In Vitro Activity, Stability and Molecular Characterization of Eight Potent Bacteriophages Infecting Carbapenem-Resistant Klebsiella pneumoniae.

BACKGROUND: Members of the genus Klebsiella are among the leading microbial pathogens associated with nosocomial infection. The increased incidence of antimicrobial resistance in these species has propelled the need for alternate/combination therapeutic regimens to aid clinical treatment, including bacteriophage therapy. Bacteriophages are considered very safe and effective in treating bacterial infections. In this study, we characterize eight lytic bacteriophages that were previously isolated by our team against carbapenem-resistant Klebsiella pneumoniae. METHODS: The one-step-growth curves, stability and lytic ability of eight bacteriophages were characterized. Restriction fragment length polymorphism (RFLP), random amplification of polymorphic DNA (RAPD) typing analysis and protein profiling were used to characterize the microbes at the molecular level. Phylogenetic trees of four important proteins were constructed for the two selected bacteriophages. RESULTS AND CONCLUSIONS: All eight bacteriophages showed high efficiency for reducing bacterial concentration with high stability under different physical and chemical conditions. We found four major protein bands out of at least ten 15-190 KDa bands that were clearly separated by SDS-PAGE, which were assumed to be the major head and tail proteins. The genomes were found to be dsDNA, with sizes of approximately 36-87 Kb. All bacteriophages reduced the optical density of the planktonic K. pneumoniae abruptly, indicating great potential to reduce K. pneumoniae infection. In this study, we have found that tail fiber protein can further distinguished closely related bacteriophages. The characterised bacteriophages showed promising potential as candidates against carbapenem-resistant Klebsiella pneumoniae via bacteriophage therapy.

Disruption of methicillin-resistant Staphylococcus aureus protein synthesis by tannins.

INTRODUCTION: Methicillin-resistant Staphylococcus aureus (MRSA) is a worldwide public health threat, displaying multiple antibiotic resistance that causes morbidity and mortality. Management of multidrug-resistant (MDR) MRSA infections is extremely difficult due to their inherent resistance to currently used antibiotics. New antibiotics are needed to combat the emergence of antimicrobial resistance. METHODS: The in vitro effect of tannins was studied against MRSA reference strain (ATCC 43300) and MRSA clinical strains utilizing antimicrobial assays in conjunction with both scanning and transmission electron microscopy. To reveal the influence of tannins in MRSA protein synthesis disruption, we utilized next-generation sequencing (NGS) to provide further insight into the novel protein synthesis transcriptional response of MRSA exposed to these compounds. RESULTS: Tannins possessed both bacteriostatic and bactericidal activity with minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of 0.78 and 1.56 mg/mL, respectively, against all tested MRSA. Scanning and transmission electron microscopy of MRSA treated with tannins showed decrease in cellular volume, indicating disruption of protein synthesis. CONCLUSION: Analysis of a genome-wide transcriptional profile of the reference strain ATCC 43300 MRSA in response to tannins has led to the finding that tannins induced significant modulation in essential ribosome pathways, which caused a reduction in the translation processes that lead to inhibition of protein synthesis and obviation of bacterial growth. These findings highlight the potential of tannins as new promising anti-MRSA agents in clinical application such as body wash and topical cream or ointments.

Transcriptome analysis of methicillin-resistant Staphylococcus aureus in response to stigmasterol and lupeol.

OBJECTIVES: Methicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen with multiple antibiotic resistance that causes morbidity and mortality worldwide. Multidrug-resistant (MDR) MRSA with increased resistance to currently available antibiotics has challenged the world to develop new therapeutic agents. Stigmasterol and lupeol, from the plant Phyllanthus columnaris, exhibit antibacterial activities against MRSA. The aim of this study was to utilise next-generation sequencing (NGS) to provide further insight into the novel transcriptional response of MRSA exposed to stigmasterol and lupeol. METHODS: Time-kill analysis of one MRSA reference strain (ATCC 43300) and three clinical isolates (WM3, BM1 and KJ7) for both compounds was first performed to provide the bacteriostatic/bactericidal profile. Then, MRSA ATCC 43300 strain treated with both compounds was interrogated by NGS. RESULTS: Both stigmasterol and lupeol possessed bacteriostatic properties against all MRSA tested; however, lupeol exhibited both bacteriostatic and bactericidal properties within the same minimum inhibitory concentration and minimum bactericidal concentration values against BM1 (12.5mg/mL). Transcriptome profiling of MRSA ATCC 43300 revealed significant modulation of gene expression with multiple desirable targets by both compounds, which caused a reduction in the translation processes leading to inhibition of protein synthesis and prevention of bacterial growth. CONCLUSIONS: This study highlights the potential of both stigmasterol and lupeol as new promising anti-MRSA agents.

Synthesis of silver nanoparticles with antibacterial activity using the lichen Parmotrema praesorediosum.

Development of a green chemistry process for the synthesis of silver nanoparticles has become a focus of interest. This would offer numerous benefits, including ecofriendliness and compatibility for biomedical applications. Here we report the synthesis of silver nanoparticles from the reduction of silver nitrate and an aqueous extract of the lichen Parmotrema praesorediosum as a reductant as well as a stabilizer. The physical appearance of these silver nanoparticles was characterized using ultraviolet-visible spectroscopy, electron microscopy, energy-dispersive spectroscopy, and X-ray diffraction techniques. The results show that silver nanoparticles synthesized using P. praesorediosum have an average particle size of 19 nm with a cubic structure. The antibacterial activity of the synthesized silver nanoparticles was tested against eight micro-organisms using the disk diffusion method. The results reveal that silver nanoparticles synthesized using P. praesorediosum have potential antibacterial activity against Gram-negative bacteria.