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Key Message: We report the size dependent uptake of dsRNA loaded MSNPs into the leaves and roots of Nicotiana benthamiana plants and accessed for their relative reduction in Tomato leaf curl New Delhi viral load. A non-GMO method of RNA interference (RNAi) has been recently in practice through direct delivery of double stranded RNA into the plant cells. Tomato leaf curl New Delhi virus (ToLCNDV), a bipartitie begomovirus, is a significant viral pathogen of many crops in the Indian subcontinent. Conventional RNAi cargo delivery strategies for instance uses viral vectors and Agrobacterium-facilitated delivery, exhibiting specific host responses from the plant system. In the present study, we synthesized three different sizes of amine-functionalized mesoporous silica nanoparticles (amino-MSNPs) to mediate the delivery of dsRNA derived from the AC2 (dsAC2) gene of ToLCNDV and showed that these dsRNA loaded nanoparticles enabled effective reduction in viral load. Furthermore, we demonstrate that amino-MSNPs protected the dsRNA molecules from nuclease degradation, while the complex was efficiently taken up by the leaves and roots of Nicotiana benthamiana. The real time gene expression evaluation showed that plants treated with nanoparticles of different sizes ~ 10 nm (MSNP DEA ), ~ 32 nm (MSNP TEA ) and ~ 66 nm (MSNP NH3 ) showed five-, eleven- and threefold reduction of ToLCNDV in N. benthamiana, respectively compared to the plants treated with naked dsRNA. This work clearly demonstrates the size dependent internalization of amino-MSNPs and relative efficacy in transporting dsRNA into the plant system, which will be useful in convenient topical treatment to protect plants against their pathogens including viruses. Mesoporous silica nanoparticles loaded with FITC, checked for its uptake into Nicotiana benthamiana., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Multiple begomovirus species are known to cause leaf curl disease in tomato in India. In order to develop specific and generic PCR based diagnostics for the tomato-infecting begomoviruses, in this study, we attempted to design primers initially based on the multiple alignment of the complete genome sequence of DNA-A component. However, the specific nucleotide stretches adequate for preparing specific primers could not be obtained. Alternatively, the online Primer-BLAST tool that offers designing of target-specific PCR primers was attempted to prepare specific primers targeting three clones (DNA-A) of tomato-infecting begomovirus species ( Tomato leaf curl New Delhi virus , Tomato leaf curl Palampur virus and Tomato leaf curl Joydebpur virus ) selected based on their sequence identity and phylogenetic relatedness. The primers derived from Primer-BLAST tool showed high level of cross-reaction among these begomovirus species and therefore were not able to differentiate these target begomovirus species. In order to understand the reason of cross-reactivity further sequence analysis revealed the high occurrence of single nucleotide variations (SNVs) compared to the multi-nucleotide stretches. There was no SNV hot-spot in the genome, rather the SNVs were randomly distributed throughout the genome of these begomovirus species. This pattern of nucleotide diversities among these tomato-infecting begomoviruses seriously implicated on developing specific PCR diagnostics. On the contrary, sequence analysis showed high sequence conservancy, which enabled to develop a generic PCR diagnostic for these begomoviruses. Our study, thus showed that the genome sequence diversity pattern among the tomato-infecting begomoviruses in India poses challenges in developing PCR based specific diagnostics., Supplementary Information: The online version contains supplementary material available at 10.1007/s13337-022-00785-9., (© The Author(s), under exclusive licence to Indian Virological Society 2022, Springer Nature or its licensor holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.)

In order to fulfil the growing need to replace fossil fuels, investigations exploring the production of biodiesel from agricultural biomass have gained attention. In this study, biodiesels were produced from Madhuca longifolia and Jatropha curcas by means of pre-treatment followed by a two-step acid-base homogeneous catalyst method. These biodiesels were blended with diesel at different percentages. The efficacy of the process was examined using various characterization methods while the efficiency of the produced biodiesels was examined by their engine performance and emission tests. Both Madhuca and Jatropha-based biodiesels exhibited physiochemical properties like that of diesel. Biodiesels were produced by pre-treating with orthophosphoric acid and toluene. The second step involves acid esterification, followed by base transesterification. Raman spectra exhibited C=O stretching at 1725 cm -1 indicating conversion of Madhuca and Jatropha oil into biodiesel. Fourier transform infrared spectroscopy showed a strong presence of fatty acid profile and triglyceride ester linkage at 1744 cm -1 . Ultraviolet-visible (UV) spectra confirmed the presence of conjugated dienes in the extracted biodiesels. UV absorbance at 320 nm decreased linearly with blend percentage. 1 H and 13 C nuclear magnetic resonance (NMR) confirmed the presence of methyl ester moiety at 3.6 δ (ppm) and methoxy carbon at 51.2 δ in biodiesel, distinguishing it from diesel. In the engine performance tests, the variations of brake specific fuel consumption, exhaust gas temperature and brake thermal efficiency versus brake power were studied. The emission tests of different blends were done in terms of carbon monoxide, nitrous oxide and unburnt hydrocarbon. The Jatropha biodiesel exhibited lower mean brake specific fuel consumption, exhaust gas temperature, emitted less carbon monoxide and unburnt hydrocarbon than Madhuca biodiesel. The average decrease in brake thermal efficiency was more in Jatropha biodiesel than Madhuca biodiesel. The present work uses for the first time treatment of ortho phosphoric acid and toluene to produce biodiesel followed by a two-step homogeneous acid-base catalyst method, drastically reducing free fatty acid value., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Biogeochemical release of soil-bound arsenic (As) governs mobilization of the toxic metalloid into the groundwater. The present study has examined As V -reduction ability of bacteria from anoxic aquatic sediments that might contribute to arsenic mobilization in the Bengal Delta. Arsenic-reducing bacteria from deep layers of pond sediment were enriched and isolated in anaerobic environments and As V reduction was assessed in culture medium. The pond sediment enrichments harboured As V -reducing bacteria belonging to the phyla Firmicutes and Proteobacteria with dominance of Paraclostridium benzoelyticum and P. bifermentans. Among total 17 isolates, the respiratory reductase genes were not detected by the most common primers and only 3 strains had arsenic reductase ArsC gene suggesting involvement of resistance and some unknown mechanisms in As V reduction. Presence of high levels of organic matter, As, and As-reducing bacteria might make deep aquatic sediments a hot spot of As mobilization and aquifer contamination., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Tomato leaf curl New Delhi virus (ToLCNDV), a bipartite begomovirus, is the most important among the 14 species of begomoviruses infecting tomato in Indian subcontinent. Begomovirus is known to  evade RNA silencing of host plants through suppressor proteins. However, in case of ToLCNDV, the suppressor proteins have not been studied well. The objective of the study is to know the sub-cellular localization of three suppressor proteins encoded by AV2, AC2 and AC4 ORFs of ToLCNDV in Nicotiana benthamiana . AV2, AC2 and AC4 ORFs of ToLCNDV were  cloned and sequenced (accession numbers MW423574, MW423576, MW423575, respectively) from a ToLCNDV isolate characterized earlier (accession number MW429271) and GFP tagged constructs were prepared in a plant expressing binary vector pEarleygate103. Bioinformatics analysis using Peptide 2.0 server predicted that all these proteins have more basic amino acid residues then acidic amino acid and AV2 protein has more hydrophobic amino acid residues. ScanProsite server predicted presence of different fuctional motifs in these proteins amongst which presence of kinase motif was observed in all of them. Virus mPLoc server predicted their subcellular localization. The suppressor gene constructs were agroinfiltrated on to leaves of one month old N. benthamiana plants and their subcellular localization has been studied through confocal microscopy. Results have shown that AV2 localizes in the host cell membrane and nucleus, AC2 in the nucleus and AC4 in the host cell membrane. Earlier reports with other begomoviruses also showed similar localization behaviour of these suppressor protein except AV2, where it was shown to be present in cytoplasm. Such localization study will help understand the mechanism of their suppression activity., (© Indian Virological Society 2021.)

RNA silencing plays a key role in shielding plant and animal hosts against viral invasion and infection. Viruses encode RNA silencing suppressors (RSS) to block small RNA guided silencing of viral transcripts. The B2 protein encoded by Flock House virus (FHV) is a well-characterized RSS that facilitates infection in insects. It has been shown to act as a functional RSS in plants. FHVB2 over-expressing tobacco plants were used to study the effect of RSS on plant susceptibility to Tobacco mosaic virus (TMV) , its natural pathogen. The major symptoms observed in TMV-infected transgenic plants were greenish mosaic, puckering and distortion of leaves, but the infected transgenic leaves were able to resist chlorophyll loss. The infected leaves of transgenic plants showed no significant difference in accumulation of virus when compared with that of the wild type plants. FHVB2 plants showed higher levels of H 2 O 2 and the ROS scavenging enzymes, APX and SOD. This suggests that interference of FHVB2 with RNA silencing machinery may activate alternative defense pathways in the plants so that they are not overly sensitive to TMV infection., Supplementary Information: The online version contains supplementary material available at 10.1007/s13337-020-00644-5., Competing Interests: Conflicts of interestThe authors declare no competing financial interests., (© Indian Virological Society 2021.)

Characterization of the subgenomic RNA (sgRNA) promoter of many plant viruses is important to understand the expression of downstream genes and also to configure their genome into a suitable virus gene-vector system. Cucumber green mottle mosaic virus (CGMMV, genus Tobamovirus ) is one of the RNA viruses, which is extensively being exploited as the suitable gene silencing and protein expression vector. Even though, characters of the sgRNA promoters (SGPs) of CGMMV are yet to be addressed. In the present study, we predicted the SGP for the movement protein (MP) and coat protein (CP) of CGMMV. Further, we identified the key regulatory elements in the SGP regions of MP and CP, and their interactions with the core RNA dependent RNA polymerase (RdRp) domain of CGMMV was deciphered. The modeled structure of core RdRp contains two palm (1-41 aa, and 63-109 aa), one finger (42-62 aa) subdomains with three conserved RdRp motifs that played important role in binding to the SGP nucleic acids. RdRp strongly preferred the double helix form of the stem region in the stem and loop (SL) structures, and the internal bulge elements. In MP-SGP, a total of six elements was identified; of them, the affinity of binding to - 26 nt to - 17 nt site (CGCGGAAAAG) was higher through the formation of strong hydrogen bonds with LYS16, TYR17, LYS19, SER20, etc. of the motif A in the palm subdomain of RdRp. Similar strong interactions were noticed in the internal bulge (CAACUUU) located at + 33 to + 39 nt adjacent to the translation start site (TLSS) (+ 1). These could be proposed as the putative core promoter elements in MP-SGP. Likewise, total five elements were predicted within - 114 nt to + 144 nt region of CP-SGP with respect to CP-TLSS. Of them, RdRp preferred to bind at the small hairpin located at - 60 nt to - 43 nt (UUGGAGGUUUAGCCUCCA) in the upstream region, and at the complex duplex structure spanning between + 99 and + 114 nt in the downstream region, thus indicating the distribution of core promoter within - 60 nt to + 114 nt region of CP-SGP with respect to TLSS (+ 1) of the CP; whereas, the - 114 nt to + 144 nt region of CP-SGP might be necessary for the full activity of the CP-SGP. Our in silico prediction certifies the gravity of these nucleotide stretches as the RNA regulatory elements and identifies their potentiality for binding with of palm and finger sub-domain of RdRp. Identification of such elements will be helpful to anticipate the critical length of the SGPs. Our finding will not only be helpful to delineate the SGPs of CGMMV but also their subsequent application in the efficient construction of virus gene-vector for the expression of foreign protein in plant., Competing Interests: Conflict of interestThe authors declare that they have no conflict of interest., (© Indian Virological Society 2020.)

Increasing bacterial resistance to common drugs is a major public health concern for the treatment of infectious diseases. Certain naturally occurring compounds of plant sources have long been reported to possess potential antimicrobial activity. This study was aimed to investigate the antibacterial activity and possible mechanism of action of andrographolide (Andro), a diterpenoid lactone from a traditional medicinal herb Andrographis paniculata. Extent of antibacterial action was assessed by minimal bactericidal concentration method. Radiolabeled N-acetyl glucosamine, leucine, thymidine, and uridine were used to determine the effect of Andro on the biosyntheses of cell wall, protein, DNA, and RNA, respectively. In addition, anti-biofilm potential of this compound was also tested. Andro showed potential antibacterial activity against most of the tested Gram-positive bacteria. Among those, Staphylococcus aureus was found to be most sensitive with a minimal inhibitory concentration value of 100 μg/mL. It was found to be bacteriostatic. Specific inhibition of intracellular DNA biosynthesis was observed in a dose-dependent manner in S. aureus. Andro mediated inhibition of biofilm formation by S. aureus was also found. Considering its antimicrobial potency, Andro might be accounted as a promising lead for new antibacterial drug development.

microRNAs (miRs) are 21- to 24-nucleotide-long RNA molecules that are mainly involved in regulating the gene expression at the post-transcriptional levels. They are present in a variety of organisms from algae to plants and play an important role in gene regulation. The identification of several diverging and converging functions of miRs indicates that they play versatile roles in regulating plant development including differentiation, organ development, phase change, signalling, disease resistance and response to environmental stresses. This article provides a concise update on the plant miR functions and their targets in the auxin pathway with focus on the interactions between miRs and auxin signalling to intricately regulate the plant responses.

In previous studies with mice the oxygen radical generating neurotoxin tertiary butylhydroperoxide (t-BuOOH) was used to mimic the oxidative injury that has been implicated in neurodegenerative diseases. In addition, previous studies have shown that the poly (ADP-ribose) polymerase (PARP) inhibitor nicotinamide is able to prevent DNA fragmentation and apoptosis that is induced by t-BuOOH in mouse brain. However, the molecular mechanism(s) by which nicotinamide is able to protect human brain cells at the cellular level is not clear. Therefore in this study a cell culture model system with human cortical neuronal cells (HCN2 cells) has been employed where the molecular mechanism(s) of nicotinamide action, both in the presence and absence of t-BuOOH has been studied. Human cortical neurons (HCN2 cells) have been shown to differentiate to a neuron-like morphology. In this study, exposure of HCN2 cells to t-BuOOH resulted in altered morphology and disruption of neuronal differentiation leading to cell death. However, in neurons, which were treated with nicotinamide before being exposed to t-BuOOH, neuronal differentiation was preserved; morphological disruption was prevented and cell death was reduced significantly. Moreover, our studies indicate that nicotinamide is able to prevent the up-regulation of the pro-apoptotic proteins p53 and p21/WAF-1, and the down-regulation of the anti-apoptotic protein bcl-2 that is induced by t-BuOOH in HCN2 cells. Thus this study indicates that nicotinamide protects human brain cells from the toxic effects of free radical generating toxins by regulating the levels of various pro- and anti-apoptotic proteins.

Effects of an inhibitor of membrane anion-exchange transport processes, 4-acetamido-4'-isothiocyano-2,2'-disulfonic stilbene (SITS), on urate transport by isolated, perfused snake (Thamnophis spp.) proximal renal tubules were studied. SITS (10(-4) mol/l) in the luminal perfusate had absolutely no effect on net urate secretion (Jneturate) or on net fluid absorption (JV). This observation is compatible with other data that give no support to the concept of a mediated transport step for urate from the cells to the lumen. SITS (10(-4) mol/l) in the bathing medium reversibly inhibited Jneturate without affecting JV. At the time of maximum inhibition of Jneturate, the concentration of urate in the cell water was increased and the apparent permeability of the luminal membrane to urate was decreased, but the urate efflux across the peritubular membrane and the apparent permeability of the peritubular membrane to urate were unchanged. There was no evidence of significant intracellular binding or trapping of urate. Although an increase in the initial rate of urate transport into the cells across the peritubular membrane could not be demonstrated conclusively in nonperfused tubules, the results still suggest that SITS in the bathing medium may inhibit Jneturate by inhibiting urate movement from the cells to the lumen while actually enhancing transport from the bathing medium into the cells.