Polymerase chain reaction

The polymerase chain reaction forts appeared in 1985 as a method for the prenatal diagnosis of sickle cell anemia. It was based on the remarkable insight of Kary Mullis, who realized that repetition of a DNA extension reaction bounded by two synthetic oligonucleotide primers would generate a large quantity of any specified DAN sequence. Since then, PCR has been used in more than 275,000 scientific publications.

This technique has been applied in different areas due to its versatility, specificity and sensitivity. Methods based on the polymerase chain reaction (PCR) have been proved to be very efficient and applicable in foods.

Accordingly, PCR has been successfully used for microorganism identification , for the detection of ingredients of food products, e.g cereal, vegetables, animal and fish species. Detection and identification of pathogens like Shigella sonnei, S. Flexneri, S. boydii, S. dysenteriae, Salmonella paratyphi A & B, Aeromonas hydrophila, Staphylococcus aureus, Clostridium perferinges, type A Clostridium botulinum in canned peas, corn and lima beans, Vibrio cholera in seafood, enteric virus on oysters, toxigenic S. aureus in beef, pork, cheese and milk, enteroinvasive E. coli in raw milk and Listeria monocytogenes in poultry have been successful using PCR.

The minimum detectable level of organisms, type of interferences associated with each food sample, precautions to be taken in sample preparation, need for pre-enrichment and correlation of the results with the conventional methods are all important considerations in standardization of the methods.

PCR is a simple, versatile, sensitive specific and reproducible assay. It consists of an exponential amplification of an DNA fragment, and its principle is based on the mechanism of DNA, replication in vivo: dsDNA is denatured to ssDNA, duplicated and this process is repeated along the reaction according to the formula.
Polymerase chain reaction

Food analysis using DNA probes

A DNA probes is a fragment of DNA. Each probe can be used to reveal the presence of a specific ‘target’ DNA sequence within the rest of the organism’s DNA.

DNA probes have been highly successful in ‘fingerprinting’ people, in forensic medicine and in clinical medicine, and in isolating specific genes from DNA libraries.

These techniques should be adaptable to answering a number of questions related to food safety and quality.

Probe based methods have been developed for detection and enumeration of foodborne pathogens like Salmonella, Staphylococcus species, Listeria spp and hepatitis A virus.

In this method colonies that have grown up in selective agar are transferred to a membrane, which is then treated with reagents that lyse the cells. The denatured DNA is released and sticks to the membrane, which is then reacted with the DNA probe.

Lactic bacteria in wines and grape must have been detected by using DNA probes. L monocytogenes in artificially inoculated soft cheese and ground chicken have been detected using a hydrophobic grid membrane filter DNA probe.

Probes for typical plant pathogens like avocado sun blotch viroid, potato tuber spindle viroid and Erwinia amylovora can replace lengthy bioassays and give a clear-cut diagnosis of these plant diseases.

DNA probe technology has the potential to significantly shorten the sample analysis time in food borne pathogen detection. However, the continued requirement for a culture enrichment period will reduce the effectiveness of the technology, since it is the rate limiting step in the process.

The DNA probe must be highly selective for the organism to be determined. Selectivity is a function of the size of the DNA fragment used, the homogeneity of the DNA in terms of purity of strain, and the associating conditions.
Food analysis using DNA probes

Biosensors in food analysis

Biosensors are analytical devices based on the direct spatial coupling containing a biological active material recognition element like an enzyme, antibody or microbe coupled to a chemical or physical transducer including and electrochemical (electrode), mass (piezoelectric crystals, or acoustic wave devices), optical (optrodes) and thermal detector.

It converts the biochemical signal into a quantifiable electrical signal.

As biological recognition elements, enzymes and antibodies certainly still dominate in the commercial market of biosensors.

Biosensors have a tremendous potential for the detection of microbial contamination in foodstuff and the microarray technology leads to simultaneously detection of different pathogens.

Among the many advantages offered by biosensors in food quality control and processing, these devices can be used for online-line processes or discrete sampling.

A wide range of enzyme biosensors for applications in food analysis have been describe, e.g. for detection of glucose, carbohydrates., ethanol, starch and phenol. In contaminant analysis, enzyme biosensors have largely been used for organophosphorus and carbamate pesticide and herbicide analysis.

The more remarkable characteristics of biosensing devices which converts them in unique attractive options to compete with other technologies in the agriculture and food market including: high selectivity, high sensitivity, short time of analysis, ability to be included in integrated systems, automation easiness, capability of response in real time, versatility allowing the design of ‘a la crate’ devices and low cost.

Biosensors in food analysis