Keywords: Yersinia ruckeri, the Enteric redmouth (ERM) disease, rainbow trout (Oncorhynchus mykiss Walbaum), gills, oral vaccination, 2-thiobarbituric-acid-reacting substances, aldehydic and ketonic derivatives of oxidatively modified proteins
The aim of the study was the evaluation of the content of oxidative stress biomarkers (2-thiobarbituric-acid-reacting substances as a biomarker of lipid peroxidation, aldehydic and ketonic derivatives of oxidatively modified proteins) in the gills of rainbow trout (Oncorhynchus mykiss Walbaum) vaccinated by a vaccine against Yersiniaruckeri.
Rainbow trout (Oncorhynchus mykiss Walbaum) with a mean body mass of (107.9±3.1) g were used in the experiments. The study was carried out in a Department of Salmonid Research, Inland Fisheries Institute in Rutki (Poland). Experiments were performed at a water temperature of 14.5±0.5°C and the pH was 7.5. The dissolved oxygen level was about 12 ppm with additional oxygen supply with a water flow of 25 L per min, a photoperiod of 12 hours per day. The fish were fed with a commercial pelleted diet at an optimal level, using 12-hour belt feeders for fish. All enzymatic assays were carried out at the Department of Zoology and Animal Physiology, Institute of Biology and Earth Sciences, Pomeranian University in Słupsk (Poland). The fish were kept for 60 days after vaccination at a water temperature of 14.5±0.5°C and pH 7.5. In our study, 15 rainbow trout from unhandled control and 15 vaccinated trout were used. Two months after immunization, samples from rainbow trout were collected. The fish were captured and killed 61 days post-vaccination (n = 15 in each group). Gills were removed in situ. The organs were rinsed clear of blood with cold isolation buffer and homogenized using a glass homogenizer H500 with a motor-driven pestle immersed in an ice water bath to yield a homogenate in proportion 1:9 (weight/volume). The isolation buffer contained 100 mMTris-HCl; a pH of 7.2 was adjusted with HCl. Homogenates were centrifuged at 3,000g for 15 min at 4°C. After centrifugation, the supernatant was collected and frozen at −20°C until analyzed. Protein contents were determined using the method of Bradford (1976) with bovine serum albumin as a standard. Absorbance was recorded at 595 nm. All enzymatic assays were carried out at 22±0.5°C using a Specol 11 spectrophotometer (Carl Zeiss Jena, Germany) in duplicate. The enzymatic reactions were started by the addition of the tissue supernatant.
Our results demonstrated that immunization by the anti-Yersinia vaccine does not alter the gills of rainbow trout. Oxidative stress parameters examined in gills homogenate, i.e., lipid peroxidation as measured by the amount of TBARS, as well as aldehydic (increased by 18.9%) and ketonic derivatives of OMP (decreased by 6.5 %) were non-significantly changed (p>0.05) in gills of vaccinated fish. Thus, immunization by anti-Yersinia vaccine does not alter oxidative stress markers compared to unhandled control in the second month after immunization. Our results confirm that the vaccine against Y. ruckeri has no adverse effect on the condition and metabolism in the gills of the fish. Alterations in the content of oxidative stress biomarkers recorded in our studies are proof that the vaccine against Y. ruckeri has no negative effects.
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