|Black grouper (Myctoperca bonaci)|
Grouper species from the genera Epinephelus and Mycteroperca are very popular food fish. Their flesh is lean and firm, but the skin is strongly flavored and should be removed before cooking. Popular species are the black grouper (Mycteroperca bonaci), Nassau grouper (Epinephelus striatus), red grouper (Epinephelus morio) and yellowmouth grouper (Mycteroperca interstitialis). Unfortunately, as a result of high demand, sporadic availability, and therefore cost, grouper is commonly substituted with Nile perch (Lates niloticus), wreck fish (Polyprio americanus), and others.
The identification of such cases of fraudulent substitution is an easy task using DNA Barcoding but the technology is currently limited to expensive molecular biology labs and trained lab technicians. Less expensive methods are indeed currently tested such as MALDI-Tof-MS or HRM but even those methods require a minimum of costly instrumentation such as a thermocycler which usually is also rather cumbersome rarely portable, and it requires trained personal. A method performed by individuals with moderate technical skills would enable much broader testing for forensic and food quality applications. Such testing could then be performed at inspection points, fish markets, and even conceivably by restaurant owners.
The molecular technique termed nucleic acid sequence based amplification (NASBA) is similar to PCR in terms of amplification of specific nucleic acid sequences via an enzymatic reaction. However, NASBA differs from PCR in some ways. The reactions are simple and quick, with an assay taking as little as about an hour including incubation. The system is based on an isothermal amplification protocol (i.e. it does not require temperature cycling) that simplifies the hardware requirements; the process also does not require a thermostable DNA polymerase or a thermal cycler, it works on RNA rather than DNA.
Researchers from Florida have now claimed a U.S. patent for a method to detect whether seafood originates from a grouper species or not:
A protocol was developed for the detection of a target seafood species, such as grouper, based or differentiation of grouper (as defined by the FDA) meat and/or tissue (fresh or frozen) from other fish meat based upon NASBA amplification of target sequences specific for grouper. RNA is extracted from a sample of muscle from a fish and amplified using nucleic acid based amplification. Specific primer sequences and molecular beacons, were designed for the amplification and detection of target sequences in the mitochondria (16 S ribosomal RNA). The resulting product of the amplified RNA is indicative of positive detection of the target fish species. The assay may be analyzed by electrophoresis, laboratory instrumentation such as benchtop luminescence detection of a molecular beacon, or through use of a field NASBA analyzer like a handheld fluorescence detection device. In specific embodiments of the invention, the assay and hardware are simplified such that semi-skilled staff can successfully differentiate grouper from other fish in situ, such as at seafood markets, restaurants, and aboard ships
Even if not a barcoding technology per se it is pretty cool as this is a first concrete step towards a handheld device that is easy to handle within a reasonable short time frame and at low cost.
Funny marginalia - when looking up NASBA I found out that the ideal temperature for the isothermal reaction condition is 41C which is exactly what a standard yogurt maker does. Something to consider for DIY fans?
h/t Bob Ward