Mango peel as an enzyme inducer in Trichoderma harzianum: a strategy for cariogenic biofilm degradation and reuse of industrial waste

Author(s)

Jéssica Silva Peixoto Bem

• Document type: Master's Dissertation
• Press: Ribeirão Preto.
• Institution: Universidade de São Paulo (USP). Faculdade de Odontologia de Ribeirão Preto (PCARP/BC)
• Defense date: 2022-07-18

Examining board members

Carolina Patricia Aires Garbellini; Maria Cristina Borsato; Cristiane Duque

• Advisor: Carolina Patricia Aires Garbellini; Carem Gledes Vargas Rechia

Abstract

The industrial use of fruits such as mango produces tons of agro-industrial waste that are mostly discarded. It has already been demonstrated that mango pulp induces the secretion of glucano-hydrolytic enzymes in the fungus Trichoderma harzianum, which may affect insoluble polysaccharides, one of the main virulence factors of cariogenic biofilms. However, the biofilm degradation potential of fungal enzymes induced from one of the main industrial wastes of mango - the peel - has not yet been explored, which is the objective of this work. In the first step, fractions of the peel (PL) and ethanol-precipitated pulp (PP) of Tommy Atkins mangoes were obtained, with an aliquot reserved for the determination of the monosaccharide composition and uronic acid content. The remaining aliquots of both fractions were sterilized and added to the culture medium containing T. harzianum for induction of hydrolytic enzymes. After 192 hours, the culture medium was centrifuged and while part of the supernatant (enzyme extract) was used for proteomic analysis, another part was reserved for the second stage of the experiment, in which S. mutans biofilms, in a \"feast and famine\" were exposed to the treatments (n=9): a) NaCl 0.9% (negative control); b) 0.12% chlorhexidine digluconate (positive control); c) extract of enzymes induced by PL or PP or insoluble polysaccharide from S. mutans (IPS, positive control for glucanases). Acidogenicity, bacterial viability, quantification of insoluble polysaccharides, and three-dimensional analysis of the biofilm by scanning electron microscopy (SEM) were determined after each experimental period, in three different replications. From the ANOVA followed by the Tukey test (&alpha;=5%), it was observed that the extracts induced by PL and PP showed lower specific activities than the IPS group (p<0.05). The percentage of uronic acid was lower in the PL fraction compared to the PP fraction, being predominantly constituted by galactose and arabinose, respectively. The hydrolytic enzymes produced from the incubation of PL or PP with T. harzianum did not alter the metabolism or bacterial viability of the biofilm (p<0.05). Although the images obtained by SEM suggest some degree of matrix degradation, the quantification of insoluble glucans for the PL and PP groups did not differ from the control group (p>0.05), suggesting a slight effect on the disorganization of the S. mutans biofilm. Considering that the matrix enzymatic degradation strategy can enhance the performance of antimicrobials, future studies should combine this initiative with the use of natural products, which may have a synergistic effect on biofilm control. (AU)

• FAPESP's process: 20/07315-0 - Mango processing by-products for cariogenic biofilm control: in vitro study
• Grantee: Jéssica Silva Peixoto Bem
• Support Opportunities: Scholarships in Brazil - Master