Genetic engineering biofilms in situ using ultrasound‐mediated DNA delivery

Abstract

The ability to directly modify native and established biofilms has enormous potential in understanding microbial ecology and application of biofilm in 'realworld' systems. However, efficient genetic transformation of established biofilms at any scale remains challenging. In this study, we applied an ultrasoundmediated DNA delivery (UDD) technique to introduce plasmid to established non-competent biofilms in situ. Two different plasmids containing genes coding for superfolder green fluorescent protein (sfGFP) and the flavin synthesis pathway were introduced into established bacterial biofilms in microfluidic flow (transformation efficiency of 3.9 AE 0.3 × 10 -7 cells in biofilm) and microbial fuel cells (MFCs), respectively, both employing UDD. Gene expression and functional effects of genetically modified bacterial biofilms were observed, where some cells in UDD-treated Pseudomonas putida UWC1 biofilms expressed sfGFP in flow cells and UDD-treated Shewanella oneidensis MR-1 biofilms generated significantly (P < 0.05) greater (61%) bioelectricity production (21.9 AE 1.2 µA cm −2 ) in MFC than a wild-type control group (~13.6 AE 1.6 µA cm −2 ). The effects of UDD were amplified in subsequent growth under selection pressure due to antibiotic resistance and metabolism enhancement. UDD-induced gene transfer on biofilms grown in both microbial flow cells and MFC systems was successfully demonstrated, with working volumes of 0.16 cm 3 and 300 cm 3 , respectively, demonstrating a significant scale-up in operating volume. This is the first study to report on a potentially scalable direct genetic engineering method for established non-competent biofilms, which can be exploited in enhancing their capability towards environmental, industrial and medical applications.

Microbe–Anode Interactions: Comparing the impact of genetic and material engineering approaches to improve the performance of microbial electrochemical systems (MES)

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“…This study highlights that our knowledge especially concerning anaerobic biofilm formation and the underlying mechanisms and networks is still very incomplete, and that more research is required in this area. Ng et al (2021) pursued a rather different strategy. The introduction of an additional copy of the gene cluster necessary for riboflavin production (ribADEHC) into S. oneidensis resulted in a 2.3-fold increased final current density and 2-fold increase in maximum power density.…”

Bacterial biofilm functionalization through Bap amyloid engineering

“…Recently, novel strategies have been developed to create biofilms as artificial platforms for self-assembling functional tags 32 , 34 , 35 . One of these approaches is based on engineering extracellular matrix components, focusing on amyloids as potential targets 8 , 9 , 11 .…”

Induction of potent antitumor immunity by intradermal DNA injection using a novel needle‐free pyro‐drive jet injector

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