Putative Mycobacterial Efflux Inhibitors from the Seeds of <i>Aframomum melegueta</i>

In order to identify new putative efflux pump inhibitors that represent an appropriate target in antimycobacterial chemotherapy, nine paradol- and gingerol-related compounds (<b>1</b>–<b>9</b>) isolated from the seeds of A<i>framomum melegueta</i> were assessed for their potential to inhibit ethidium bromide (EtBr) efflux in a <i>Mycobacterium smegmatis</i> model. Five of the compounds from <i>A. melegueta</i> and NMR spectroscopic data of the diketone 6-gingerdione (<b>2</b>) and its enolic tautomers, methyl-6-gingerol (<b>5</b>) and <i>rac</i>-6-dihydroparadol (<b>7</b>), are presented herein for the first time. After determination of their antimycobacterial activities and modulatory effects on the MIC of antibiotics as well as their synergistic effects in combination with antibiotics against <i>M. smegmatis</i> mc² 155, their impact on EtBr accumulation and efflux was evaluated using a microtiter plate-based fluorometric assay. The compounds exhibited moderate to weak antimycobacterial activities, and the best modulators induced a 4- to 16-fold decrease of the MICs of EtBr and rifampicin as well as a reduction of the MIC of isoniazid with fractional inhibitory concentration index values indicating synergistic activities in some cases. 6-Paradol (<b>3</b>), 8-gingerol (<b>6</b>), and <i>rac-</i>6-dihydroparadol (<b>7</b>) were the most potent EtBr efflux inhibitors in <i>M. smegmatis</i> mc² 155, displaying EtBr efflux inhibiting activities comparable to reference inhibitors

Metabolic profiling of rhizomes of native populations of the strictly endemic Croatian species <i>Iris adriatica</i>

<p><i>Iris adriatica</i> Trinajstić ex Mitić (Iridaceae L.) is a strictly endemic taxon from Croatia. It is a rhizomatous dwarf plant from the <i>I. pumila</i> complex with a distribution area limited to the Croatian part of the Mediterranean area, mainly central Dalmatia. The genus <i>Iris</i> is known for its richness in isoflavonoids which also play a significant role in chemotaxonomy and biological activity. Hence, in the current study, different plant batches of <i>I. adriatica</i> collected in early spring of 2016 were analysed for their phytochemical profiles and qualitatively compared. UHPLC-PDA-ESI-MS analyses of methanolic rhizome extracts were performed. Altogether, 36 compounds, representing isoflavonoids (including 6,7-methylendioxy derivatives), benzophenones and xanthones were found as aglycones or in glycosidically bound form to be the main constituent groups of <i>I. adriatica</i> rhizomes. Qualitative results were identical between different batches of plant material from collection sites in central Dalmatia, they differed only in quantity. For some phenolic compounds of <i>I. adriatica</i>, chemotaxonomic relevance was detected.</p

Influence of (-)-α-pinene on EtBr accumulation in the 16 <i>C</i>. <i>jejuni</i> strains.

<p>Relationship between EtBr accumulation in untreated cultures and in cultures treated with 62.5 mg/L (-)-α-pinene. Each symbol represents a single isolate. Solid squares, antibiotic resistant strains (resistant to three or more unrelated antibiotics); open circles, antibiotic sensitive strains.</p

Scheme of the insertional mutagenesis strategies (not to scale).

<p>Box arrows, target open reading frames; solid arrows, primers used for DNA fragment amplification (from left to right: F1-F, F1-R, F2-F, F2-R, specific for each individual mutant, as listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122871#pone.0122871.s002" target="_blank">S2 Table</a>); blue dots, target gene; blue rectangles, deleted regions substituted with a kanamycin cassette (<i>kan</i>).</p

Functional categories of the differentially expressed genes in <i>Campylobacter jejuni</i>.

<p>Functional categories of the differentially expressed genes in <i>Campylobacter jejuni</i>.</p

Antimicrobial activity and modulation of antimicrobial resistance of (-)-α-pinene for the different strains of <i>Campylobacter jejuni</i>.

<p>MIC, minimal inhibitory concentration; AP, (-)-α-pinene; CIP, ciprofloxacin; ERY: erythromycin; TC, triclosan; EtBr, ethidium bromide;-AP, in the absence of (-)-α-pinene; +AP, with 125 mg/L (-)-α-pinene, MF, modulation factor.</p><p>* In this case +AP was applied in concentration 62.5 mg/L.</p><p>Antimicrobial activity and modulation of antimicrobial resistance of (-)-α-pinene for the different strains of <i>Campylobacter jejuni</i>.</p

Influence of (-)-α-pinene on membrane integrity of <i>C</i>. <i>jejuni</i> NCTC 11168.

<p>Membrane integrity of wild-type <i>C</i>. <i>jejuni</i> NCTC 11168 cultures, untreated culture, culture treated with 125 mg/L (-)-α-pinene or 62.5 mg/L (-)-α-pinene, heat treated culture and blank.</p

Influence of (-)-α-pinene, CCCP and reserpine on ethidium bromide accumulation.

<p>(A) Reference strain <i>C</i>. <i>jejuni</i> NCTC 11168, (B) <i>C</i>. <i>jejuni</i> NCTC 11168 ΔcmeB knock-out mutant, (C) <i>C</i>. <i>jejuni</i> NCTC 11168 Δ<i>Cj1687</i> knock-out mutant.</p