Conventional, Molecular, and Biosensor-Based Detection Methods ◾ 557
function of time at a given temperature can be monitored. Impedance detection time (IDT) is
the time required by microorganisms initially present in a food to reach ≥10 6 /mL. It is inversely
proportional to the initial microbial load; a higher IDT means a lower initial load. From the
IDT and the slope of the curve, the initial load as well as generation time of a microbial population can be calculated. The major advantage of this system is that it allows detection of only
the viable cells, which is a major concern in food safety. At present, impedance instruments are
able to detect 10 5 –10 7 bacteria per milliliter. Several commercially available systems, such as the
Bactometer ® (bioMérieux), Malthus AT analyzer (Malthus Instruments), BacTrac, and μ-Trac
microorganism growth analyzer (SyLab), are used for pathogen monitoring and quality assurance
purposes. Bacterial contamination in milk has been analyzed using the Bactometer for quality
assessment purposes. The impedance-based assay has been used for detection and enumeration of
Listeria, Campylobacter, Esc. coli, Staphylococcus, and Salmonella from food samples. The impedance method has been accepted by the Association of Official Analytical Chemists, International
(AOAC) as a first-action method. This macroscale method, however, lacks sensitivity and time-toresult is long, requiring 24–48 hours.
To improve sensitivity, miniaturized microfabricated electronic devices, such as semiconductor silicone chips referred to as “biochips,” are developed. In this biology-based, microelectricalmechanical systems (Bio-MEMS) device, the cells are electrophoretically confined into a nanoliter
volume in a chamber, thus allowing rapid turnover of substrate by bacteria into electrically charged
byproducts aiding rapid and sensitive detection of bacterial growth by impedance. Lis. monocytogenes cells were successfully detected at 2 × 10 7 cells/mL chamber with only a two hour incubation
step. Pathogen-specific antibodies, receptor molecules, such as virulence proteins or bacteriophage
receptor proteins, immobilized in the detection chamber can provide specificity for target pathogen. Pathogens are also detected on a microfluidic chip by PCR.
Fourier Transform Infrared Spectroscopy
(FTIR) 29,42 and Raman 43 Spectroscopy
Both FTIR and Raman spectroscopy generate bacterial spectral scans based on the molecular
composition of the cell wall and the cytoplasm. Basically, infrared spectroscopy consists of the
infrared source, sample, and the detector. When infrared radiation is absorbed or transmitted
through the sample to the detector, it generates a “scan” or “fingerprint” profile. A library of
spectral scans can be generated for different bacterial species and strains, which can be used for
future comparison. This method requires a transfer of cells (biomass) from the growth media to
the IR-reflecting surface for spectral collection. It is a nondestructive label-free rapid method, and
future identification depends on the available spectral library. FTIR has been used for classification or identification of several foodborne pathogens: Yersinia, Staphylococcus, Salmonella, Listeria,
Klebsiella, Escherichia, Enterobacter, Citrobacter, and so forth.
Raman spectroscopy is also a label-free detection system and uses a diode laser (785 nm),
which, upon interaction with microbial cells, generates Raman scatter, known as inelastic scattering. Typically, the Raman signal is very weak, and to amplify the signal, surface-enhanced Raman
spectroscopy (SERS) has been configured using biorecognition molecules conjugated to gold or
silver nanoparticles to specifically bind to target microorganisms or toxin molecules. Raman is
used for detection of Bacillus anthracis, Yersinia pestis, Burkholderia mallei, Francisella tularensis,
Brucella abortus, Lis. monocytogenes, Esc. coli O157:H7, and Salmonella.
function of time at a given temperature can be monitored. Impedance detection time (IDT) is
the time required by microorganisms initially present in a food to reach ≥10 6 /mL. It is inversely
proportional to the initial microbial load; a higher IDT means a lower initial load. From the
IDT and the slope of the curve, the initial load as well as generation time of a microbial population can be calculated. The major advantage of this system is that it allows detection of only
the viable cells, which is a major concern in food safety. At present, impedance instruments are
able to detect 10 5 –10 7 bacteria per milliliter. Several commercially available systems, such as the
Bactometer ® (bioMérieux), Malthus AT analyzer (Malthus Instruments), BacTrac, and μ-Trac
microorganism growth analyzer (SyLab), are used for pathogen monitoring and quality assurance
purposes. Bacterial contamination in milk has been analyzed using the Bactometer for quality
assessment purposes. The impedance-based assay has been used for detection and enumeration of
Listeria, Campylobacter, Esc. coli, Staphylococcus, and Salmonella from food samples. The impedance method has been accepted by the Association of Official Analytical Chemists, International
(AOAC) as a first-action method. This macroscale method, however, lacks sensitivity and time-toresult is long, requiring 24–48 hours.
To improve sensitivity, miniaturized microfabricated electronic devices, such as semiconductor silicone chips referred to as “biochips,” are developed. In this biology-based, microelectricalmechanical systems (Bio-MEMS) device, the cells are electrophoretically confined into a nanoliter
volume in a chamber, thus allowing rapid turnover of substrate by bacteria into electrically charged
byproducts aiding rapid and sensitive detection of bacterial growth by impedance. Lis. monocytogenes cells were successfully detected at 2 × 10 7 cells/mL chamber with only a two hour incubation
step. Pathogen-specific antibodies, receptor molecules, such as virulence proteins or bacteriophage
receptor proteins, immobilized in the detection chamber can provide specificity for target pathogen. Pathogens are also detected on a microfluidic chip by PCR.
Fourier Transform Infrared Spectroscopy
(FTIR) 29,42 and Raman 43 Spectroscopy
Both FTIR and Raman spectroscopy generate bacterial spectral scans based on the molecular
composition of the cell wall and the cytoplasm. Basically, infrared spectroscopy consists of the
infrared source, sample, and the detector. When infrared radiation is absorbed or transmitted
through the sample to the detector, it generates a “scan” or “fingerprint” profile. A library of
spectral scans can be generated for different bacterial species and strains, which can be used for
future comparison. This method requires a transfer of cells (biomass) from the growth media to
the IR-reflecting surface for spectral collection. It is a nondestructive label-free rapid method, and
future identification depends on the available spectral library. FTIR has been used for classification or identification of several foodborne pathogens: Yersinia, Staphylococcus, Salmonella, Listeria,
Klebsiella, Escherichia, Enterobacter, Citrobacter, and so forth.
Raman spectroscopy is also a label-free detection system and uses a diode laser (785 nm),
which, upon interaction with microbial cells, generates Raman scatter, known as inelastic scattering. Typically, the Raman signal is very weak, and to amplify the signal, surface-enhanced Raman
spectroscopy (SERS) has been configured using biorecognition molecules conjugated to gold or
silver nanoparticles to specifically bind to target microorganisms or toxin molecules. Raman is
used for detection of Bacillus anthracis, Yersinia pestis, Burkholderia mallei, Francisella tularensis,
Brucella abortus, Lis. monocytogenes, Esc. coli O157:H7, and Salmonella.