Synthetic Biology Tools for Engineering Microbial Cells to Fight Superbugs.

With the increase in clinical cases of bacterial infections with multiple antibiotic resistance, the world has entered a health crisis. Overuse, inappropriate prescribing, and lack of innovation of antibiotics have contributed to the surge of microorganisms that can overcome traditional antimicrobial treatments. In 2017, the World Health Organization published a list of pathogenic bacteria, including Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Escherichia coli (ESKAPE). These bacteria can adapt to multiple antibiotics and transfer their resistance to other organisms; therefore, studies to find new therapeutic strategies are needed. One of these strategies is synthetic biology geared toward developing new antimicrobial therapies. Synthetic biology is founded on a solid and well-established theoretical framework that provides tools for conceptualizing, designing, and constructing synthetic biological systems. Recent developments in synthetic biology provide tools for engineering synthetic control systems in microbial cells. Applying protein engineering, DNA synthesis, and in silico design allows building metabolic pathways and biological circuits to control cellular behavior. Thus, synthetic biology advances have permitted the construction of communication systems between microorganisms where exogenous molecules can control specific population behaviors, induce intracellular signaling, and establish co-dependent networks of microorganisms.

Re-sensitizing Ampicillin and Kanamycin-Resistant E. coli and S. aureus Using Synergistic Metal Micronutrients-Antibiotic Combinations.

Due to the recent emergence of multi-drug resistant strains, the development of novel antimicrobial agents has become a critical issue. The use of micronutrient transition metals is a promising approach to overcome this problem since these compounds exhibit significant toxicity at low concentrations in prokaryotic cells. In this work, we demonstrate that at concentrations lower than their minimal inhibitory concentrations and in combination with different antibiotics, it is possible to mitigate the barriers to employ metallic micronutrients as therapeutic agents. Here, we show that when administered as a combinatorial treatment, Cu2+, Zn2+, Co2+, Cd2+, and Ni2+ increase susceptibility of Escherichia coli and Staphylococcus aureus to ampicillin and kanamycin. Furthermore, ampicillin-resistant E. coli is re-sensitized to ampicillin when the ampicillin is administered in combination with Cu2+, Cd2+, or Ni2. Similarly, Cu2+, Zn2+, or Cd2+ re-sensitize kanamycin-resistant E. coli and S. aureus to kanamycin when administered in a combinatorial treatment with those transition metals. Here, we demonstrate that for both susceptible and resistant bacteria, transition-metal micronutrients, and antibiotics interact synergistically in combinatorial treatments and exhibit increased effects when compared to the treatment with the antibiotic alone. Moreover, in vitro and in vivo assays, using a murine topical infection model, showed no toxicological effects of either treatment at the administered concentrations. Lastly, we show that combinatorial treatments can clear a murine topical infection caused by an antibiotic-resistant strain. Altogether, these results suggest that antibiotic-metallic micronutrient combinatorial treatments will play an important role in future developments of antimicrobial agents and treatments against infections caused by both susceptible and resistant strains.

Antimicrobial activity of a silver-microfibrillated cellulose biocomposite against susceptible and resistant bacteria.

Antibiotic Microbial Resistance (AMR) is a major global challenge as it constitutes a severe threat to global public health if not addressed. To fight against AMR bacteria, new antimicrobial agents are continually needed, and their efficacy must be tested. Historically, many transition metals have been employed, but their cytotoxicity is an issue and hence must be reduced, typically by combination with organic polymers. Cellulose of natural origin, especially those derived from unavoidable residues in the food supply chain, appears to be a good capping agent for the green synthesis of silver nanoparticles. Herein, we describe a green synthesis method to produce a novel biocomposite, using ascorbic acid as reducing agent and microfibrillated cellulose as a capping agent and demonstrate this material to be an efficient antimicrobial agent. Silver nanoparticles were obtained in the cellulose matrix with an average size of 140 nm and with antimicrobial activity against both sensitive and resistant Gram positive (using 1500 ppm) as well as sensitive and resistant Gram negative (using 125 ppm) bacteria. Also, an inverted disk-diffusion methodology was applied to overcome the low-solubility of cellulose compounds. This novel silver nanoparticle-cellulose biocomposite synthesized by a green methodology shows the potential to be applied in the future development of biomedical instruments and therapeutics.

Antimicrobial and antibiofilm activity of biopolymer-Ni, Zn nanoparticle biocomposites synthesized using R. mucilaginosa UANL-001L exopolysaccharide as a capping agent.

Introduction: Global increase in the consumption of antibiotics has induced selective stress on wild-type microorganisms, pushing them to adapt to conditions of higher antibiotic concentrations, and thus an increased variety of resistant bacterial strains have emerged. Metal nanoparticles synthesized by green methods have been studied and proposed as potential antimicrobial agents against both wild-type and antibiotic-resistant strains; in addition, exopolysaccharides have been used as capping agent of metal nanoparticles due to their biocompatibility, reducing biological risks in a wide variety of applications. Purpose: In this work, we use an exopolysaccharide, from Rhodotorula mucilaginosa UANL-001L, an autochthonous strain from the Mexican northeast, as a capping agent in the synthesis of Zn, and Ni, nanoparticle biopolymer biocomposites. Materials and methods: To physically and chemically characterize the synthesized biocomposites, FT-IR, UV-Vs, TEM, SAED and EDS analysis were carried out. Antimicrobial and antibiofilm biological activity were tested for the biocomposites against two resistant clinical strains, a Gram-positive Staphylococcus aureus, and a Gram-negative Pseudomonas aeruginosa. Antimicrobial activity was determined using a microdilution assay whereas antibiofilm activity was analyzed through crystal violet staining. Results: Biocomposites composed of exopolysaccharide capped Zn and Ni metal nanoparticles were synthesized through a green synthesis methodology. The average size of the Zn and Ni nanoparticles ranged between 8 and 26 nm, respectively. The Ni-EPS biocomposites showed antimicrobial and antibiofilm activity against resistant strains of Staphylococcus aureus and Pseudomonas aeruginosa at 3 and 2 mg/mL, respectively. Moreover, Zn-EPS biocomposites showed antimicrobial activity against resistant Staphylococcus aureus at 1 mg/mL. Both biocomposites showed no toxicity, as renal function showed no differences between treatments and control in the in vivo assays with male rats tests in this study at a concentration of 24 mg/kg of body weight. Conclusion: The exopolysaccharide produced by Rhodotorula mucilaginosa UANL-001L is an excellent candidate as a capping agent in the synthesis of biopolymer-metal nanoparticle biocomposites. Both Ni and Zn-EPS biocomposites demonstrate to be potential contenders as novel antimicrobial agents against both Gram-negative and Gram-positive clinically relevant resistant bacterial strains. Moreover, Ni-EPS biocomposites also showed antibiofilm activity, which makes them an interesting material to be used in different applications to counterattack global health problems due to the emergence of resistant microorganisms.