Enterococcal infections, especially bacteremia and endocarditis, are challenging to treat because E. faecium have acquired resistance to multiple classes of antimicrobials, including ampicillin, aminoglycosides, and glycopeptides. Thus, research on different modes of enterococcal pathogenicity is warranted. This study utilized a proteomic approach to identify MV-associated proteins of different nosocomial E. faecium strains representing four clinically relevant sequence types (STs), namely ST17, ST18, ST78, and ST192. The presented data suggest that E. faecium MVs are involved in virulence and antimicrobial resistance.
Background. Infections by microbes (viruses, bacteria, and fungi) and parasites can cause serious diseases in both humans and animals. Heavy use of antimicrobials has created selective pressure and caused resistance to currently available antibiotics, hence the need for finding new and better antibiotics. Natural products, especially from plants, are known for their medicinal properties, including antimicrobial and anthelmintic activities. Geoclimatic variation, together with diversity in ethnomedicinal traditions, has made the Himalayas of Nepal an invaluable repository of traditional medicinal plants. We studied antiviral, antibacterial, antifungal, and anthelmintic activities of medicinal plants, selected based upon ethnobotanical evidence. Methods. Ethanolic and methanolic extracts were tested (1) on a panel of microbes: two Gram-positive bacteria (Staphylococcus aureus and Listeria innocua), four Gram-negative bacteria (Escherichia coli, Pseudomonas aeruginosa, Salmonella enterica, and Shigella sonnei), and one fungal species: Candida albicans; (2) against three different viruses: yellow fever, chikungunya, and enterovirus; and (3) on the nematode Caenorhabditis elegans. Also, cytotoxicity was assessed on human hepatoma (Huh), rhabdosarcoma (RD), and Vero (VC) cell lines. Results. Of 18 plants studied, Ampelocissus tomentosa and Aleuritopteris anceps inhibited S. aureus (MIC 35 μg/mL and 649 μg/mL, respectively) and Pseudomonas aeruginosa (MIC 15 μg/mL and 38 μg/mL, respectively). Rhododendron arboreum and Adhatoda vasica inhibited S. enterica (MIC 285 μg/mL and 326 μg/mL, respectively). Kalanchoe pinnata, Ampelocissus tomentosa, and Paris polyphylla were active against chikungunya virus, and Clerodendrum serratum was active against yellow fever virus (EC50 15.9 μg/mL); Terminalia chebula was active against enterovirus (EC50 10.6 μg/mL). Ampelocissus tomentosa, Boenninghausenia albiflora, Dichrocephala integrifolia, and Kalanchoe pinnata significantly reduced C. elegans motility, comparable to levamisole. Conclusions. In countries like Nepal, with a high burden of infectious and parasitic diseases, and a current health system unable to combat the burden of diseases, evaluation of local plants as a treatment or potential source of drugs can help expand treatment options. Screening plants against a broad range of pathogens (bacteria, viruses, fungi, and parasites) will support bioprospecting in Nepal, which may eventually lead to new drug development.
At present, few organisms are known to and capable of naturally producing laccases and white rot fungi are one such group. In the present study, three fungal species, namely, Ganoderma lucidum-CDBT1, Ganoderma japonicum, and Lentinula edodes, isolated from their native habitat in Nepal were screened for laccase production, and G. lucidum-CDBT1 was found to express highest levels of enzyme (day 10 culture media showed 0.92 IU/mg total protein or 92 IU/mL laccase activity with ABTS as substrate). Lignin extracted from rice straw was used in Olga medium for laccase production and isolation from G. lucidum-CDBT1. Presence of lignin (5 g/L) and copper sulfate (30 μM) in the media increased the extracellular laccase content by 111% and 114%, respectively. The laccase enzyme produced by G. lucidum-CDBT1 was fractionated by ammonium sulfate and purified by DEAE Sepharose anion exchange chromatography. The purified enzyme was found to have a molecular mass of 43 kDa and exhibits optimal activity at pH 5.0 and 30°C. The isolated laccase was thermally stable for up to 70°C for 1 h and exhibited broad pH stability. The kinetic constants, K m, V max, and K cat, determined using 2,2′-azinobis-(-3-ethylbenzothiazoline-6-sulfonic acid) as substrate were found to be 110 μM, 36 μmol/min/mg, and 246 min−1, respectively. The isolated thermostable laccase will be used in future experiments for delignification process.
Bacterial extracellular vesicles (EVs) have a vital role in bacterial pathogenesis. However, to date, the small RNA-cargo of EVs released by the opportunistic pathogen Staphylococcus aureus has not been characterized. Here, we shed light on the association of small RNAs with EVs secreted by S. aureus MSSA476 cultured in iron-depleted bacteriologic media supplemented with a subinhibitory dosage of vancomycin to mimic infection condition. Confocal microscopy analysis on intact RNase-treated EVs indicated that RNA is associated with EV particles. Transcriptomic followed by bioinformatics analysis of EV-associated RNA revealed the presence of potential gene regulatory small RNAs and high levels of tRNAs. Among the EV-associated enriched small RNAs were SsrA, RsaC and RNAIII. Our finding invites new insights into the potential role of EV-associated RNA as a modulator of host-pathogen interaction.
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