Phylogenetic analysis, molecular characterization and virulence profiling based on toxoflavin gene of an Indian BG1 strain of Burkholderia glumae causing panicle blight of rice.

Bacterial panicle blight (BPB) caused by Burkholderia glumae (BG) has become significantly more prevalent in the rice-growing regions of North India. Based on virulence screening and in vitro quantification of toxoflavin, the BG strains were classified as hyper- (BG1 and BG3), moderate- (BG2, BG4, BG6, BG8, and BG9), and hypo- (BG5, BG7, and BG10) virulent. Plant inoculation assays with cell-free culture filtrate revealed strains with higher toxoflavin-producing ability had higher virulence. Based on 16S rRNA sequence, 6 isolates from Uttar Pradesh were grouped in clad C1; whereas, clad C2 exhibited 4 isolates, two each from Delhi and Uttar Pradesh. Strain BG1 being the most virulent Indian strain from Uttar Pradesh was further profiled for 11 tox genes. We found all the 11 tox genes present in strain BG1. In toxRABCDE cluster, all tox genes showed high similarity to B. glumae BGR1 except toxB, whereas in toxFGHIJ cluster toxF, toxG, toxH and toxI shared maximum similarity to B. glumae 336gr-1. tox genes of BG1 exhibited homology as well as divergence with B. gladioli. The domain prediction and protein association network analysis indicated the possible involvement of tox genes in the toxoflavin biosynthesis. As per our knowledge, this is the first report in India on characterization of tox genes cluster in B. glumae. Altogether, our study unravels a reliable method for identifying and characterizing B. glumae using tox genes and its relationship with disease production. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03660-6.

Rice transcriptome upon infection with Xanthomonas oryzae pv. oryzae relative to its avirulent T3SS-defective strain exposed modulation of many stress responsive genes.

Xanthomonas oryzae pv. oryzae (Xoo) is a destructive pathogen that causes bacterial blight disease of rice worldwide. Xoo uses T3SS (type III secretion system) effectors to subvert rice innate immunity. However, the comprehensive knowledge of rice genes involved in T3SS effectors-mediated interaction remains unclear. In this study, the transcriptome profiles of rice infected with a virulent Xoo strain from North-eastern region of India relatives to its avirulent strain (that lacks functional T3SS) were analyzed at early (2-6 hpi) and late (16-24 hpi) hours of infection. Out of total 255 differentially expressed genes (DEGs), during early infection, 62 and 70 genes were upregulated and downregulated, respectively. At late infection, 70 and 53 genes were upregulated and downregulated, respectively. The transcriptomic data identified many differentially expressed resistant genes, transposons, transcription factors, serine/threonine protein kinase, cytochrome P450 and peroxidase genes that are involved in plant defense. Pathway analysis revealed that these DEGs are involved in hormone signaling, plant defense, cellular metabolism, growth and development processes. DEGs associated with plant defense were also validated through quantitative real-time PCR. Our study brings a comprehensive picture of the rice genes that are being differentially expressed during bacterial blight infection. Nevertheless, the DEG-associated pathways would provide sensible targets for developing resistance to bacterial blight. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03193-4.

Xanthomonas axonopodis pv. punicae depends on multiple non-TAL (Xop) T3SS effectors for its coveted growth inside the pomegranate plant through repressing the immune responses during bacterial blight development.

Xanthomonas axonopodis pv. punicae (Xap), the bacterial blight pathogen of pomegranate, incurs substantial loss to yield and reduces export quality of this economically important fruit crop. During infection, the bacterium secretes six non-TAL (Xop) effectors into the pomegranate cells through a specialized type three secretion system (T3SS). Previously, we demonstrated the role of two key effectors, XopL and XopN in pathogenesis. Here, we investigate the role of rest effectors (XopC2, XopE1, XopQ and XopZ) on disease development. We generated null mutants for each individual effector and mutant bacterial suspension was infiltrated into pomegranate leaves. Compared to Xap wild, the mutant bacterial growth was reduced by 2.7-11.5 folds. The mutants produced lesser water-soaked lesions when infiltrated on leaves by 1.13-2.21 folds. Among the four effectors, XopC2 contributes highest for in planta bacterial growth and disease development. XopC2 efficiently suppressed the defense responses like callose deposition, reactive oxygen species (ROS) and the activation of immune responsive genes. Being a major contributor, we further characterize XopC2 for its subcellular localization, its protein structure and networking. XopC2 is localized to the plasma membrane of Nicotiana benthamiana like XopL and XopN. XopC2 is a 661 amino acids protein having 15 alpha and 17 beta helix. Our STRING and I-TASSER based analysis hinted that XopC2 interacts with multiple membrane localized plant proteins including transcription regulator of CCR4-NOT family, TTN of maintenance of chromosome family and serine/threonine-protein phosphatase 2A (PP2A) isoform. Based on the interaction it is predicted that XopC2 might involve in diverse functions like nuclear-transcribed mRNA catabolic process, maintenance of chromosome, hormone signaling and protein dephosphorylation activities and thereby suppress the plant immunity. Altogether, our study suggests that Xap largely depends on three non-TAL (Xop) effectors, including XopC2, XopL and XopN, to modulate pomegranate PTI for its unrestricted proliferation during bacterial blight development.