Population proteomics of hake: proteins can pinpoint the location of the fish

Tracking fish for their own good

The ever-increasing demand for food around the world is placing greater stress on fish stocks, leading to overfishing of some species. Monitoring marine stocks is by nature a difficult task, complicated further by the spread of species across different seas and oceans. This is the case for one species, the European hake (Merluccius merluccius), which is found in the Eastern Atlantic Ocean from Norway to Mauritania, as well as throughout the Mediterranean.

The hake lives near the bottom during the day but swims off at night to feed on smaller fish and squid. It has become a popular species with the consumer, so it is important to ensure that it is managed correctly, maintaining fish levels in all of its habitats. As part of this process, it is important to be able to identify the areas from which fish have been caught.

In the past, population genetic studies have shown that the genetic make up of marine fish has been moulded by many different factors, including their local marine environment. Recent studies on hake from the Atlantic Ocean led to the differentiation of northern and southern Atlantic stocks.

Now, a European study has applied population proteomics to the European hake to see if differentiation on the basis of the protein content is feasible.

Proteins place fish at particular locations

Elena Gonzalez, Jose Bautista and co-researchers from the Universidad Complutense de Madrid, ANFACO-CECOPESCA, Vigo, Spain, and the National Agricultural Research Foundation-Fisheries Research Institute, Kavala, Greece, suggested that genetic differentiation may not be entirely reliable due to the influence of many factors. Fish migration and the existence of homogenous habitats were cited as two of the potential complications.

So, they applied a proteomics approach to hake caught at one location in the Mediterranean Sea and two in the Atlantic Ocean. Fish from the North Aegean Sea in the Mediterranean, the Bay of Biscay (the Cantabrian Sea) and the Cies Islands off Vigo in northern Spain were examined.

Two types of tissue were extracted on board the fishing vessel and placed on dry ice to prevent premature degradation. Brain tissue was selected on the basis that it is less susceptible to environmental variables, whereas the opposite is true for liver tissue, which is affected by factors such as nutrition, salinity and temperature.

The proteins were extracted from all samples and subjected to two-dimensional differential gel electrophoresis with fluorescent dyes to measure the protein abundances. A total of 84 liver proteins and 145 brain proteins had different abundances between populations and mass spectrometric analysis unambiguously identified 20 liver and 35 brain proteins.

The protein expression patterns of the 55 identified proteins were visualised using principal components analysis. The first two components accounted for more than 75% of the data variance for liver proteins and 78% for brain proteins and a plot of these two components displayed clear clustering between three fish populations.

The first component discriminated between the Atlantic Ocean and Bay of Biscay fish within the liver gels, while the second discriminated between the Mediterranean Sea and Bay of Biscay fish for the brain gels. For all of the gels together, separation was still clear but there was a small amount of overlap between the populations within some of the outliers.

A second statistical method, hierarchical clustering, was also applied to the data to produce heat maps based on the levels of protein expression and these separated the populations into two groups for the liver and three for the brain.

The protein classes of the identified proteins covered cell metabolism and energy, cell signalling, protein fate, protein structure and protein transport. Within these classes, the most discriminating functions for the brain proteins were cell signalling and cell metabolism/energy whereas that for the liver proteins was protein fate.

Extending population proteomics to other fish species

So, the different levels of the liver and brain proteins were able to assign fish to one of three discrete locations with a high degree of accuracy. The hake is not a model species (like the mouse or rat) used in proteomics studies, so the results suggest that the technique could be used in a broader sense for population studies on marine fish.

This is good news for the FishPopTrace Consortium, a European project set up to examine product traceability and policy related monitoring, control and surveillance in the fisheries sector. All of the participants in the hake research study are affiliated to the organisation and their results will help to achieve one of the declared aims: to provide end-user tools in the areas of fish population analysis and fish product traceability.

The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.