The Effect of Iodine-Containing Nano-Micelles, FS-1, on Antibiotic Resistance, Gene Expression and Epigenetic Modifications in the Genome of Multidrug Resistant MRSA Strain Staphylococcus aureus ATCC BAA-39.
ABSTRACT: Application of supplementary drugs which increase susceptibility of pathogenic bacteria to antibiotics is a promising yet unexplored approach to overcome the global problem of multidrug-resistant infections. The discovery of a new drug, an iodine-containing nano-molecular complex FS-1, which has proven to improve susceptibility to antibiotics in various pathogens, including MRSA strain Staphylococcus aureus ATCC BAA-39TM, allowed studying this phenomenon. Chromosomal DNA and total RNA samples extracted from the FS-1 treated strain (FS) and from the negative control (NC) cultures were sequenced by PacBio SMRT and Ion Torrent technologies, respectively. PacBio DNA reads were used to assemble chromosomal DNA of the NC and FS variants of S. aureus BAA-39 and to perform profiling of epigenetically modified nucleotides. Results of transcriptional profiling, variant calling and detection of epigenetic modifications in the FS variant were compared to the NC variant. Additionally, the genetic alterations caused by the treatment of S. aureus BAA-39 with FS-1 were compared to the results of a similar experiment conducted with another model organism, E. coli ATCC BAA-196. Several commonalities in responses of these phylogenetically distant microorganisms to the treatment with FS-1 were discovered, which included metabolic transition toward anaerobiosis and oxidative/osmotic stress response. S. aureus culture appeared to be more sensitive to FS-1 due to a higher penetrability of cells by iodine bound compounds, which caused carbonyl stress associated with nucleotide damaging by FS-1, abnormal epigenetic modifications and an increased rate of mutations. It was hypothesized that the disrupted pattern of adenine methylated loci within methicillin-resistance chromosome cassettes (SCCmec) may promote excision of this antibiotic resistance determinant from chromosomes while the altered pattern of cytosine methylation was behind the adaptive gene regulation in the culture FS. The selection against the antibiotic resistance in bacterial populations caused by abnormal epigenetic modifications exemplifies possible mechanisms of antibiotic resistance reversion induced by iodine-containing compounds. These finding will facilitate development of therapeutic agents against multidrug-resistant infections.
Project description:Staphylococcus aureus ATCC BAA-39 is the reference organism for a multidrug-resistant Staphylococcus aureus (MRSA) strain that was used to study drug-induced resistance reversion by an iodine-containing nanomolecular complex, FS-1. PacBio sequencing was performed on both the experimental and control strains, followed by genome assembly, variant calling, and DNA modification profiling.
Project description:Here, we report the complete genome sequence of the multidrug-resistant <i>Escherichia</i> <i>coli</i> strain ATCC BAA-196, a model organism used for studying possible antibiotic resistance reversion induced by FS-1, an iodine-containing complex. Two genomes, representing FS-1-treated and negative-control variants and composed of a chromosome and several plasmids, were assembled.
Project description:The strain Acinetobacter baumannii ATCC BAA-1790 was sequenced as a model for nosocomial multidrug-resistant infections. Long-read PacBio sequencing revealed a circular chromosome of 3,963,235?bp with two horizontally transferred genomic islands and a 67,023-bp plasmid. Multiple antibiotic resistance genes and genome methylation patterns were identified.
Project description:In this study, a drug discovery programme that sought to identify novel dual bacterial topoisomerase II inhibitors (NBTIs) led to the selection of six optimized compounds. In enzymatic assays, the molecules showed equivalent dual-targeting activity against the DNA gyrase and topoisomerase IV enzymes of Staphylococcus aureus and Escherichia coli. Consistently, the compounds demonstrated potent activity in susceptibility tests against various Gram-positive and Gram-negative reference species, including ciprofloxacin-resistant strains. The activity of the compounds against clinical multidrug-resistant isolates of S. aureus, Clostridium difficile, Acinetobacter baumannii, Neisseria gonorrhoeae, E. coli and vancomycin-resistant Enterococcus spp. was also confirmed. Two compounds (1 and 2) were tested in time-kill and post-antibiotic effect (PAE) assays. Compound 1 was bactericidal against all tested reference strains and showed higher activity than ciprofloxacin, and compound 2 showed a prolonged PAE, even against the ciprofloxacin-resistant S. aureus BAA-1720 strain. Spontaneous development of resistance to both compounds was selected for in S. aureus at frequencies comparable to those obtained for quinolones and other NBTIs. S. aureus BAA-1720 mutants resistant to compounds 1 and 2 had single point mutations in gyrA or gyrB outside of the quinolone resistance-determining region (QRDR), confirming the distinct site of action of these NBTIs compared to that of quinolones. Overall, the very good antibacterial activity of the compounds and their optimizable in vitro safety and physicochemical profile may have relevant implications for the development of new broad-spectrum antibiotics.
Project description:The rise of antibiotic resistance (ABR) and the drying up of the pipeline for the development of new antibiotics demands an urgent search for new antibiotic leads. While the majority of clinically available antibiotics were discovered from terrestrial Streptomyces, related species from marine sediments as a source of antibiotics remain underexplored. Here, we utilized culture-dependent isolation of thirty-five marine sediment-derived actinobacterial isolates followed by a screening of their antibacterial activity against multidrug-resistant S. aureus ATCC BAA-44. Our results revealed that the crude extract of Streptomyces griseorubens strain DSD069 isolated from marine sediments collected in Romblon, Philippines displays the highest antibacterial activity, with 96.4% growth inhibition. The S. aureus ATCC BAA-44 cells treated with crude extract of Streptomyces griseorubens strain DSD069 showed cell membrane damage as demonstrated by (a) leakage and loss of vital cell constituents, including DNA and proteins, (b) irregular shrinkage of cells, and (c) increase membrane permeability. The antibiotic compounds were identified as Bisanhydroaklavinone and 1-Hydroxybisanhydroaklavinone with MIC value of 6.25 ?g/mL and 50.00 ?g/mL, respectively. Bisanhydroaklavinone and 1-Hydroxybisanhydroaklavinone are shunt metabolites in the biosynthesis of anticancer anthracycline derivatives namely doxorubicin, daunorubicin, and cinerubins. It is rare, however, that shunt metabolites are accumulated during fermentation of marine sediment-derived Streptomyces strain without genetic modification. Thus, our study provides evidence that natural bacterial strain can produce Bisanhydroaklavinone and 1-Hydroxybisanhydroaklavinone as antibiotic leads to combat ABR.
Project description:BACKGROUND:Evaluating the antibiotic susceptibility and resistance genes is essential in the clinical management of bloodstream infections (BSIs). Nevertheless, there are still limited studies in Northern Vietnam. This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International License (CC BY-NC 4.0). AIM:This study aimed to determine the antibiotic resistance profile and methicillin-resistant encoding genes of Staphylococcus aureus (S. aureus) causing BSIs in Northern Vietnam. METHODS:The cross-sectional study was done from December 2012 to June 2014 in two tertiary hospitals in Northern Vietnam. Tests performed at the lab of the hospital. RESULTS:In 43 S. aureus strains isolating, 53.5 % were MRSA. Distribution of gene for overall, MRSA, and MSSA strains were following mecA gene (58.1 %; 95.7%, and 15%), femA gene (48.8%, 47.8%, and 50%), femB gene (88.4%, 82.6%, and 95%). Antibiotic resistance was highest in penicillin (100%), followed by erythromycin (65.1%) and clindamycin (60.5%). Several antibiotics were susceptible (100%), including vancomycin, tigecycline, linezolid, quinupristin/dalfopristin. Quinolone group was highly sensitive, include ciprofloxacin (83.7%), levofloxacin (86%) and moxifloxacin (86%). CONCLUSION:In S. aureus causing BSIs, antibiotic resistance was higher in penicillin, erythromycin, and clindamycin. All strains were utterly susceptible to vancomycin, tigecycline, linezolid, quinupristin/dalfopristin.
Project description:OBJECTIVE:Panton-Valentine leukocidin (PVL)-positive methicillin-resistant Staphylococcus aureus (MRSA) is a healthcare problem worldwide. There are no reports on the virulence characteristics of MRSA in Northern Cyprus (NC). This study aimed to determine the presence of pvl among MRSA isolates from patients admitted to a university hospital in NC using molecular methods. Fifty S. aureus strains were included in this study. BD Phoenix automated identification system was used for bacterial identification and antibiotic susceptibility testing. Methicillin resistance was confirmed by disc diffusion assay. Presence of nuc and mecA genes was tested by multiplex PCR. Detection of pvl gene was performed by single-target PCR. RESULTS:Out of 50 S. aureus isolates identified as MRSA by BD Phoenix system, 3 were susceptible to cefoxitin with disc diffusion assay and were confirmed as methicillin-sensitive S. aureus (MSSA). All isolates (n?=?50, 100%) tested positive for the presence nuc gene and 68% (n?=?34/50) were mecA positive. pvl was detected in 27.7% (n?=?13/47) of the MRSA isolates. Among PVL-positive MRSA isolates, 69.2% (9/13) were inpatients. PVL-MRSA was more common in isolates from deep tracheal aspirate (30.8%, 4/13) and abscess/wound (23.1%, 3/13). This represents the first study of PVL presence among MRSA in hospital setting in NC.
Project description:Staphylococcus aureus is a notorious human bacterial pathogen with considerable capacity to develop antibiotic resistance. We have observed that human infections caused by highly drug-resistant S. aureus are more prolonged, complicated, and difficult to eradicate. Here we describe a metabolic adaptation strategy used by clinical S. aureus strains that leads to resistance to the last-line antibiotic, daptomycin, and simultaneously affects host innate immunity. This response was characterized by a change in anionic membrane phospholipid composition induced by point mutations in the phospholipid biosynthesis gene, cls2, encoding cardiolipin synthase. Single cls2 point mutations were sufficient for daptomycin resistance, antibiotic treatment failure, and persistent infection. These phenotypes were mediated by enhanced cardiolipin biosynthesis, leading to increased bacterial membrane cardiolipin and reduced phosphatidylglycerol. The changes in membrane phospholipid profile led to modifications in membrane structure that impaired daptomycin penetration and membrane disruption. The cls2 point mutations also allowed S. aureus to evade neutrophil chemotaxis, mediated by the reduction in bacterial membrane phosphatidylglycerol, a previously undescribed bacterial-driven chemoattractant. Together, these data illustrate a metabolic strategy used by S. aureus to circumvent antibiotic and immune attack and provide crucial insights into membrane-based therapeutic targeting of this troublesome pathogen.
Project description:Bovine mastitis is a worldwide disease of dairy cattle associated with significant economic losses for the dairy industry. One of the most common pathogens responsible for mastitis is Staphylococcus (S.) aureus. Due to the development and spreading of antibiotic resistance, the search for novel antimicrobial substances against S. aureus is of great importance. The aim of this study was to evaluate two dihydroxybenzaldehydes for the prevention of bovine mastitis. Therefore we determined the minimal inhibitory concentration (MICs) of gentisaldehyde (2,5-dihydroxybenzaldehyde) and 2,3-dihydroxybenzaldehyde of a diverse set of 172 bovine mastitis S. aureus isolates using an automated robot-based microdilution method. To characterize the bovine isolates we determined the genotype by spa-typing, the antimicrobial resistance to eight antibiotic classes using the disk diffusion method and the MICs of three commonly used antiseptics (benzalkonium chloride, chlorhexidine, and iodine). Further we investigated the cytotoxicity of gentisaldehyde and 2,3-dihydroxybenzaldehyde in bovine mammary epithelial MAC-T cells using the XTT assay. The S. aureus strains showed a high genetic diversity with 52 different spa-types, including five novel types. Antibiotic susceptibility testing revealed that 24% of isolates were resistant to one antimicrobial agent and 3% of isolates were multi-resistant. The occurrence of antibiotic resistance strongly correlated with the spa-type. Both dihydroxybenzaldehydes showed antimicrobial activities with a MIC50 of 500 mg/L. The MIC of gentisaldehyde significantly correlated with that of 2,3-dihydroxybenzaldehyde, whereas no correlation was observed with the MIC of the three antiseptics. Cytotoxicity testing using bovine mammary epithelial MAC-T cells revealed that gentisaldehyde and 2,3-dihydroxybenzaldehyde show low toxicity at MIC50 and MIC90 concentrations. In conclusion, gentisaldehyde and 2,3-dihydroxybenzaldehyde exhibited antimicrobial activities against a diverse range of bovine mastitis S. aureus strains at low-cytotoxic concentrations. Therefore, both compounds are potential candidates as antiseptics to prevent bovine mastitis and to reduce the use of antibiotics in dairy cows.
Project description:This study aims to detect Staphylococcus aureus (S. aureus) resistance in the veterinary hospital environment. S. aureus are one of the components of the microbiota, and they may be present in patients in a veterinary hospital environment. Methicillin resistance is determined by a chromosomal gene (mecA), which codes for modifications in the beta-lactam antibiotic receptor, where the penicillin-binding protein will have a low affinity for the antibiotic. Samples were collected through swabs of materials and equipment at the hospital. S. aureus was identified in 7.6% (21/276) of the samples collected, and of the 21 strains isolated, 4 (19.0%) carried the mecA gene. MRSA are all strains of S. aureus that express the mecA gene. Four strains harbor the mecA gene; however, only two expressed the phenotypic resistance to cefoxitin and were characterized as MRSA. An isolate (strain 18) present on a patient care table was identified as methicillin-resistant S. aureus with intermediate sensitivity to vancomycin (VISA). Our observations suggest the need for containment measures (good antisepsis practices) to avoid the possible transmission of resistant bacterial agents for the veterinary hospital environment.