R. solani infected OE lines, but not in the atrav1 mutant plants. In-vitro phosphorylation assay suggests AtRAV1 to be a novel phosphorylation target of AtMPK3. The bimolecular fluorescence complementation and yeast two-hybrid assay support physical interactions between AtRAV1 and AtMPK3. Overexpression of the native as well as phospho-mimic but not the phospho-defective variant of AtRAV1 impart disease resistance in the atrav1 mutant A. thaliana lines. On the other hand, overexpression of AtRAV1 fails to impart disease resistance in the atmpk3 mutant. These analyses emphasize that AtMPK3-mediated phosphorylation of AtRAV1 is important for the elaboration of defense response in A. thaliana. Considering that RAV1 homologs are conserved in diverse plant species, we propose that they can be gainfully deployed to impart disease resistance in agriculturally important crop plants. Indeed, overexpression of SlRAV1 (a member of the RAV1 family) imparts disease tolerance against not only fungal (R. solani and B. cinerea) but also against bacterial (Ralstonia solanacearum) pathogens in tomato, while silencing of the gene enhances disease susceptibility.
Silencing of tomato CTR1 provides enhanced tolerance against Tomato leaf curl virus infection
(Taylor & Francis Group, 2019) Chandan, Ravindra K.; Singh, Achuit K.; Patel, Sunita; Swain, Durga Madhab; Tuteja, Narendra; Jha, Gopaljee
Tomato leaf curl virus(ToLCV) belonging to Begomovirus family of Geminivirus is known to cause one of the most destructive diseases in tomato. Amongst variousToLCVs, a monopartite Tomato leaf curl Joydebpur virus (ToLCJoV) is most prevalent in eastern part of India. In the present study, we observed induced expression of one of the negative regulators of ethylene signaling pathway gene (LeCTR1)in ToLCJoVinfected plants. TheTobacco rattle virus(TRV) induced silencing of theLeCTR1gene provided enhanced tolerance toToLCJoVinfections. The leaf curling as well as ROS accumulation was significantly reduced in the viral infectedLeCTR1silenced plants. Induction of several defense marker genes (NPR1, PR1, PR5, AOS2, EIN2, EIN3and ERF5) reinforced enhanced tolerance againstToLCJoVinfection in the LeCTR1silenced tomato. Overall, the present study provides evidence that silencing ofLeCTR1can be deployed to protect tomato fromToLCJoVinfections.