*2.1.3 Phage-SERS-based sensors*

An innovative Raman method, i.e., surface-enhanced Raman spectroscopy (SERS), is enhancing the intensity by vibrational absorbance of definite elements when they are near the surface of nano-organized noble metals by the influence of numerous orders of magnitude. The improved intensity of SERS method is dependent on the molecules' capability to release a Raman signal and the contained fields of plasmon in their neighborhood [21]. For instance, a report stated a phage-SERS biosensor for *E. coli* detection using phage immobilization on nano-figured thin sheet of silver over substrates of silica (**Figure 2**) [22] established by exploitation of metallic nanosculptured thin silver film. The silver film exterior is activated by self-assembled monolayer of 4-aminothiophenol and glutaraldehyde for T4 immobilization to screen *E. coli*. As a reporter molecule, 4-aminothiophenol monitored the Raman band enhancement. Other reports of phage-SERS-based biosensors have been reported and are briefed in **Table 1**.

#### **Figure 2.**

*Schematic representation of phage-SERS-based sensor. Adapted from [22].*


**179**

[23] and 104

*2.1.4 Phage-fluorescent sensor*

*aureus; LB, Luria-Bertani broth.*

**Table 1.**

*Applications of Phage-Based Biosensors in the Diagnosis of Infectious Diseases, Food Safety…*

SERS T4 phage *E. coli B* Buffer 150 CFU/mL [22]

A511 phage *L. monocytogenes* — 6.1 × 107

Phage *S. typhimurium* — 1.5 × 103

Amperometric B1-7064 phage *B. cereus* — 10 CFU/mL [70]

T2 phage *E. coli* B Broth 103

Lytic phage *S. Newport* — 103

*Gamma* phage *B. anthracis Str* Water 103

*SPR, surface plasmon resonance; scFv, single-chain variable fragment; MRSA, methicillin-resistant Staphylococcus aureus; PBS, phosphate-buffered saline; TNB, trinitrobenzene; TNT, trinitrotoluene; QCM, quartz crystal microbalance; QD, quantum dot; SERS, surface-enhanced Raman spectroscopy; LFA, lateral flow assay; HRP, horseradish peroxidase; CFU, colony-forming unit; PFU, plaque-forming unit; E. coli, Escherichia coli; S. arlettae, Staphylococcus arlettae; B. anthracis, Bacillus anthracis; P. aeruginosa, Pseudomonas aeruginosa; S. flexneri, Shigella flexneri; S. Newport, Salmonella Newport; S. typhimurium, Salmonella typhimurium; S. aureus, Staphylococcus* 

T4 phage *E. coli* B Water, milk 800 CFU/mL

*L. monocytogenes* Milk 105

Phage 12,600 MRSA — — [63]

JRB7 phage *B. anthracis* — Spores [67]

M13 phage *E. coli TG1* — 1 CFU/mL [71]

lettuce

samples

**Target bacteria/ analyte**

P9b phage *P. aeruginosa* Clinical

E2 phage *S. typhimurium* Romaine

Magnetoelastic E2 phage *S. typhimurium* — 5 × 102

Impedimetric T4 phage *E. coli* — 104

In fluorescent-phage-based sensor techniques, fluorescently stained phages are utilized as marking agents for the detection of bacterial cells. Fluorescently labeled phages are identified followed by binding to specific host bacterial cell. The composite of bacteriophage-bacteria is then sensed by means of flow cytometry or epi-fluorescent filter approach. A combination of immunomagnetic separation

*Applications of phage/phage components in detection of infectious pathogen and other deadly analytes related to food safety and environmental monitoring, where transduction platform used, target analyte/bacteria,* 

bacteria *E. coli* O157:H7 after 10 h augmentation in artificially contaminated milk

in the sensitivity of this method was reported by using fluorescent quantum dots (QDs) for phage labeling [25]. Also fluorescent-based sensors have been used for bacterial toxin recognition. Phage display was applied to choose a peptide (12-mer) that was able to attach to *staphylococcal enterotoxin B* (SEB) that is responsible for food poisoning [26]. This approach permitted toxin sensing and detected 1.4 ng of SEB/sample well with the help of fluorescence immunoassay and involved a fluorescently stained SEB binding bacteriophage. Also array-based sensors have been

established following the same principle for simultaneous detection of

CFU/mL in sample of broth medium [24]. Additional improvement

CFU/mL of pathogenic

**Sample Detection limit Ref.**

103

5 × 102

CFU/mL [64]

pfu/mL [65]

CFU/mL [66]

CFU/mL [68]

CFU/mm2 [69]

CFU/mL [72]

CFU/mL [73]

CFU/mL [74]

CFU/mL [75]

CFU/mL [77]

[76]

100 CFU/mL

with fluorescent method is detected between 10 and 102

*sample processed, and limit of detection are briefed with reported literature.*

*DOI: http://dx.doi.org/10.5772/intechopen.88644*

**bio-probe**

Endolysin Ply500

**Transducer Phage-based** 


*Applications of Phage-Based Biosensors in the Diagnosis of Infectious Diseases, Food Safety… DOI: http://dx.doi.org/10.5772/intechopen.88644*

*SPR, surface plasmon resonance; scFv, single-chain variable fragment; MRSA, methicillin-resistant Staphylococcus aureus; PBS, phosphate-buffered saline; TNB, trinitrobenzene; TNT, trinitrotoluene; QCM, quartz crystal microbalance; QD, quantum dot; SERS, surface-enhanced Raman spectroscopy; LFA, lateral flow assay; HRP, horseradish peroxidase; CFU, colony-forming unit; PFU, plaque-forming unit; E. coli, Escherichia coli; S. arlettae, Staphylococcus arlettae; B. anthracis, Bacillus anthracis; P. aeruginosa, Pseudomonas aeruginosa; S. flexneri, Shigella flexneri; S. Newport, Salmonella Newport; S. typhimurium, Salmonella typhimurium; S. aureus, Staphylococcus aureus; LB, Luria-Bertani broth.*

#### **Table 1.**

*Biosensors for Environmental Monitoring*

**Transducer Phage-based** 

**Figure 2.**

QCM Filamentous

phage

**bio-probe**

**Target bacteria/ analyte**

T4 phage *E. coli O157*:H7 PBS 103

BP14 phage MRSA PBS 103

scFv phages *L. monocytogenes* — 2.106

12,600 phage *S. aureus* — 104

Luminescence *lacZ* T4 phage *E. coli* B Water 10 CFU/mL [7] SJ2 phage *S. enteritidis* — 103

Lytic phage *Listeria innocua* — >104

Shfl25875 *S. flexneri* Stool 103

*Gamma* phage *B. anthracis* — 2.5 × 104

T7 phage *E. coli* Broth 103

Fluorescent P22 phage *S. typhimurium* Milk 1 CFU/24 mL [57] P-*S. aureus*-9 *S. aureus* PBS 2.47 × 103

Wβ phage *B. anthracis* Soil 104

O157-IOV 4 *E. coli O157*:H7 Milk 4.9 × 104

Wild-type *E. coli* K12 — 103

Pap1 phage *P. aeruginosa* Milk, urine 56 CFU/mL [3]

PP01 phage *E. coli* O157:H7 Apple juice 1 CFU/mL [60] PDPs TNT — 10 μg/mL [61] T7 phage *E. coli* LB broth 10 CFU/mL [62]

*S. typhimurium* — 102

T4 phage *E. coli* Milk Few CFU/mL [9]

SPR T4 phage *E. coli K12* PBS 7 × 102

*Schematic representation of phage-SERS-based sensor. Adapted from [22].*

LFA B4 phage *B. cereus* Buffer 1 × 104

**Sample Detection limit Ref.**

CFU/mL [15]

CFU/mL [17]

CFU/mL [17]

CFU/mL [53]

CFU/mL [16]

CFU/mL [20]

CFU/mL [54]

CFU/g [55]

CFU/mL [11]

CFU/mL [56]

CFU/L [58]

CFU/g [50]

CFU/mL [59]

CFU/mL [35]

CFU/mL [6]

CFU/mL [47]

**178**

*Applications of phage/phage components in detection of infectious pathogen and other deadly analytes related to food safety and environmental monitoring, where transduction platform used, target analyte/bacteria, sample processed, and limit of detection are briefed with reported literature.*

#### *2.1.4 Phage-fluorescent sensor*

In fluorescent-phage-based sensor techniques, fluorescently stained phages are utilized as marking agents for the detection of bacterial cells. Fluorescently labeled phages are identified followed by binding to specific host bacterial cell. The composite of bacteriophage-bacteria is then sensed by means of flow cytometry or epi-fluorescent filter approach. A combination of immunomagnetic separation with fluorescent method is detected between 10 and 102 CFU/mL of pathogenic bacteria *E. coli* O157:H7 after 10 h augmentation in artificially contaminated milk [23] and 104 CFU/mL in sample of broth medium [24]. Additional improvement in the sensitivity of this method was reported by using fluorescent quantum dots (QDs) for phage labeling [25]. Also fluorescent-based sensors have been used for bacterial toxin recognition. Phage display was applied to choose a peptide (12-mer) that was able to attach to *staphylococcal enterotoxin B* (SEB) that is responsible for food poisoning [26]. This approach permitted toxin sensing and detected 1.4 ng of SEB/sample well with the help of fluorescence immunoassay and involved a fluorescently stained SEB binding bacteriophage. Also array-based sensors have been established following the same principle for simultaneous detection of

*Bacillus globigii*, MS2 bacteriophage, and also SEB [27]. The typically reported sensitivity until now is about 20 CFU/mL by epi-fluorescent microscopic platform [25] and is 1 CFU/mL by flow cytometric recognition approach [28].
