Role of a Novel I1781T Mutation and Other Mechanisms in Conferring Resistance to Acetyl-CoA Carboxylase Inhibiting Herbicides in a Black-Grass Population

Background

Knowledge of the mechanisms of herbicide resistance is important for designing long term sustainable weed management strategies. Here, we have used an integrated biology and molecular approach to investigate the mechanisms of resistance to acetyl-CoA carboxylase inhibiting herbicides in a UK black-grass population (BG2).

Methodology/Principal Findings

Comparison between BG2 phenotypes using single discriminant rates of herbicides and genotypes based on ACCase gene sequencing showed that the I1781L, a novel I1781T, but not the W2027C mutations, were associated with resistance to cycloxydim. All plants were killed with clethodim and a few individuals containing the I1781L mutation were partially resistant to tepraloxydim. Whole plant dose response assays demonstrated that a single copy of the mutant T1781 allele conferred fourfold resistance levels to cycloxydim and clodinafop-propargyl. In contrast, the impact of the I1781T mutation was low (Rf = 1.6) and non-significant on pinoxaden. BG2 was also characterised by high levels of resistance, very likely non-target site based, to the two cereal selective herbicides clodinafop-propargyl and pinoxaden and not to the poorly metabolisable cyclohexanedione herbicides. Analysis of 480 plants from 40 cycloxydim resistant black grass populations from the UK using two very effective and high throughput dCAPS assays established for detecting any amino acid changes at the 1781 ACCase codon and for positively identifying the threonine residue, showed that the occurrence of the T1781 is extremely rare compared to the L1781 allele.

Conclusion/Significance

This study revealed a novel mutation at ACCase codon position 1781 and adequately assessed target site and non-target site mechanisms in conferring resistance to several ACCase herbicides in a black-grass population. It highlights that over time the level of suspected non-target site resistance to some cereal selective ACCase herbicides have in some instances surpassed that of target site resistance, including the one endowed by the most commonly encountered I1781L mutation.     

Global Wild Annual Lens Collection: A Potential Resource for Lentil Genetic Base Broadening and Yield Enhancement

Crop wild relatives (CWRs) are invaluable gene sources for various traits of interest, yet these potential resources are themselves increasingly threatened by the impact of climate change as well as other anthropogenic and socio-economic factors. The prime goal of our research was to cover all aspects of wild Lens genetic resource management like species characterization, agro-morphological evaluation, diversity assessment, and development of representative sets for its enhanced utilization in lentil base broadening and yield improvement initiatives. We characterized and evaluated extensively, the global wild annual Lens taxa, originating from twenty seven counties under two agro-climatic conditions of India consecutively for three cropping seasons. Results on various qualitative and quantitative characters including two foliar diseases showed wide variations for almost all yield attributing traits including multiple disease resistance in the wild species, L. nigricans and L. ervoides accessions. The core set developed from the entire Lens taxa had maximum representation from Turkey and Syria, indicating rich diversity in accessions originating from these regions. Diversity analysis also indicated wide geographical variations across genepool as was reflected in the core set. Potential use of core set, as an initial starting material, for genetic base broadening of cultivated lentil was also suggested.     

Antiplasmodial activity of novel keto-enamine chalcone-chloroquine based hybrid pharmacophores

A series of novel keto-enamine chalcone-chloroquine based hybrids were synthesized following new methodology developed in our laboratory. The synthesized compounds were screened against chloroquine sensitive strain (3D7) of Plasmodium falciparum in an in vitro model. Some of the compounds were showing comparable antimalarial activity at par with chloroquine. Compounds with significant in vitro antimalarial activity were then evaluated for their in vivo efficacy in Swiss mice against Plasmodium yoelii (chloroquine resistant N-67 strain), wherein compounds 25 and 27 each showed an in vivo suppression of 99.9% parasitaemia on day 4. Biochemical studies reveal that inhibition of hemozoin formation is the primary mechanism of action of these analogues.     

Recent advances in development of marker-free transgenic plants: Regulation and biosafety concern

During the efficient genetic transformation of plants with the gene of interest, some selectable marker genes are also used in order to identify the transgenic plant cells or tissues. Usually, antibiotic- or herbicide-selective agents and their corresponding resistance genes are used to introduce economically valuable genes into crop plants. From the biosafety authority and consumer viewpoints, the presence of selectable marker genes in released transgenic crops may be transferred to weeds or pathogenic microorganisms in the gastrointestinal tract or soil, making them resistant to treatment with herbicides or antibiotics, respectively. Sexual crossing also raises the problem of transgene expression because redundancy of transgenes in the genome may trigger homology-dependent gene silencing. The future potential of transgenic technologies for crop improvement depends greatly on our abilities to engineer stable expression of multiple transgenic traits in a predictable fashion and to prevent the transfer of undesirable transgenic material to non-transgenic crops and related species. Therefore, it is now essential to develop an efficient marker-free transgenic system. These considerations underline the development of various approaches designed to facilitate timely elimination of transgenes when their function is no longer needed. Due to the limiting number of available selectable marker genes, in future the stacking of transgenes will be increasingly desirable. The production of marker-free transgenic plants is now a critical requisite for their commercial deployment and also for engineering multiple and complex trait. Here we describe the current technologies to eliminate the selectable marker genes (SMG) in order to develop marker-free transgenic plants and also discuss the regulation and biosafety concern of genetically modified (GM) crops.     

Evidence of a humoral immune response against the prokaryotic expressed N-terminal autoprotease (N<Superscript>pro</Superscript>) protein of bovine viral diarrhoea virus

Bovine viral diarrhoea virus (BVDV) is an economically important pathogen of cattle and sheep belonging to the genus Pestivirus of the family Flaviviridae. Although the BVDV non-structural N-terminal protease (N^pro) acts as an interferon antagonist and subverts the host innate immunity, little is known about its immunogenicity. Hence, we expressed a recombinant BVDV N^pro-His fusion protein (28 kDa) in E. coli and determined the humoral immune response generated by it in rabbits. The antigenicity of the N^pro protein was confirmed by western blot using anti-BVDV hyperimmune cattle, sheep and goat serum, and anti-N^pro rabbit serum. When rabbits were immunized with the N^pro protein, a humoral immune response was evident by 4 weeks and persisted till 10 weeks post immunization as detected by ELISA and western blot. Despite N^pro-specific antibodies remaining undetectable in 80 serum samples from BVDV-infected sheep and goats, BVDV hyperimmune sera along with some of the field cattle, sheep and goat sera with high BVDV neutralizing antibody titres were found positive for N^pro antibodies. Our results provide evidence that despite the low immunogenicity of the BVDV N^pro protein, a humoral immune response is induced in cattle, sheep and goats only with repeated BVDV exposure.     

Antimicrobial Efficacy of Synthetic Pyranochromenones and (Coumarinyloxy)acetamides

Four (1, 2, 4 and 6) synthetic quaternary ammonium derivatives of pyranochromenones and (coumarinyloxy)acetamides were synthesized and investigated for their antimicrobial efficacy on MRSA (Methicillin-resistant Staphylococcus aureus), and multi-drug resistant Pseudomonas aeruginosa, Salmonella enteritidis and Mycobacterium tuberculosis H37Rv strain. One of the four compounds screened i.e. N,N,N-triethyl-10-((4,8,8-trimethyl-2-oxo-2,6,7,8-tetrahydropyrano[3,2-g]chromen-10-yl)oxy)decan-1-aminium bromide (1), demonstrated significant activity against S. aureus, P. aeruginosa and M. tuberculosis with MIC value of 16, 35, and 15.62 µg/ml respectively. The cytotoxicity evaluation of compound 1 on A549 cell lines showed it to be a safe antimicrobial molecule, TEM study suggested that the compound led to the rupture of the bacterial cell walls.