BLT Stages

This study is examining the effect of food quality (in terms of fatty acid composition) on the survival of harpacticoid copepods and on their fatty acid composition (and possibility bioconversion of short-chain fatty acids to long-chain PUFA). This is achieved by examining the effect of different diets on fatty acid composition and copepod survival. Long-chain PUFA are fatty acids that have twenty or more carbon atoms and have three or more double bounds. These fatty acids are important for the health and stability of the ecosystem.

Previous research has shown that bioconversion of fatty acids by harpacticoid copepods is possible. Research is still being performed to better understand this process. On the copepod Platychelipus littoralis, there has not yet been much research done on the long-chain PUFA bioconversion in copepods. This harpacticoid copepod is used as test organism in this research. This research is essential because this species is an important primary consumer that feeds on primary producers. The research aims to better understand how energy is transferred through the food web. Fatty acids are used to estimate this energy transfer between organisms.

The copepods are fed during eight days with nothing (negative), Saccharomyces cerevisiae, Nitzschia species and Dunaliella tertiolecta. These food sources are chosen because they have a different fatty acid composition. The fatty acid profiles of these diets are also compared with the fatty acid profiles of Platychelipus littoralis that came directly from the field. The information about mortality was measured by counting the copepods at the end of the treatments. Fatty acids profiles of the different diets and the copepods that were fed with it were compared. First, the fatty acids were extracted from the copepods and methylated. The samples were then analysed using a gas chromatography coupled to a flame ionisation detector for quantification and to a mass spectrophotometer for identification. Possible difference between the copepods, fed with different diets, were analyzed with one-way ANOVA or Kruskal-Wallis tests and post-hoc Tukey HSD tests or Dunn tests.

In this study, the copepods survival was always very high, regardless of diet. In the obtained fatty acid profiles, it is remarkable that Platychelipus littoralis fed with the negative diet (without food) still contained a lot of DHA. This may be caused by the large amount of reserve fatty acids in the copepods. It is also remarkable that the copepods fed with the Dunaliella tertiolecta diet contain a high relative amount of 20:3w3 (0.23 %) while this is not present in the diet itself. It can therefore be concluded that Platychelipus littoralis is most likely using the side branch from 18:3w3 to form long-chain fatty acids.

The aim of this study is to provide scientific based advice regarding the diet of shrimp to the fisheries in order to support a sustainable way of fishing. The information about the diet was obtained by means of fatty acid analysis. Fatty acid profiles of the shrimps of different life stages and different areas and seasons were compared. During the extraction step a methylation protocol was used to make the fatty acids volatile. Gas chromatography coupled to a mass spectrometer was used for their identification and a flame ionization detector for quantification. An internal standard was utilized to correct for losses during the extraction protocol and to calculate the concentration of the fatty acids. Possible differences between life stages, areas and seasons were statistically tested with multivariate ANOVA and univariate Kruskal-Wallis tests.

The obtained results show that shrimp samples from the pristine area display more diversity in their diet than those originating from the disturbed area, but no significant difference was observed. The absolute total fatty acid content was significantly higher in juveniles than in adults during the long dry season and the small rainy season. Other seasons showed no significant difference. A significant higher amount of oleic acid (18:1ω9) was observed in adult shrimp, compared to juveniles. Significant differences between different seasons were observed, especially the small dry season differed from the other seasons in multiple aspects such as lower total fatty acid concentration, lower DHA, lower omega-3 and omega-6 fatty acids and higher oleic acid.

Juvenile shrimp contained more fatty acids than adults, as they need more energy to grow. The higher amount of oleic acid in adult shrimp can be explained by the need to lower cholesterol. The diet of the shrimp was significantly different during the small dry season possibly caused by a difference in food availability. The quality of the shrimp during the small dry season is lower due to a lower concentration of omega-3 and omega-6 fatty acids. Therefore fishing during the small dry season is not recommended but further research is needed to provide the fisheries with useful information in order to gain a sustainable way of fishing.

Marine invertebrates are organisms that form an important food source for other marine organisms such as fish. These fish eventually end up on a plate for us, humans. A big problem with marine invertebrates such as copepods is that they are subjected to sorts of stress due to climate change and anthropogenic factors. These stressor factors can be measured on many levels and in many ways. For instance, the appearance of species growing up with stress can be measured. Further, quantifying the densities of species is a way of analyzing the condition of the population of species. In this research we applied a more functional approach by analyzing fatty acid content and by looking at the development time of juveniles to investigate stress effects on marine organisms.

We used both field data and experimental research in this thesis. The first part of my investigation is to see whether marine invertebrates from the coastal area of Peru are still safe to eat and if they will preserve their food quality (in terms of omega-3-fatty acids) in the future as the environment is changing. For the species from Peru we aimed to profile their fatty acids and to contrast the profiles for stressed organisms to control organisms without stress. In a second part of my investigation, the effect of stressors to copepods in the Belgian part of the North Sea was analyzed by means of a lab experiment. Earlier research of my mentor Yana Deschutter showed that PCBs might have an effect on copepods species in the Belgian part of the North Sea (Deschutter, Everaert, De Schamphelaere, & De Troch, 2017). To test this, we measured the mortality and time of transitioning to copepodite stage one of copepods submitted to an elevated temperature and exposed to spiked solutions of PCBs.

The field samples from Sechura Bay, Peru were collected in 2016 (by mentor I. Loaiza in the frame of his PhD research) from environments with different levels of pollution and analyzed in February and March 2018. These samples consisted of six species that are listed further in the thesis. For the fatty acid profiling of the species from Peru, the extracted fatty acids were injected in a gas chromatograph – mass spectrometer. All data was statistically stored and analyzed with ‘Agilent Technologies Chem Data Analysis’. The main results for the field samples of Peru is that samples from the Illescas Reserve have better food quality than samples taken from Sechura Bay.

The lab samples were collected in April 2018. These consisted of one copepod species (Temora longicornis) from the Belgian part of the North Sea. For this lab research, the time between the hatching of a nauplius and the molting to copepodite stage one was the aim to investigate. Unfortunately, no results were found because the eggs did not hatch.

Copepods are planktonic organisms that are very common in oceans, rivers and lakes. They form an important source of food for other marine organisms like fish larvae. Copepods are very sensitive to changes in the environment. Therefore, they are good indicators in research about climate change. Many research has been done on the influence of single stressors on copepods, but there’s a lack of data concerning the influence of multiple stressors.

The aim of this research was to see whether or not there’s a seasonal and spatial difference in fatty acid profiling in two species of copepods common in the Belgian part of the North Sea: Acartia clausi and Temora longicornis. Samples were taken all year round from February 2015 until February 2016 on five different locations. There were two harbor stations (ZB1 and NP1) and three sea stations (ZG02, 120 and 700). Another way to look at the influence of stressors on copepods is to directly expose them to it. In a second experiment T. longicornis nauplii were exposed to high concentrations of polycyclic hydrocarbons (PCB’s) and different temperatures to see how the molting process would differ.

For the fatty acid profiling of the copepods, A. clausi and T. longicornis were isolated from samples. Then, a fatty acid extraction was performed to analyze the fatty acids with a gas chromatographer and mass spectrometer. Using multiple standards, the concentration of each fatty acid was calculated. The data were statistically analyzed with MDS, ANOSIM and SIMPER.

For T. longicornis a significant difference was found in fatty acid composition between samples from winter and spring and also samples from the sea and harbor. In A. clausi samples these differences were not found. For T. longicornis there were a lot of significant differences between stations. For A. clausi only three stations were significantly different from each other. Looking closer to the profiles of fatty acids docosahexaenoic acid and eicosapentaenoic acid, it was notable that the fatty acid content in T. longicornis was higher than the content in A. clausi and that for both species the content peaked during spring. Also, copepods from the sea stations had higher fatty acid content than the copepods from the harbors.

Both species showed seasonal and spatial differences. In T. longicornis these differences were more visible and significant.

For the second experiment, nine female and six male T. longicornis were kept in a jar to produce eggs. There were four replicates of these jars at 15 °C and four at 18 °C. The nauplii that were produced were transferred to a glass vial with 3 mL of an Isochrysis solution and a certain concentration of PCB’s. There were three PCB solutions: blanc, 90 percentiles, and 10 x 90 percentiles. Each concentration was also divided over the 15 °C and the 18 °C incubator. For each treatment there were 15 replicates. Every two days the nauplii were checked to see if they molted from the last nauplii stage to the first copepodite stage. The time between the hatching of the eggs and the molting from nauplii to copepodite was used as a parameter.

For the second experiment no data was obtained in time for this report.

• Het beste aantal nematoden bepalen voor het opstellen van een standaardcurve
• Nagaan of er een verschil is tussen standaardcurves opgesteld met levende nematoden en ingevroren nematoden
• Nagaan of de kwantificatie beter is wanneer adulten en juvenielen gescheiden worden