Isolation of two triterpenoids from Phlomis purpurea, one of them with anti-oomycete activity against Phytophthora cinnamomi, and insights into its biosynthetic pathway.

Phytophthora cinnamomi is an important plant pathogen responsible for dieback diseases in plant genera including Quercus, Fagus, Castanea, Eucalyptus, and Pinus, among others, all over the world. P. cinnamomi infection exerts tremendous ecological and economic losses. Several strategies have been developed to combat this pathogenic oomycete, including the search for novel anti-oomycete compounds. In this work, a Mediterranean vascular plant, Phlomis purpurea, has been screened for secondary bioactivity against this pathogen. The genus Phlomis includes a group of herbaceous plants and shrubs described as producers of many different bioactive compounds, including several triterpenoids. Triterpenoids are well-known molecules synthesized by plants and microorganisms with potent antioxidant, antitumoral, and antimicrobial activities. We have isolated by HPLC-DAD and characterized by HPLC-MS and NMR two nortriterpenoid compounds (phlomispentaol A and phlomispurtetraolone) from the root extracts of P. purpurea. One of them (phlomispentaol A) is active against the plant pathogenic oomycete P. cinnamomi (based on in vitro inhibition bioassays). Based on their chemical structure and their relationship to other plant triterpenoids, oleanolic acid is proposed to be the common precursor for these molecules. The anti-oomycete activity shown by phlomispentaol A represents a promising alternative to counteract the worldwide-scale damage caused to forest ecosystems by this pathogen.

A diet based on cured acorn-fed ham with oleic acid content promotes anti-inflammatory gut microbiota and prevents ulcerative colitis in an animal model.

BACKGROUND: Diets based on meat products are not recommended in the case of ulcerative colitis (UC). The objective here is to test if some traditional cured meat products, as acorn-fed ham (high levels of oleic acid), may be useful for controlling inflammatory diseases as UC in animal models, which could represent a new dietary complementary intervention in the prevention of this inflammatory disease in humans. METHODS: Two rat cohorts have been used: conventional vegetable rat feed and acorn-fed ham. UC was induced with DSS in drinking water ad libitum for 1 week. Short-chain fatty acids (SCFAs) and 16S rRNA metagenomics from bacterial populations were analyzed in cecum samples. Colon samples were analyzed for histological parameters. RESULTS: Acorn-fed ham diet induced changes in gut microbiota composition, with pronounced enrichments in anti-inflammatory bacterial genera (Alistipes, Blautia, Dorea, Parabacteroides). The animals with this diet showed a strong reduction in most parameters associated to ulcerative colitis: disease activity index, macroscopic score of colitis, epitelium alteration in colon mucosa, inflammatory cell density in colon, myeloperoxidase titers in colon, proinflammatory cytokines (IL-17, IFN-γ). Also, acorn-fed ham diet animals showed increased total antioxidant activity an oleic acid levels in plasma, as well as higher short-chain fatty acid concentrations in cecum (isobutyric, isovaleric and valeric). CONCLUSIONS: In the acorn-fed ham cohort, as a result of the dietary intake of oleic acid and low intake of omega-6 fatty acids, a strong preventive effect against UC symptoms was observed.

Mapping of the mouse 86-kDa heat-shock protein expressed gene (Hsp86-1) on chromosome 12 and related genes on chromosomes 3, 4, 9, and 11.

The HSP86 gene family in BALB/c, AKR/J, C58/J, and NFS/N inbred mice comprises an intron-containing expressed gene and, depending on the strain, two to four other HSP86-related members that are apparently processed pseudogenes. The expressed gene locus, Hsp86-1, was identified by its sequence identity with the mouse HSP86 cDNA coding region together with the presence of an intron at the same position as in the homologous human gene. Hsp86-1 was mapped 11.6 cM from the immunoglobulin heavy chain gene IgH on Chromosome 12 using an intersubspecies backcross. Two of the other loci that were common to all inbred strains tested, designated Hsp86-ps1 and Hsp86-ps2, were mapped to positions on Chromosomes 11 and 3, respectively. An HSP86-related locus specific to NFS/N and C58/J mice, designated Hsp86-ps3, was mapped on Chromosome 9. Also, an HSP86-related locus that was unique to NFS/N mice, designated Hsp86-ps4, was mapped to Chromosome 4.

Formation of an adduct between fosfomycin and glutathione: a new mechanism of antibiotic resistance in bacteria.

Plasmid-borne resistance to fosfomycin in bacteria is due to modification of the antibiotic molecule by a glutathione S-transferase that catalyzes the formation of a covalent bond between the sulfhydryl residue of the cysteine in glutathione and the C-1 of fosfomycin. This reaction results in opening of the epoxide ring of the antibiotic to form an inactive adduct, the structure of which was confirmed by nuclear magnetic resonance. Dialyzed extracts prepared from resistant Escherichia coli strains were unable to modify fosfomycin unless exogenous glutathione was added to the reaction mixtures. Similarly, mutants defective in glutathione biosynthesis were susceptible to fosfomycin, despite harboring a resistance plasmid. Extracts of resistant but not susceptible strains could join glutathione to 1-chloro-2,4-dinitrobenzene, confirming the nature of the enzymatic activity. Adduct formation appeared to be specific for glutathione: none of the other thiols tested (cysteine, N-acetylcysteine, and dithiothreitol) could modify fosfomycin.

Cloning and molecular epidemiology of plasmid-determined fosfomycin resistance.

The plasmid determinant of resistance to fosfomycin (For) was cloned into pBR322 and located in a 0.7-kilobase segment of DNA by transposon mutagenesis and in vitro deletion analysis. It encodes an 18-kilodalton protein located in the cytoplasm of resistant cells. Its synthesis is constitutive. The For genetic determinant is common to all plasmids isolated since 1975 in an hospital environment as determined by DNA-DNA hybridization. However, plasmids which carry For can be divided into two groups on the basis of size, pattern of antibiotic resistances, incompatibility specificity, and restriction and hybridization properties.

Plasmid-mediated fosfomycin resistance is due to enzymatic modification of the antibiotic.

The molecular mechanism of plasmid-mediated resistance to fosfomycin is described. The antibiotic was inactivated intracellularly and remained inside the cells. Modification was also obtained from cell extracts and was not energy dependent. The modifying enzyme seems to have sulfhydryl groups in its active center.

[Prevalence and dispersion of plasmids conferring fosfomycin resistance in enterobacteria]. / Prédominance et dispersion des plasmides conférant la résistance á la fosfomycine chez des entérobactéries.

The mechanisms of resistance to fosfomycin which, at the present, predominate among clinical isolates of Serratia marcescens as well as the incidence and dispersion of For plasmids among other species of Enterobacteriaceae were studied. It was found that that 23 of the 29 strains under study phenotypically behaved as glp T- mutants, another as glp T- and uhp- and five of them showed no alterations in their transport system. Self-transferable plasmids involved in For were isolated from the latter. For plasmids were also found in strains of Klebsiella pneumoniae but not in other enterobacteria. Plasmids were studied in order to establish their resistance phenotype, molecular weight and incompatibility group. The data obtained along with the restriction pattern allow us to conclude that the trait For is linked to, at least, two different replicons.