Towards sustainable antimicrobials from plants: some ways to abridge current methodological approaches

Plants are potential sources for new antimicrobials that might be helpful in combating antimicrobial resistance. Screening of plant materials for antimicrobial activities is an essential step towards discovery of new alternatives to current antibiotics. This article presents strategies allowing to simplify some methodological approaches used in initial testing of plant materials for their antimicrobial properties. Particular attention is given to sensitivity testing without extraction based on utilisation of agarose hydrogel tablets and dry powders. Other aspects relate to testing fractionated extracts and rapid determination of inhibition modes directly from zones of inhibition in diffusion assays. Using these approaches could facilitate standardised preliminary screening for new antimicrobials to make this process more productive

Absence of photosynthetic state transitions in alien chloroplasts

MAIN CONCLUSION:The absence of state transitions in a Nt(Hn) cybrid is due to a cleavage of the threonine residue from the misprocessed N-terminus of the LHCII polypeptides. The cooperation between the nucleus and chloroplast genomes is essential for plant photosynthetic fitness. The rapid and specific interactions between nucleus-encoded and chloroplast-encoded proteins are under intense investigation with potential for applications in agriculture and renewable energy technology. Here, we present a novel model for photosynthesis research in which alien henbane (Hyoscyamus niger) chloroplasts function on the nuclear background of a tobacco (Nicotiana tabacum). The result of this coupling is a cytoplasmic hybrid (cybrid) with inhibited state transitions-a mechanism responsible for balancing energy absorption between photosystems. Protein analysis showed differences in the LHCII composition of the cybrid plants. SDS-PAGE analysis revealed a novel banding pattern in the cybrids with at least one additional 'LHCII' band compared to the wild-type parental species. Proteomic work suggested that the N-terminus of at least some of the cybrid Lhcb proteins was missing. These findings provide a mechanistic explanation for the lack of state transitions-the N-terminal truncation of the Lhcb proteins in the cybrid included the threonine residue that is phosphorylated/dephosphorylated in order to trigger state transitions and therefore crucial energy balancing mechanism in plants

Checkpoint-dependent phosphorylation of Exo1 modulates the DNA damage response

Exo1 is a nuclease involved in mismatch repair, DSB repair, stalled replication fork processing and in the DNA damage response triggered by dysfunctional telomeres. In budding yeast and mice, Exo1 creates single-stranded DNA (ssDNA) at uncapped telomeres. This ssDNA accumulation activates the checkpoint response resulting in cell cycle arrest. Here, we demonstrate that Exo1 is phosphorylated when telomeres are uncapped in cdc13-1 and yku70Δ yeast cells, and in response to the induction of DNA damage. After telomere uncapping, Exo1 phosphorylation depends on components of the checkpoint machinery such as Rad24, Rad17, Rad9, Rad53 and Mec1, but is largely independent of Chk1, Tel1 and Dun1. Serines S372, S567, S587 and S692 of Exo1 were identified as targets for phosphorylation. Furthermore, mutation of these Exo1 residues altered the DNA damage response to uncapped telomeres and camptothecin treatment, in a manner that suggests Exo1 phosphorylation inhibits its activity. We propose that Rad53-dependent Exo1 phosphorylation is involved in a negative feedback loop to limit ssDNA accumulation and DNA damage checkpoint activation

Co-operative inhibitory effects of hydrogen peroxide and iodine against bacterial and yeast species.

BACKGROUND: Hydrogen peroxide and iodine are powerful antimicrobials widely used as antiseptics and disinfectants. Their antimicrobial properties are known to be enhanced by combining them with other compounds. We studied co-operative inhibitory activities (synergism, additive effects and modes of growth inhibition) of hydrogen peroxide and iodine used concurrently against 3 bacterial and 16 yeast species. RESULTS: Synergistic or additive inhibitory effects were shown for hydrogen peroxide and iodine mixtures against all 19 species used in the study. Both biocides were mostly cidal individually and in mixtures against Pseudomonas aeruginosa and Staphylococcus aureus. Both compounds manifested static inhibitory effects individually, but their mixtures were synergistically cidal for Saccharomyces cerevisiae and Escherihia coli. Cells of S. cerevisiae treated with hydrogen peroxide and iodine-hydrogen peroxide mixture produced increased numbers of respiratory deficient mutants indicating genotoxic effects. CONCLUSION: Iodine and hydrogen peroxide used concurrently interact synergistically or additively against a range of prokaryotic and eukaryotic microorganisms. The study provides an insight as to how these traditional antimicrobials could be used more effectively for disinfection and antisepsis. In addition, a simple approach is proposed for scoring genotoxicity of different biocides by using the budding yeast system

Mitochondrial tuning fork in nuclear homeotic functions

Homeotic-like flower morphologies in plants with cytoplasmic male sterility (CMS) are maternally inherited and associated with rearrangements in mitochondrial DNA. Recent studies allow an interpretation of dramatic CMS morphologies in the light of the floral ABC model. They uncover new nuclear targets for interactions with mitochondrial genes. GLOBOSA-, DEFICIENS- and APETALA3-like genes were transcriptionally down-regulated in carpelloid CMS flowers of tobacco, carrot and wheat. These results allow cooperation between nuclear and cytoplasmic genetic compartments to be considered as a developmental function and an evolutionary mechanism of speciation

Linear chromosome maintenance in the absence of essential telomere-capping proteins

Belgium Herbarium image of Meise Botanic Garden

Self-fertile cybrids nicotiana tabacum (+hyoscyamus aureus) with a nucleo-plastome incompatibility

Cytoplasmic hybrids (cybrids) in a novel inter-generic combination, Nicotiana tabacum (+Hyoscyamus aureus), were generated by fusion of protoplasts from a plastome tobacco albino mutant (line R100a1) and %-irradiated green protoplasts of H. aureus. Cybrids possessed a plastome of H. aureus and a rearranged mitochondrial DNA. The cybrids displayed a syndrome of nucleo-plastome incompatibility expressed as a partial chlorophyll-deficiency of cotyledonary and true leaves at the early stage of vegetative development of plants grown from seeds in soil. During later development, the plants restored a normal green coloration. This character is phenotypically indistinguishable from the same syndrome in previously generated cybrids N. tabacum (+H. nigrum). In contrast to the cybrids N. tabacum (+H. nigrum), cybrids N. tabacum (+H. aureus) were self-fertile, and did not manifest other features that were interpreted as nucleo-mitochondrial incompatibilities in N. tabacum (+H. nigrum) plants. Therefore, the cybrids N. tabacum (+H. aureus) present a self-propagating system of Nicotiana (+Hyoscyamus) nucleo-plastome incompatibility in its pure form

Differential regulation of genes transcribed by nucleus-encoded plastid RNA polymerase, and DNA amplification, within ribosome-deficient plastids in stable phenocopies of cereal albino mutants

We isolated stable albino plants of barley and maize by inhibiting plastid protein synthesis with streptomycin and propagating bleached seedlings in the absence of antibiotics in vitro. Albino plants are deficient in plastid translation products and plastid ribosomal RNAs, and are stable phenocopies of the barley albostrians and maize iojap mutants, which contain ribosome-free plastids. Once plastid ribosomes are lost they cannot be re-synthesized because about one-third of plastid ribosomal proteins are themselves plastid encoded. The group II/subgroup IIA intron in plastid rpl2 transcripts was not processed in albinos, providing strong evidence for the absence of plastid translation. Photosynthesis-related plastid mRNAs and plastid tRNAs were down-regulated in albino leaves. A differential influence of plastid ribosome deficiency on mRNA levels allowed us to divide genes transcribed by nucleus-encoded plastid RNA polymerase into two groups. Northern analysis revealed increases in the levels of clpP, rpl2, rpl23, rps15 and rpoB mRNAs in total RNA from albino leaves relative to those in green leaves. In contrast, albinism did not increase the band intensities of rps2 and rps4 messages. Plastid ribosome-associated factor(s) or plastid-encoded product(s) play a role in the initiation, termination, processing or stability of transcripts containing trnG(UCC) and rps4. Excision and 100-fold amplification of a 5.2-kb region of plastid DNA encompassing the trnG(UCC) and trnE(UUC) genes was observed in one of four albino barley plants. Gene amplification was correlated with the accumulation of abundant novel transcripts derived from regions flanking the trnG(UCC) gene

Exo1 and Rad24 Differentially Regulate Generation of ssDNA at Telomeres of Saccharomyces cerevisiae cdc13-1 Mutants

Cell cycle arrest in response to DNA damage depends upon coordinated interactions between DNA repair and checkpoint pathways. Here we examine the role of DNA repair and checkpoint genes in responding to unprotected telomeres in budding yeast cdc13-1 mutants. We show that Exo1 is unique among the repair genes tested because like Rad9 and Rad24 checkpoint proteins, Exo1 inhibits the growth of cdc13-1 mutants at the semipermissive temperatures. In contrast Mre11, Rad50, Xrs2, and Rad27 contribute to the vitality of cdc13-1 strains grown at permissive temperatures, while Din7, Msh2, Nuc1, Rad2, Rad52, and Yen1 show no effect. Exo1 is not required for cell cycle arrest of cdc13-1 mutants at 36° but is required to maintain arrest. Exo1 affects but is not essential for the production of ssDNA in subtelomeric Y′ repeats of cdc13-1 mutants. However, Exo1 is critical for generating ssDNA in subtelomeric X repeats and internal single-copy sequences. Surprisingly, and in contrast to Rad24, Exo1 is not essential to generate ssDNA in X or single-copy sequences in cdc13-1 rad9Δ mutants. We conclude that Rad24 and Exo1 regulate nucleases with different properties at uncapped telomeres and propose a model to explain our findings