Thesis

Effect of mycotoxin, deoxynivalenol, in aquaculture reared rainbow trout (Oncorhynchus mykiss) metabolism

Citation

Mendes Goncalves R (2020) Effect of mycotoxin, deoxynivalenol, in aquaculture reared rainbow trout (Oncorhynchus mykiss) metabolism. Doctor of Philosophy. University of Stirling. http://hdl.handle.net/1893/31248

Abstract
The decreasing supply and high cost of fishmeal, as well as the sustainability issues of this finite raw material, will continue to push the industry to concentrate their efforts on finding alternative sources of protein to substitute fishmeal in aquafeeds. From all the possible alternatives, e.g. animal by-products, fishery by-products, single-cell protein and the recent insect meal, plant-based meals seem to be one of the major contributors to reduce reliance upon marine sources. However, when considering plant-based meals for aquafeeds it is commonly agreed that one of the negative aspects is the presence of anti-nutritional factors (ANF’s; e.g. cyanogens, saponins, tannins, etc.) that are detrimental to fish and shrimp (Krogdahl et al., 2010). Although there are processes to remove or inactivate many of these ANF’s, the same does not apply for mycotoxins which are reasonably stable to processing conditions (Cheli et al., 2013). The effects of mycotoxins in fish and shrimps are diverse, varying from immunosuppression to death, depending on toxin-related (type of mycotoxin consumed, level and duration of intake), animal-related (animal species, sex, age, general health, immune status, nutritional standing) and environmental related (farm management, biosecurity, hygiene, temperature) factors. Therefore, it is often difficult to trace observed problems back to mycotoxins. This thesis main goal is to increase the awareness of mycotoxin contamination in aquafeeds and its consequences to aquaculture species, especially characterizing the impact of deoxynivalenol in rainbow trout (Onchorhynchus mykiss). In Chapter 1 the known mycotoxin occurrence and co-occurrence in aquaculture finished feeds are described and critically reviewed by correlating the extent of mycotoxin contamination in aquaculture feeds to its impact in aquaculture species. This chapter also contributes to understanding the risk of mycotoxin carry-over to aquaculture seafood products. Additionally, this chapter targets to expose the scientific community, the regulatory authorities and the aquaculture industry, to the main challenges and myths that the industry faces in developing mycotoxin management strategies. Chapter 2 presents the results the impact of deoxynivalenol in rainbow trout and the difficulties to diagnose DON ingestion. Here was explored two different DON contamination scenarios, i.e., the effect of short-term feeding of high levels of DON and the effects of long-term feeding of low levels of DON (a more common scenario in aquaculture industry). Was concluded that the ingestion of DON by trout over shortterm periods at high dosages (50 days; 1,166 µg kg -1 and 2,745 µg kg-1 ) impactsgrowth performance, especially feed intake, with minor or variable biochemical changes in trout blood and histopathological changes. In this case, some fish did exhibit clinical symptoms (i.e., anal papilla), which could be attributed to the DON treatment (reported for the very first time). This chapter also reports, for the first time, the effects of the long-term exposure of rainbow trout to low concentrations of DON (168 days; 367 µg kg -1 DON). Ingestion of DON in the long-term study was asymptomatic; however, the fish started to reduce their growth performance 92 days after ingesting DON. Such low contamination levels, which might be unnoticed by farmers, may have economic consequences for aquaculture. Here is concluded that the short-term effect of DON ingestion cannot be extrapolated to other contamination scenarios, e.g., long-term exposure. In Chapter 3 aimed to elucidate the impact of DON on rainbow trout and study the reasons behind the apparent lack and/or high variability of clinical signs during DON ingested (as reported in chapter 2). With this experiment, we further confirmed that ingestion of DON by rainbow trout is mainly characterised by impaired growth performance and reduced feed intake, with the total absence of any visible clinical signs (up to 11,412 ± 1,141 µg kg -1). Proteolytic enzyme activities (pepsin, trypsin and chymotrypsin) in trout were altered by DON ingestion. The impact of DON on the abundance of specific measured mRNA transcripts was unexpected with higher expression levels for insulin-like growth factors, igf1 and igf2, which are directly related to elevated insulin levels in plasma. This can also in part be influenced by the trypsin activity and by npy, given its higher mRNA expression levels. The apparent digestibility of dry matter, protein and energy was not affected by dietary levels of DON, however, nutrient retention, protein, fat and energy retention were significantly affected in animals fed DON. Adenylate cyclase-activating polypeptide (PACAP) expression seems to play an important role in controlling feed intake in DON fed trout. In the present study, we have shown for the first time that DON is metabolized to DON-3sulfate in trout. DON-3-sulfate is much less toxic than DON, which helps to explain the lack of clinical signs in fish fed DON. Being a novel metabolite identified in trout makes it a potential biomarker of DON exposure. Is suggested that the suppression of appetite due to DON contamination in feeds might be a defence mechanism in order to decrease the exposure of the animal to DON, therefore reducing the potential negative impacts of DON. In Chapter 4, this thesis further explored the current knowledge of DON toxicokinetics and rainbow trout DON metabolism, accessing the organs assimilation rates, excretion and possible biotransformation kinetics. Here we found that hour after tube-feeding, [3H]-DON was detected in the liver samples of fish, indicating a rapid absorption of DON. In the first hour, [ 3H]-DON was present in the GIT (20.56 ± 8.30 ng). However, 6.19 ± 0.83 ng was also detected in the water at this sampling point. The fast excretion of [3H]-DON (faster than the average trout gastric emptying time) suggests that DON, as a water-soluble compound, is excreted with the liquid phase of the chyme. The presence of [3H]-DON in the GIT was stable during the first six hours. This long residence time of DON in the GIT may compromise the health of the GIT and favour absorption. Our data suggest that an effective DON detoxifying method should have a period of action of ≤ 6 h. Furthermore, as most of the excretion can be expected to happen after six hours, the detoxification should be irreversible at GIT conditions. Results of this PhD study contributes to better understand the importance and the risk of mycotoxin occurrence and co-occurrence in aquaculture finished feeds, highlighting concerns regarding the unvalued risk of mycotoxin carry-over to aquaculture seafood products. Here is highlighted that long-term exposure to DON is normally asymptomatic which might be unnoticed by farmers, however, representing economic consequences for aquaculture (reduce growth performance). Suppression of appetite due to DON contamination in feeds seems to be a defence mechanism in order to decrease the exposure of the animal to DON, therefore reducing the potential negative impacts of DON. The biotransformation of DON to DON-3-sulfate helps to explain the lack of clinical signs in fish fed DON and may be used as a novel biomarker of DON exposure.

Keywords
Mycotoxins; Aquaculture; deoxynivalenol; Rainbow trout (Onchorhynchus mykiss); anti-nutritional factors; plant-based meals

StatusUnpublished
FundersBiomin Gmbg
SupervisorsSimon Mackenzie and Amaya Albalat
InstitutionUniversity of Stirling
QualificationDoctor of Philosophy
Qualification levelPhD
Publisher URLhttp://hdl.handle.net/1893/31248