Multiplexed detection and differentiation of bacterial enzymes and bacteria by color-encoded sensor hydrogels

Abstract

We report on the fabrication and characterization of color-encoded chitosan hydrogels for the rapid, sensitive and specific detection of bacterial enzymes as well as the selective detection of a set of tested bacteria through characteristic enzyme reactions. These patterned sensor hydrogels are functionalized with three different colorimetric enzyme substrates affording the multiplexed detection and differentiation of α-glucosidase, β-galactosidase and β-glucuronidase. The limits of detection of the hydrogels for an observation time of 60 min using a conventional microplate reader correspond to concentrations of 0.2, 3.4 and 4.5 nM of these enzymes, respectively. Based on their different enzyme expression patterns, Staphylococcus aureus strain RN4220, methicillin-resistant S . aureus (MRSA) strain N315, both producing α-glucosidase, but not β-glucuronidase and β-galactosidase, Escherichia coli strain DH5α, producing β-glucuronidase and α-glucosidase, but not β-galactosidase, and the enterohemorrhagic E . coli (EHEC) strain E32511, producing β-galactosidase, but none of the other two enzymes, can be reliably and rapidly distinguished from each other. These results confirm the applicability of enzyme sensing hydrogels for the detection and discrimination of specific enzymes to facilitate differentiation of bacterial strains. Patterned hydrogels thus possess the potential to be further refined as detection units of a multiplexed format to identify certain bacteria for future application in point-of-care microbiological diagnostics in food safety and medical settings.

An Overview on Recent Progress of the Hydrogels: From Material Resources, Properties, to Functional Applications

“…Color-coded chitosan hydrogels are used for rapid, sensitive, and specific detection of bacterial enzymes [142] Preparation and characterization of hydrogels obtained from chitosan and carboxymethyl chitosan Preparation and characterization of hydrogels [156] Chitosan and cellulose-based hydrogels for wound management Wound dressing materials [157] Agarose Nanosilicate embedded agarose hydrogels with improved bioactivity Nanocomposite agarose hydrogel to promote cell binding, growth, and proliferation [108] Facile formation of agarose hydrogel and electromechanical responses as electro-responsive hydrogel materials in actuator applications…”

“…For example, Xiao et al reported color-coded chitosan hydrogels that can be used for rapid, specific, and sensitive detection of bacterial enzymes, as well as selective detection of bacteria through the characteristic enzyme reactions. [142] Nano-cellulose is a famous reproducible biopolymer which is released from cellulose fiber through multiple processes, such comments as acid hydrolysis, chemical fiber formation or mechanical treatment. [143] Because of its rich surface charges and surface functional groups, Nano-cellulose has potential application prospects as carriers/dispersants.…”

Potential Economic Value of Chitin and Its Derivatives as Major Biomaterials of Seafood Waste, with Particular Reference to Southeast Asia

“…Chitosan is also playing a very important role in the area of sensor development, especially in the topic of biological evaluation. The versatility of chitosan in forming various forms of structures such as films, microgel/hydrogel and nanocomposites, and its adhesive properties are the main reasons for its being a popular candidate for sensor development [191,192]. These chitosan nanocomposite-based biosensors have shown high sensitivity, selectivity, and stability for detecting a variety of targets, ranging from biomolecules, DNA, and microorganisms.…”

Advanced Functional Membranes for Sensor Technologies

“…Furthermore, these membranes must be non-toxic and biocompatible to minimize the risk of epithelialization or allergic reactions. In this context, the use of enzymes [54][55][56][57], hydrogels [6,41,[58][59][60], and conductive polymers such as polyaniline (PANI) [61][62][63][64], polypyrrole (PPy) [65][66][67], polythiophene (PT) [68][69][70], or polyethylene dioxide thiophene (PEDOT) [43,[71][72][73][74] has been reported for the development of bioelectrodes due to their biocompatibility, conductivity, and the presence of tunable functionalities. However, its application in the development of biosensors is limited by different reasons and challenges to overcome, such as its relative fragility; its decrease in properties against variations in the pH of the medium, possible degradation in toxic or reactive compounds; and in some cases the use of synthesis processes and complex manufacturing procedures that are difficult to implement outside the laboratory scale [70][71][72][73][74][75][76].…”