Phosphine is a small redox-active gas that is used to protect global grain reserves, which are threatened by the emergence of phosphine resistance in pest insects. We find that polymorphisms responsible for genetic resistance cluster around the redox-active catalytic disulfide or the dimerization interface of dihydrolipoamide dehydrogenase (DLD) in insects (Rhyzopertha dominica and Tribolium castaneum) and nematodes (Caenorhabditis elegans). DLD is a core metabolic enzyme representing a new class of resistance factor for a redox-active metabolic toxin. It participates in four key steps of core metabolism, and metabolite profiles indicate that phosphine exposure in mutant and wild-type animals affects these steps differently. Mutation of DLD in C. elegans increases arsenite sensitivity. This specific vulnerability may be exploited to control phosphine-resistant insects and safeguard food security.
Biofilm is a natural form of bacterial growth ubiquitously in environmental niches. The biofilm formation results in increased resistance to negative environmental influences including resistance to antibiotics and antimicrobial agents. Quorum sensing (QS) is cell-to-cell communication mechanism, which plays an important role in biofilm development and balances the environment when the bacteria density becomes high. Due to the prominent points of biofilms implicated in infectious disease and the spread of multi-drug resistance, it is urgent to discover new antibacterial agents that can regulate biofilm formation and development. Accumulated evidences demonstrated that natural products from plants had antimicrobial and chemo-preventive properties in modulation of biofilm formation in the last two decades. This review will summarize recent studies on the discovery of natural anti-biofilm agents from plants with clear-cut mechanisms or identified molecular addresses, as well as some herbs with unknown mechanisms or unidentified bioactive ingredients. We also focus on the progression of techniques on the extraction and identification of natural anti-biofilm substances. Besides, anti-biofilm therapeutics undergoing clinical trials are discussed. These newly discovered natural anti-biofilm agents are promising candidates which could provide novel strategies for biofilm-associated infections.
Phosphine (hydrogen phosphide, PH3) is the fumigant most widely used to protect stored products from pest infestation. Despite the importance of this chemical, little is known about its mode of action. We have created three phosphine-resistant lines (pre-1, pre-7, pre-33) in the model organism C. elegans, with LC50 values 2, 5, and 9 times greater than the fully susceptible parental strain. Molecular oxygen was shown to be an extremely effective synergist with phosphine as, under hyperoxic conditions, 100% mortality was observed in wild-type nematodes exposed to 0.1 mg/l phosphine, a nonlethal concentration in air. All three mutants were resistant to the synergistic effects of oxygen in proportion to their resistance to phosphine with one mutant, pre-33, showing complete resistance to this synergism. We take the proportionality of cross-resistance between phosphine and the synergistic effect of oxygen to imply that all three mutants circumvent a mechanism of phosphine toxicity that is directly coupled to oxygen metabolism. Compared with the wild-type strain, all three mutants have an extended average life expectancy of from 12.5 to 25.3%. This is consistent with the proposed involvement of oxidative stress in both phosphine toxicity and ageing. Because the wild-type and mutant nematodes develop at the same rate, the longevity is unlikely to be caused by a clk-type reduction in oxidative metabolism, a potential alternative mechanism of phosphine resistance.
Streptococcus pyogenes (Group A Streptococcus or GAS) is a Gram-positive bacterial pathogen that has shown complex modes of regulation of its virulence factors to cause diverse diseases. Bacterial small RNAs are regarded as novel widespread regulators of gene expression in response to environmental signals. Recent studies have revealed that several small RNAs (sRNAs) have an important role in S. pyogenes physiology and pathogenesis by regulating gene expression at the translational level. To search for new sRNAs in S. pyogenes, we performed a genomewide analysis through computational prediction followed by experimental verification. To overcome the limitation of low accuracy in computational prediction, we employed a combination of three different computational algorithms (sRNAPredict, eQRNA and RNAz). A total of 45 candidates were chosen based on the computational analysis, and their transcription was analyzed by reverse-transcriptase PCR and Northern blot. Through this process, we discovered 7 putative novel trans-acting sRNAs. Their abundance varied between different growth phases, suggesting that their expression is influenced by environmental or internal signals. Further, to screen target mRNAs of an sRNA, we employed differential RNA sequencing analysis. This study provides a significant resource for future study of small RNAs and their roles in physiology and pathogenesis of S. pyogenes.
BackgroundIn recent years, dozens of Arabidopsis and rice CCCH-type zinc finger genes have been functionally studied, many of which confer important traits, such as abiotic and biotic stress tolerance, delayed leaf senescence and improved plant architecture. Switchgrass (Panicum virgatum) is an important bioenergy crop. Identification of agronomically important genes and/or loci is an important step for switchgrass molecular breeding. Annotating switchgrass CCCH genes using translational genomics methods will help further the goal of understanding switchgrass genetics and creating improved varieties.ResultsTaking advantage of the publicly-available switchgrass genomic and transcriptomic databases, we carried out a comprehensive analysis of switchgrass CCCH genes (PvC3Hs). A total of 103 PvC3Hs were identified and divided into 21 clades according to phylogenetic analysis. Genes in the same clade shared similar gene structure and conserved motifs. Chromosomal location analysis showed that most of the duplicated PvC3H gene pairs are in homeologous chromosomes. Evolution analysis of 19 selected PvC3H pairs showed that 42.1% of them were under diversifying selection. Expression atlas of the 103 PvC3Hs in 21 different organs, tissues and developmental stages revealed genes with higher expression levels in lignified cells, vascular cells, or reproductive tissues/organs, suggesting the potential function of these genes in development. We also found that eight PvC3Hs in Clade-XIV were orthologous to ABA- or stress- responsive CCCH genes in Arabidopsis and rice with functions annotated. Promoter and qRT-PCR analyses of Clade-XIV PvC3Hs showed that these eight genes were all responsive to ABA and various stresses.ConclusionsGenome-wide analysis of PvC3Hs confirmed the recent allopolyploidization event of tetraploid switchgrass from two closely-related diploid progenitors. The short time window after the polyploidization event allowed the existence of a large number of PvC3H genes with a high positive selection pressure onto them. The homeologous pairs of PvC3Hs may contribute to the heterosis of switchgrass and its wide adaptation in different ecological niches. Phylogenetic and gene expression analyses provide informative clues for discovering PvC3H genes in some functional categories. Particularly, eight PvC3Hs in Clade-XIV were found involved in stress responses. This information provides a foundation for functional studies of these genes in the future.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1328-4) contains supplementary material, which is available to authorized users.
BackgroundPerennial ryegrass (Lolium perenne L.) is an important temperate grass used for turf and forage purposes. With the increasing accumulation of genomic and transcriptomic data of perennial ryegrass, an efficient protoplast and transient gene expression protocol is highly desirable for in vivo gene functional studies in its homologous system. ResultsIn this report, a highly efficient protoplast isolation (5.6 × 107 protoplasts per gram of leaf material) and transient expression (plasmid transformation efficiency at 55.2%) was developed and the detailed protocol presented. Using this protocol, the subcellular locations of two ryegrass proteins were visualized in chloroplasts and nuclei, respectively, and protein–protein interaction between two chlorophyll catabolic enzymes (LpNOL and LpNYC1) was recorded in its homologous system for the first time.ConclusionThis efficient protoplast isolation and transformation protocol is sufficient for studies on protein subcellular localization and protein–protein interaction, and shall be suitable for many other molecular biology applications where the mesophyll protoplast system is desirable in perennial ryegrass.
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