Begomovirus (family-Geminiviridae) infects a wide range of commercial crops like tomato, bean, cassava, cotton, cucurbits and chilli. Purification of begomoviruses from the infected plants, in particular from vegetable crops remains challenging. The conventional process of begomovirus purification requires sophisticated instruments and moreover, it is time-consuming. Herein, we used antibody-conjugated magnetic nanoparticles (Ab-MNPs) to purify begomoviruses from the infected plants. MNPs were prepared using the co-precipitation method (at pH between 8 and 12 & size 25 nm). The prepared MNPs were functionalized with APTES (at pH 7) and confirmed with FTIR. Thus functionalized MNPs were conjugated with polyclonal antibodies (pAbs) using EDC–NHS chemistry (size = 80 nm). The crude extract prepared from the infected plants was suspended in the solution of Ab-MNPs and separated using a magnet. The captured virus particles were released into the aqueous solution (at pH 10). SDS-PAGE analysis and PCR analysis were done to confirm the presence of viral infection.

The DNA templated self-assembly of gold nanoparticles clustered in different configurations (nn = 2–∞) was investigated in the colorimetric detection of ToLCNDV DNA using a gold nanoparticle conjugated bifunctional oligonucleotide probe.

Colletotrichum gloeosporioides is the most destructive endophytic plant-pathogenic fungi causing anthracnose disease in a wide number of economically important plants throughout the world. Presently there are no existing methods for effective disease management over the outbreak of anthracnose disease and it thus leads to huge economic losses for farmers and plant tissue culture laboratories. In order to find a new and effective control over these endophytes, we synthesized and characterized bioactive bile salt sodium deoxycholate (NaDC) capped silver nanoparticles. These nanoparticles were employed to control the endophytic fungus through in vitro direct and indirect model systems with time dependant/light mediated incubation manner. In our findings we achieved fivefold synergistic effect of NaDC-capped AgNPs with their bioactive capping agent against Colletotrichum gloeosporioides upon analyzing different parameters. Moreover, it was evident that NaDC-capped AgNPs did not cause any phytotoxicity to treated plants as revealed by molecular marker studies.

Scanometric detection of tomato leaf curl New Delhi viral DNA using AuNP-conjugated mono- and bifunctional oligo probes through direct DNA hybridization assay (DDH assay) and sandwich DNA hybridization assay (SDH assay) with silver enhancement was developed. Tomato leaf curl New Delhi virus (ToLCNDV) coat protein gene-specific thiol-modified ssoligo probes were used for the preparation of mono- and bifunctional AuNP-ssoligo probe conjugates (signal probes). ssDNA arrays were prepared using polymerase chain reaction (PCR), rolling circle amplification (RCA), genomic DNAs fragments, and phosphate-modified positive control/capture probes through 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide/1-methylimidazole conjugation on the amine-modified glass slide (GS) surface. In the DDH assay, signal probes were directly hybridized with ssDNA array of positive control and ToLCNDV DNA samples and the detection signals were amplified by silver enhancement. Dark black/gray colors were developed on the GS by the result of Ag enhancement, which can be visualized and discriminated by the naked eye. The images were captured using a simple flatbed scanner, and the determined amounts of signal probes were hybridized with their target DNA. Similarly, the SDH assay also performed through two rounds of hybridization between capture probes and target DNA; target DNA and signal probes followed by silver enhancement. The detection signals were found higher in the PCR sample than the RCA and genomic DNA samples because of the presence of increased copy numbers of complementary DNAs in PCR samples. Further, bifunctional AuNP-ssoligo probe shows higher intensity of detection signal than monofunctional probes because it can be hybridized with both strands of dsDNA targets. Moreover, the DDH-based scanometric method showed higher detection sensitivity than the SDH assay-based scanometric method. Overall, bifunctional signal probes showed more detection sensitivity than monofunctional probes in scanometric methods based on both DDH and SDH assays. The limit of detection of this developed scanometric method was optimized (100 zM to 100 pM concentration). Further, DDH assay-based scanometric method shows significant advantages over the SDH assay method, such as cost-effectiveness, because it requires only single probes (signal probes), less time-consuming by the need of only single-step hybridization, and higher detection sensitivity (up to zM). To the best of our knowledge, this is the first attempt made to develop a scanometric-based nanoassay method for the detection of plant viral DNA. This approach will be a remarkable milestone for the application of nanotechnology in the development of nanobiosensor for plant pathogen detection.