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Experimental evolution in morbidostat reveals converging genomic trajectories on the path to triclosan resistance.
Leyn, SA, Zlamal, JE, Kurnasov, OV, Li, X, Elane, M, Myjak, L, Godzik, M, de Crecy, A, Garcia-Alcalde, F, Ebeling, M, et al
Microbial genomics. 2021;(5)
Abstract
Understanding the dynamics and mechanisms of acquired drug resistance across major classes of antibiotics and bacterial pathogens is of critical importance for the optimization of current anti-infective therapies and the development of novel ones. To systematically address this challenge, we developed a workflow combining experimental evolution in a morbidostat continuous culturing device with deep genomic sequencing of population samples collected in time series. This approach was applied to the experimental evolution of six populations of Escherichia coli BW25113 towards acquiring resistance to triclosan (TCS), an antibacterial agent in various consumer products. This study revealed the rapid emergence and expansion (up to 100% in each culture within 4 days) of missense mutations in the fabI gene, encoding enoyl-acyl carrier protein reductase, the known TCS molecular target. A follow-up analysis of isolated clones showed that distinct amino acid substitutions increased the drug IC90 in a 3-16-fold range, reflecting their proximity to the TCS-binding site. In contrast to other antibiotics, efflux-upregulating mutations occurred only rarely and with low abundance. Mutations in several other genes were detected at an earlier stage of evolution. Most notably, three distinct amino acid substitutions were mapped in the C-terminal periplasmic domain of CadC protein, an acid stress-responsive transcriptional regulator. While these mutations do not confer robust TCS resistance, they appear to play a certain, yet unknown, role in adaptation to relatively low drug pressure. Overall, the observed evolutionary trajectories suggest that the FabI enzyme is the sole target of TCS (at least up to the ~50 µm level), and amino acid substitutions in the TCS-binding site represent the main mechanism of robust TCS resistance in E. coli. This model study illustrates the potential utility of the established morbidostat-based approach for uncovering resistance mechanisms and target identification for novel drug candidates with yet unknown mechanisms of action.
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2.
Toxicity and biochemical action of the antibiotic fungicide tetramycin on Colletotrichum scovillei.
Gao, Y, He, L, Li, X, Lin, J, Mu, W, Liu, F
Pesticide biochemistry and physiology. 2018;:51-58
Abstract
Tetramycin, a novel polyene macrolide antibiotic, has strong activity against a broad spectrum of fungi and may have potential uses in future agricultural applications. Thus, the antifungal activity and biochemical action of tetramycin on Colletotrichum scovillei were investigated in this study. The experimental results indicated that tetramycin had strong inhibitory activity against the mycelial growth, spore germination and germ tube elongation of C. scovillei. The baseline sensitivity curves were unimodal, with mean EC50 values of 1.98 ± 0.078 μg/mL and 0.003 ± 0.005 μg/mL for mycelial growth and spore germination inhibition, respectively. Tetramycin also inhibited the germination of spores and formation of appressoria. After tetramycin treatment, the edge of the mycelial diaphragm showed protuberances, with decreased offshoots at the top. Additionally, disruption of the membrane was detected through an increase in membrane permeability, leakage of sugars and a reduction in the ergosterol content. Tetramycin effectively controlled C. scovillei on detached pepper fruits. These results will contribute to our evaluation of the potential of tetramycin for successful management of pepper anthracnose and to our understanding of the possible biochemical action of tetramycin against C. scovillei.
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3.
Magnetic biochar-based manganese oxide composite for enhanced fluoroquinolone antibiotic removal from water.
Li, R, Wang, Z, Zhao, X, Li, X, Xie, X
Environmental science and pollution research international. 2018;(31):31136-31148
Abstract
Magnetic biochar-based manganese oxide composite (MMB) and raw biochar (BC) were synthesized via pyrolysis at a temperature of 500 °C under anoxic conditions of potato stems and leaves, characterized, and successfully used for the removal of norfloxacin (NOR), ciprofloxacin (CIP), and enrofloxacin (ENR) as representative compounds of fluoroquinolone antibiotics (FQs). Characterization results suggested that Fe3O4 and MnOx are the dominant crystals in MMB. MMB possessed large surface area and pore volume than BC. Batch adsorption experiments showed that the maximum adsorption abilities of MMB for norfloxacin (NOR), ciprofloxacin (CIP), and enrofloxacin (ENR) were 6.94, 8.37, and 7.19 mg g-1. In comparison to BC, the adsorption abilities of MMB increased 1.2, 1.5, and 1.6 times for NOR, CIP, and ENR, respectively. The pseudo-second-order kinetic model and the Langmuir model correlated satisfactorily to the experimental data. Thermodynamic studies revealed that the adsorption processes were spontaneous and endothermic. The adsorption capacity of MMB decreased with increasing solution pH (between 3.0 and 10.0) and increasing ionic strength (0.001-0.1). The MMB with high FQ removal efficiency, easy separation, and desirable regeneration ability may have promising environmental applications for the removal of fluoroquinolone antibiotics from water environment.
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4.
A systems biology approach to investigate the antimicrobial activity of oleuropein.
Li, X, Liu, Y, Jia, Q, LaMacchia, V, O'Donoghue, K, Huang, Z
Journal of industrial microbiology & biotechnology. 2016;(12):1705-1717
Abstract
Oleuropein and its hydrolysis products are olive phenolic compounds that have antimicrobial effects on a variety of pathogens, with the potential to be utilized in food and pharmaceutical products. While the existing research is mainly focused on individual genes or enzymes that are regulated by oleuropein for antimicrobial activities, little work has been done to integrate intracellular genes, enzymes and metabolic reactions for a systematic investigation of antimicrobial mechanism of oleuropein. In this study, the first genome-scale modeling method was developed to predict the system-level changes of intracellular metabolism triggered by oleuropein in Staphylococcus aureus, a common food-borne pathogen. To simulate the antimicrobial effect, an existing S. aureus genome-scale metabolic model was extended by adding the missing nitric oxide reactions, and exchange rates of potassium, phosphate and glutamate were adjusted in the model as suggested by previous research to mimic the stress imposed by oleuropein on S. aureus. The developed modeling approach was able to match S. aureus growth rates with experimental data for five oleuropein concentrations. The reactions with large flux change were identified and the enzymes of fifteen of these reactions were validated by existing research for their important roles in oleuropein metabolism. When compared with experimental data, the up/down gene regulations of 80% of these enzymes were correctly predicted by our modeling approach. This study indicates that the genome-scale modeling approach provides a promising avenue for revealing the intracellular metabolism of oleuropein antimicrobial properties.