Enzymatic disruption of dental biofilm by mutanase/dextranase in bioencapsulated plant cells for oral care at home during the pandemic

Pooja A. Patel

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Patel, Pooja A.
Faculty / Advisor: Daniell, Henry
University of Pennsylvania School of Dental Medicine, Department of Basic and Translational Sciences

Introduction

Caries develops as a result of plaque biofilm development on teeth surface. Such biofilm contains alpha-1,3 and 1,6-glycosidic bonds, which allow for the development of tight clusters and extracellular matrices (ECM). Matrix degrading enzymes are effective against dental plaque due to their ability to break alpha linkages. However, their clinical feasibility is hindered by their high cost, as they are produced in prohibitively expensive microbial fermentation systems. Additionally, these enzymes require expensive purification and cold storage to preserve their activity. Therefore, this study investigated enzymatic disruption of biofilm using dextranase and mutanase expressed in lettuce cells because they don’t require purification, can be stored at ambient temperature, and can be topically delivered using chewing gum. Enhancing oral healthcare at home is feasible using chewing gum, especially during the pandemic.

Methods

Several chloroplast vectors were developed containing dextranase gene from Streptococcus mutans and mutanase gene from Paenibacillus sp. These vector constructs were bombarded into WT Lactuca sativa (lettuce) to transform the chloroplast DNA. Successful bombardment was confirmed using PCR and gel electrophoresis. Southern blot was done to determine whether homoplasmy was achieved and if the antibiotic resistance gene was removed. Plants that reached this stage of selection were grown, then lyophilized for preservation. The dried plant material was further analyzed for enzymatic activity.

Results

My investigations using Southern blot confirmed that both dextranase and mutanase genes were stably integrated into the lettuce chloroplast genome. They also confirmed removal of the antibiotic resistance gene (aadA) used for the selection process. The absence of untransformed genome fragments in the Southern blots confirmed that all 10,000 copies of chloroplast genomes contained insertions of dextranase or mutanase genes and thereby achieved homoplasmy. Other lab investigators confirmed the effectiveness of plant extracts in the disruption of biofilm. Plant powder crude extracts without purification performed equal to or better than commercial mutanase/dextranase enzymes in disrupting dental biofilm matrix.

Conclusion

When completed, the development of a chewing gum protein drug from lyophilized plants with EPS degrading enzymes would provide a low-cost, low-maintenance drug that is easily preserved and can be used at home to disrupt the pathogenic biofilm. Current approaches for oral health care are unaffordable to impoverished populations. Additionally, aerosolized droplets in the dental clinic and poor oral hygiene may contribute to the spread of several infectious diseases including global pandemic COVID-19, requiring new solutions for dental biofilm/plaque treatment at home.