Browsing by Subject "spesifisyystestaus"

Water moulds belonging to genus Saprolegnia are known to be opportunistic fish pathogens and cause a deadly disease called saprolegniosis. Saprolegniosis is found in both wild and farmed fish as fish farms use natural waters as their fish-farming waters. Majority of cases of saprolegniosis found in fish farms are caused by Saprolegnia parasitica which is currently identified from dead fish by microscopic methods. This master's thesis was done for Finnish Food Authority. The aim of this thesis was to compare quantitative PCR (qPCR) methods in detection of S. parasitica and to evaluate the applicability of the best method. qPCR allows for better species identification than the microscopic method and it also enables the quantification of water mould DNA. The study was performed for three qPCR methods amplifying the gene region encoding S. parasitica aquatic mould 28S rRNA. Of the three methods, the best was selected based on specificity and sensitivity tests. The best method was used to detect and quantify S. parasitica in water samples from four Finnish fish farms. By this, the suitability of the method for water samples was confirmed. Also, data about the occurrence of S. parasitica water mould on fish farms was also collected. Based on the studies performed, the Roc-qPCR method based on the method developed by Rocchi et al. (2017) was selected as the best. The Roc-method was highly specific in identification of S. parasitica. With Roc- method, very low concentrations of S. parasitica DNA was quantified from fish farm water samples within good confidence. DNA amount of S. parasitica was statistically significantly higher in the effluent waters of fish farms, in which S. parasitica DNA levels were highest when water mould was detected in the fish farm fish. These results confirmed the notion that S. parasitica increases in fish. This increase of spores could be reliably detected by the Roc method. In the study, water temperature was not found to correlate with S. parasitica DNA, which may be due to the lack of data in the study. On the other hand, the occurrence of water mould is influenced by several other factors in addition to temperature, which were not considered in this study. Based on the results of this study, the Roc-qPCR method is a specific and sensitive method for the detection of S. parasitica from water samples. Roc-method is a potential method for use when designing of saprolegniosis control measures in fish farms. Further research is needed about the factors associated with the presence of S. parasitica in fish farms.