Genomic and Kinetic Modeling Involving Nanoparticle-Mediated Delivery of a Novel Chitinase Enzyme to Outpace Tuta absoluta Damage

Abstract

The tomato leafminer, Tuta absoluta, poses a severe global agricultural threat due to its rapid leaf-mining behavior and swift development of resistance to conventional chemical pesticides. While microbial chitinases are potent biopesticides, their field efficacy is limited by environmental degradation and the short exposure window before larvae penetrate leaf tissues. This study evaluates a stimuli-responsive, controlled-release nanobiopesticide system utilizing a novel chitinase from newly characterized Serratia marcescens GBS19. A 61.1 kDa chitinase (GBS19_ChiA) was heterologously expressed in Escherichia coli and purified to a specific activity of 215.01 U/mg. The enzyme was immobilized onto starch-coated silica nanoparticles designed for target-triggered release via host alpha-amylase. Genomic profiling and R-based kinetic modeling were integrated to evaluate the efficacy of purified and immobilized forms against T. absoluta. Immobilization enhanced thermal and pH stability, with the nanocarrier maintaining 85% activity over 10 weeks. In larval bioassays, immobilization increased mortality from 21.9% to 59.4% (5000 U/mL) by day 3, reaching 62.5% by day 6. Genomic analysis identified an expansive secretome and a Type VI Secretion System (T6SS), characterizing GBS19 as a multi-pronged pathogen. Kinetic modeling established that while immobilized enzymes are effective, the 2.5-hour exposure time on T. absoluta requires the synergistic action of chitinases (ChiA/B/C) to reach the lethal desiccation threshold before larvae establish protective mines. Starch-coated silica nanoparticles significantly improve chitinase stability and delivery. However, overcoming the rapid penetration of T. absoluta necessitates a whole-cell or multi-enzyme synergistic approach to outpace larval behavioural defences.