Pathogenic bacterial contamination is a major threat to human health and safety. In this review, we summarize recent strategies for the integration of recognition elements with nanomaterials for the detection and sensing of pathogenic bacteria. Nanoprobes can provide sensitive and specific detection of bacterial cells, which can be applied across multiple applications and industries.

In this study, we demonstrate a bacteriophage (phage)-based magnetic separation scheme for the rapid detection of Escherichia coli (E. coli) in drinking water. T7 phage is a lytic phage with a broad host range specificity for E. coli. Our scheme was as follows: (1) T7 bacteriophage-conjugated magnetic beads were used to capture and separate E. coli BL21 from drinking water; (2) subsequent phage-mediated lysis was used to release endemic β-galactosidase (β-gal) from the bound bacterial cells; (3) the release of β-gal was detected using chlorophenol red-β-d-galactopyranoside (CRPG), a colorimetric substrate which changes from yellow to red in the presence of β-gal. Using this strategy, we were able to detect E. coli at a concentration of 1 × 10(4) CFU·mL(-1) within 2.5 h. The specificity of the proposed magnetic probes toward E. coli was demonstrated against a background of competing bacteria. By incorporating a pre-enrichment step in Luria-Bertani (LB) broth supplemented with isopropyl β-d-thiogalactopyranoside (IPTG), we were able to detect 10 CFU·mL(-1) in drinking water after 6 h of pre-enrichment. The colorimetric change can be determined either by visual observation or with a reader, allowing for a simple, rapid quantification of E. coli in resource-limited settings.

A novel enzyme-induced metallization colorimetric assay was developed to monitor and measure beta-galactosidase (β-gal) activity, and was further employed for colorimetric bacteriophage (phage)-enabled detection of Escherichia coli (E. coli). This assay relies on enzymatic reaction-induced silver deposition on the surface of gold nanorods (AuNRs). In the presence of β-gal, the substrate p-aminophenyl β-D-galactopyranoside (PAPG) is hydrolyzed to produce p-aminophenol (PAP). Reduction of silver ions by PAP generates a silver shell on the surface of AuNRs, resulting in the blue shift of the longitudinal localized surface plasmon resonance (LSPR) peak and multi-color changes of the detection solution from light green to orange-red. Under optimized conditions, the detection limit for β-gal was 128 pM, which was lower than the conventional colorimetric assay. Additionally, the assay had a broader dynamic range for β-gal detection. The specificity of this assay for the detection of β-gal was demonstrated against several protein competitors. Additionally, this technique was successfully applied to detect E. coli bacteria cells in combination with bacteriophage infection. Due to the simplicity and short incubation time of this enzyme-induced metallization colorimetric method, the assay is well suited for the detection of bacteria in low-resource settings.

Research in microfluidic biosensors has led to dramatic improvements in sensitivities. Very few examples of these devices have been commercially successful, keeping this methodology out of the hands of potential users. In this study, we developed a method to fabricate a flexible microfluidic device containing electrowetting valves and electrochemical transduction. The device was designed to be amenable to a roll-to-roll manufacturing system, allowing a low manufacturing cost. Microchannels with high fidelity were structured on a PET film using UV-NanoImprint Lithography (UV-NIL). The electrodes were inkjet-printed and photonically sintered on second flexible PET film. The film containing electrodes was bonded directly to the channel-containing layer to form sealed fluidic device. Actuation of the multivalve system with food dye in PBS buffer was performed to demonstrate automated fluid delivery. The device was then used to detect Salmonella in a liquid sample.

A pragmatic method to deposit silver nanoparticles on polydopamine-coated nanoimprinted pillars for use as surface-enhanced Raman scattering (SERS) substrates was developed. Pillar arrays consisting of poly(methyl methacrylate) (PMMA) that ranged in diameter from 300 to 500 nm were fabricated using nanoimprint lithography. The arrays had periodicities from 0.6 to 4.0 μm. A polydopamine layer was coated on the pillars in order to facilitate the reduction of silver ions to create silver nucleation sites during the electroless deposition of sliver nanoparticles. The size and density of silver nanoparticles were controlled by adjusting the growth time for the optimization of the SERS performance. The size of the surface-adhered nanoparticles ranged between 75 and 175 nm, and the average particle density was ∼30 particles per μm(2). These functionalized arrays had a high sensitivity and excellent signal reproducibility for the SERS-based detection of 4-methoxybenzoic acid. The substrates were also able to allow the SERS-based differentiation of three types of bacteriophages (λ, T3, and T7).

The application of bacteriophage combined with the use of magnetic separation techniques has emerged as a valuable tool for the sensitive identification and detection of bacteria. In this study, bacteriophage T7 labelled magnetic beads were developed for the detection of viable bacterial cells. Fusion of the biotin acceptor peptide (BAP) with the phage capsid protein gene and the insertion of the biotin ligase (BirA) gene enabled the display of the BAP ligand and the expression protein BirA during the replication cycle of phage infection. The replicated Escherichia coli specific bacteriophage was biotinylated in vivo and coated on magnetic beads via streptavidin-biotin interaction. Immobilization efficiency of the recombinant phage was investigated on magnetic beads and the phage-bead complex was evaluated by detecting E. coli from inoculated broth. When compared to the wild type phage, the recombinant phage T7birA-bap had a high immobilization density on streptavidin-coated magnetic beads and could capture 86.2% of E. coli cells from broth within 20 min. As this phage-based biomagnetic detection approach provided a low detection limit of 10(2) CFU mL(-1) without pre-enrichment, we believe this assay could be further developed to detect other bacteria of interest by applying host-specific phages. This would be of particular use in detecting bacteria which are difficult to grow or replicate slowly in culture.

Here we presented a simple, rapid and label-free surface-enhanced Raman spectroscopy (SERS) based mapping method for the detection and discrimination of Salmonella enterica and Escherichia coli on silver dendrites. The sample preparation was first optimized to maximize sensitivity. The mapping method was then used to scan through the bacterial cells adsorbed on the surface of silver dendrites. The intrinsic and distinct SERS signals of bacterial cells were used as the basis for label-free detection and discrimination. The results show the developed method is able to detect single bacterial cells adsorbed on the silver dendrites with a limit of detection as low as 10(4) CFU mL(-1), which is two orders of magnitude lower than the traditional SERS method under the same experimental condition. The time needed for collecting a 225 points map was approximately 24 minutes. Moreover, the developed SERS mapping method can realize simultaneous detection and identification of Salmonella enterica subsp. enterica BAA1045 and Escherichia coli BL21 from a mixture sample using principle component analysis. Our results demonstrate the great potential of the label-free SERS mapping method to detect, identify and quantify bacteria and bacterial mixtures simultaneously.

A novel and pragmatic method was developed to detect the concentration of nitrite ions using Fe3O4@SiO2/Au magnetic nanoparticles (MNPs) by surface-enhanced Raman scattering (SERS). The as-prepared bifunctional nanocomposites can be used to simultaneously purify target molecules using external magnetic field and produce Raman fingerprint spectrum with trace level of target molecules. In acidic media, 4-aminothiophenol (4-ATP) molecules conjugated on Fe3O4@SiO2/Au MNPs were triggered by nitrite ions to form azo bonds, resulting in three new marker peaks on the SERS spectrum. Under optimized conditions, the detection limit based on the three marker peaks were 15.63, 13.69, and 17.77μM, which was much lower than the maximum NO2(-) concentration of 1.0mgL(-1) (71.4μM) allowed in drinking water as defined by U.S. Environmental Protection Agency (EPA). The specificity of this proposed method to detect nitrite ions was demonstrated using common ions as competitors. In addition, the SERS-based technique was successfully employed to detect nitrite ions in pond water, a synthetic urine solution, and pickle brine. Considering its good sensitivity and selectivity, the detection method is well suited for the detection of nitrite ions in real samples without pretreatment steps.