A small cysteine-rich fungal effector, BsCE66 is essential for the virulence of Bipolaris sorokiniana on wheat plants.

The Spot Blotch (SB) caused by hemibiotrophic fungal pathogen Bipolaris sorokiniana is one of the most devastating wheat diseases leading to 15-100% crop loss. However, the biology of Triticum-Bipolaris interactions and host immunity modulation by secreted effector proteins remain underexplored. Here, we identified a total of 692 secretory proteins including 186 predicted effectors encoded by B. sorokiniana genome. Gene Ontology categorization showed that these proteins belong to cellular, metabolic and signaling processes, and exhibit catalytic and binding activities. Further, we functionally characterized a cysteine-rich, B. sorokiniana Candidate Effector 66 (BsCE66) that was induced at 24-96 hpi during host colonization. The Δbsce66 mutant did not show vegetative growth defects or stress sensitivity compared to wild-type, but developed drastically reduced necrotic lesions upon infection in wheat plants. The loss-of-virulence phenotype was rescued upon complementing the Δbsce66 mutant with BsCE66 gene. Moreover, BsCE66 does not form homodimer and conserved cysteine residues form intra-molecular disulphide bonds. BsCE66 localizes to the host nucleus and cytosol, and triggers a strong oxidative burst and cell death in Nicotiana benthamiana. Overall, our findings demonstrate that BsCE66 is a key virulence factor that is necessary for host immunity modulation and SB disease progression. These findings would significantly improve our understanding of Triticum-Bipolaris interactions and assist in the development of SB resistant wheat varieties.

Biotechnological Interventions in Tomato (Solanum lycopersicum) for Drought Stress Tolerance: Achievements and Future Prospects.

Tomato production is severely affected by abiotic stresses (drought, flood, heat, and salt) and causes approximately 70% loss in yield depending on severity and duration of the stress. Drought is the most destructive abiotic stress and tomato is very sensitive to the drought stress, as cultivated tomato lack novel gene(s) for drought stress tolerance. Only 20% of agricultural land worldwide is irrigated, and only 14.51% of that is well-irrigated, while the rest is rain fed. This scenario makes drought very frequent, which restricts the genetically predetermined yield. Primarily, drought disturbs tomato plant physiology by altering plant-water relation and reactive oxygen species (ROS) generation. Many wild tomato species have drought tolerance gene(s); however, their exploitation is very difficult because of high genetic distance and pre- and post-transcriptional barriers for embryo development. To overcome these issues, biotechnological methods, including transgenic technology and CRISPR-Cas, are used to enhance drought tolerance in tomato. Transgenic technology permitted the exploitation of non-host gene/s. On the other hand, CRISPR-Cas9 technology facilitated the editing of host tomato gene(s) for drought stress tolerance. The present review provides updated information on biotechnological intervention in tomato for drought stress management and sustainable agriculture.

Development of agro-infectious clones for screening resistance against recombinant mungbean yellow mosaic India virus causing golden mosaic disease in vegetable cowpea.

Begomovirus associated with golden mosaic disease on vegetable cowpea has been characterized through rolling circle amplification. The genomic components (DNA A and DNA B) were cloned and sequenced. Nucleotide sequence analysis of DNA A (MT671430) and DNA B (MT671431) component had > 98% identity toward the mungbean yellow mosaic India virus (MYMIV) reported previously from India on various legumes. In phylogenetic analysis, study isolate shared common ancestry with MYMIV isolates of India, Pakistan and Nepal infecting legumes. Based on the recombination analysis, this cowpea isolate appears to be evolved through recombination of MYMIV sequences both at DNA A (Major parent: AF481855; Minor parent: AF416742) and DNA B (Major parent: AF416741; Minor parent: MN698281) level. Furthermore, Agrobacterium-based dimeric clone constructs were found highly infectious on cowpea host upon co-inoculation of DNA-A and DNA-B components by producing typical golden mosaic symptoms 42 days post-inoculation. Upon inoculation of these agro-infectious clones, vegetable cowpea germplasm lines were categorized as resistant, moderately resistant and susceptible to golden mosaic disease. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03206-2.

Plant Secondary Metabolites as Defense Tools against Herbivores for Sustainable Crop Protection.

Plants have evolved several adaptive strategies through physiological changes in response to herbivore attacks. Plant secondary metabolites (PSMs) are synthesized to provide defensive functions and regulate defense signaling pathways to safeguard plants against herbivores. Herbivore injury initiates complex reactions which ultimately lead to synthesis and accumulation of PSMs. The biosynthesis of these metabolites is regulated by the interplay of signaling molecules comprising phytohormones. Plant volatile metabolites are released upon herbivore attack and are capable of directly inducing or priming hormonal defense signaling pathways. Secondary metabolites enable plants to quickly detect herbivore attacks and respond in a timely way in a rapidly changing scenario of pest and environment. Several studies have suggested that the potential for adaptation and/or resistance by insect herbivores to secondary metabolites is limited. These metabolites cause direct toxicity to insect pests, stimulate antixenosis mechanisms in plants to insect herbivores, and, by recruiting herbivore natural enemies, indirectly protect the plants. Herbivores adapt to secondary metabolites by the up/down regulation of sensory genes, and sequestration or detoxification of toxic metabolites. PSMs modulate multi-trophic interactions involving host plants, herbivores, natural enemies and pollinators. Although the role of secondary metabolites in plant-pollinator interplay has been little explored, several reports suggest that both plants and pollinators are mutually benefited. Molecular insights into the regulatory proteins and genes involved in the biosynthesis of secondary metabolites will pave the way for the metabolic engineering of biosynthetic pathway intermediates for improving plant tolerance to herbivores. This review throws light on the role of PSMs in modulating multi-trophic interactions, contributing to the knowledge of plant-herbivore interactions to enable their management in an eco-friendly and sustainable manner.

Overexpression of AtDREB1 and BcZAT12 genes confers drought tolerance by reducing oxidative stress in double transgenic tomato (Solanum lycopersicum L.).

KEY MESSAGE: Double transgenic tomato developed by AtDREB1A and BcZAT12 genes pyramiding showed significant drought tolerance by reducing oxidative stress with enhanced yield. Although a large number of efforts have been made by different researchers to develop abiotic stress tolerance tomato for improving yield using single gene, however, no reports are available which targets AtDREB1 and BcZAT12 genes together. Hence, in the present study, double transgenic plants were developed using AtDREB1 and BcZAT12 genes to improve yield potential with better drought tolerance. Double transgenic (DZ1-DZ5) tomato lines showed enhanced drought tolerance than their counterpart non-transgenic and single transgenic plants at 0, 07, 14, and 21 days of water deficit, respectively. Double transgenic plants showed increased activity of antioxidant enzymes, like catalase (CAT), superoxide dismutase (SOD), glutathione reductase (GR), ascorbate peroxidase (APX), dehydroascorbate reductase (DHAR), monodehydroascorbate reductase (MDHAR) and guaiacol peroxidase (POD), and accumulation of non-enzymatic antioxidants like ascorbic acid, glutathione as compared to non-transgenic and single transgenic. Additionally, the transcript analysis of antioxidant enzymes revealed the increased level of gene expression in double transgenic tomato lines. Developed double-transgenic tomato plants co-over-expressing both genes exhibited more enzymatic and non-enzymatic anti-oxidative activities as compared to the non-transgenic and single transgenic control, respectively. This is the preliminary report in tomato, which forms the basis for a multigene transgenic approach to cope with drought stress.

Characterization of high-temperature stress-tolerant tomato (Solanum lycopersicum L.) genotypes by biochemical analysis and expression profiling of heat-responsive genes.

High-temperature stress severely impacts both yield and quality of tomato fruits, and therefore, it is required to develop stress-tolerant cultivars. In the present study, two tomato genotypes, H88-78-1 and CLN-1621, identified through preliminary phenotypic screening were characterized by analysis of molecular, physiological, and biochemical traits in comparison with a susceptible genotype Punjab Chhuhara. Phenotypic stress tolerance of both the genotypes was validated at biochemical level as they showed higher amount of relative water content, photosynthetic pigments, free cellular proline, and antioxidant molecules while less amount of H2O2 and electrolyte leakage. Expression analysis of 67 genes including heat shock factors, heat shock proteins, and other stress-responsive genes showed significant up-regulation of many of the genes such as 17.4 kDa class III heat shock protein, HSF A-4a, HSF30, HSF B-2a, HSF24, HSF B-3 like, 18.1 kDa class I HSP like, and HSP17.4 in H88-78-1 and CLN-1621 after exposure to high-temperature stress. These candidate genes can be transferred to cultivated varieties by developing gene-based markers and marker-assisted breeding. This confirms the rapid response of these genotypes to high-temperature stress. All these traits are characteristics of a stress-tolerance and establish them as candidate high-temperature stress-tolerant genotypes that can be effectively utilized in stress tolerance improvement programs. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-020-02587-6.

Complexity of begomovirus and betasatellite populations associated with chilli leaf curl disease in India.

Chilli, which encompasses several species in the genus Capsicum, is widely consumed throughout the world. In the Indian subcontinent, production of chilli is constrained due to chilli leaf curl disease (ChiLCD) caused by begomoviruses. Despite the considerable economic consequences of ChiLCD on chilli cultivation in India, there have been scant studies of the genetic diversity and structure of the begomoviruses that cause this disease. Here we report on a comprehensive survey across major chilli-growing regions in India. Analysis of samples collected in the survey indicates that ChiLCD-infected plants are associated with a complex of begomoviruses (including one previously unreported species) with a diverse group of betasatellites found in crops and weeds. The associated betasatellites neither enhanced the accumulation of the begomovirus components nor reduced the incubation period in Nicotiana benthamiana. The ChiLCD-associated begomoviruses induced mild symptoms on Capsicum spp., but both the level of helper virus that accumulated and the severity of symptoms were increased in the presence of cognate betasatellites. Interestingly, most of the begomoviruses were found to be intra-species recombinants. The betasatellites possess high nucleotide variability, and recombination among them was also evident. The nucleotide substitution rates were determined for the AV1 gene of begomoviruses (2.60 × 10- 3 substitutions site- 1 year- 1) and the ßC1 gene of betasatellites [chilli leaf curl betasatellite (ChiLCB), 2.57 × 10- 4 substitution site- 1 year- 1; tomato leaf curl Bangladesh betasatellite (ToLCBDB), 5.22 × 10- 4 substitution site- 1 year- 1]. This study underscores the current understanding of Indian ChiLCD-associated begomoviruses and also demonstrates the crucial role of betasatellites in severe disease development in Capsicum spp.