Scientists at the University of Nottingham have engineered a self-assembling, living material resembling human sputum, or phlegm, which supports the growth of microbial 3D biofilms. The new platform will help researchers better understand how infections develop in the lungs of patients with cystic fibrosis, and the effects of antibiotics.
“The capacity to create complex 3D biofilms in the lab in a simple manner will lead to practical tools to better understand how these living structures form and how to treat them better,” said research lead Yuanhao Wu, MD, PhD. “The technology developed in this study will revolutionise the way we study biofilm-mediated infections and assess the effectiveness of novel antimicrobials using different in vivo-like infection environments,” added Miguel Cámara, PhD, at the National Biofilms Innovation Centre in the School of Life Sciences at the University of Nottingham.
The researchers described their development of the living material platform in Matter, in a paper titled “Co-assembling living material as an in vitro lung epithelial infection model.” In their report the team concluded, “Our study offers a living material capable of growing functional 3D biofilms that simulate in vitro the nutritional and mechanical properties of these systems in vivo.” The study was carried out as a collaboration with Alvaro Mata, PhD, in the School of Pharmacy and Department of Chemical Engineering.
Biofilms are strong living 3D materials that play key roles in nature, but also cause major problems in the real world, such as in the contamination of hospital surfaces, or in our tolerance to antibiotic treatment, the authors noted. However, one of the biggest challenges in antimicrobial discovery is the lack of biofilm models that reflect the complexity of natural environments, such as those encountered in the lung of cystic fibrosis (CF) patients. “… growing 3D biofilms in vitro is difficult primarily because of the limitations in developing matrices that mimic the inherent structural and compositional complexity of their extracellular milieu,” the team further wrote.
Individuals with cystic fibrosis are unable to clear infections in their lungs, where complex communities of disease-causing microbes accumulate within thick mucus and form 3D biofilms. These natural biofilms are highly resilient to antibiotics and there is a critical need to develop in vitro models which can reliably reproduce them in the lab, so that experts can better understand their biology discover new ways to tackle the resulting issues. “Recreation of these living structures in vitro is critical to understand their biology and develop solutions to the problems they cause,” the team stated.
The availability of in vitro models will enable a more consistent evaluation of novel therapeutic interventions before being taking into preclinical studies. Such models will also be key to answering fundamental research questions about the interactions between polymicrobial biofilms and their host which lead to chronic infections.
Living materials consist of extracellular matrix materials (ECMMs) embedding living components, such as engineered cells and bacteria, which are able to modify the structure and properties of the ECMMs. For their reported study the researchers engineered a living material resembling natural sputum from CF patients, which could support and grow 3D polymicrobial biofilms in a controlled manner, resembling those found in the CF lung.
The team was able to achieve this by combining peptides with an artificial culture medium that is known to recreate natural sputum and which can be easily infected. The living material incorporates multiple microbial communities and key nutritional and chemical factors that promote bacterial growth and exhibit physical properties mimicking those of biofilms from CF sputum.
The authors further explained, “Artificial sputum medium (ASM) has been used to recreate the environment found in the lung of cystic fibrosis (CF) patients because it includes key nutritional components of native sputum, such as salts, DNA, and key proteins such as mucin, which enable the growth of bacteria found in these patients as biofilms.” The material has been used to build an infected in vitro lung epithelial model that was used to study the impact of antibiotics. “The main goal of the reported study was to develop a technology that would allow practical and relevant investigations of real-life microbial communities for lab testing, “which is the ultimate objective and innovative aspect of our work,” the authors continued.
The 3D living material supported the growth and enabled production of 3D biofilms of the three major CF pathogens, Pseudomonas aeruginosa PAO1-L, Staphylococcus aureus SH1000, and Candida albicans SC5314, individually and as a polymicrobial community of the species. In their reported work the team used the platform to study the effects of the antibiotic ciprofloxacin. “Altogether, we developed a ‘‘biocooperative’’ platform that harnesses a complex biofluid (ASM) to engineer a living material that can recreate features of 3D biofilms and build an interkingdom in vitro model of polymicrobial communities and living tissue cells,” the team stated. “We demonstrate its capacity to support the growth of 3D polymicrobial biofilms and build an interkingdom infected lung epithelial model to study the impact of the antibiotic ciprofloxacin.”