engleski

METABOLOMIC ANALYSIS OF SERUM SAMPLES IN CANINE BABESIOSIS BY UHPLC-MS

Canine babesiosis is an important worldwide tick- borne disease caused by the intra-erythrocyte protozoal parasites Babesia canis or Babesia gibsoni. The main complications are the development of an excessive inflammatory response called SIRS and MODS. The innovative, post-genomic technologies, has led to the development of strategies aimed at identifying specific and sensitive biomarkers in biological fluids and tissues. Metabolomics is one of the most frequently applied approaches in the field of systems biology. [1] Blood and urine contains a multitude of unstudied and unknown biomarkers. The goal was to examine the difference of serum metabolome between dogs infected with B. canis and healthy dogs using mass spectrometry analysis. Serum was collected from 12 dogs infected with B. canis and 12 healthy dogs. Briefly, 25 μL serum aliquots were prepared, and 1000 μL of 1:3:1 chloroform:methanol:water was added to precipitate the proteins. The samples were allowed to cool on ice for 30 minutes, vortexed at 4˚C for 5 minutes, and then centrifuge for 3 minutes at 13.000 g at 4˚C. The supernatant (200 μL) was transferred to a screw-top vial and stored at -80˚C until analysis. Samples were analysed on Orbitrap Q-EXACTIVE MS operating in alternating positive and negative modes with mass resolution 70.000 at m/z range 70 – 1050. Analyses were performed using PiMP software. The metabolomics analysis resulted in the annotation of 1802 peaks, 158 of which showed significant differences (p < 0.05) between dogs with B. canis infection and healthy controls. 22 identified metabolites were significantly changed. The most significant metabolites were Inosine, Hypoxanthine, Choline phosphate, L-Kynurenine, and L-Cystine. The study confirmed that host pathogen interactions can be studied by metabolomics to assess chemical changes in the host, respectively that the differences in serum metabolome between dogs with B. canis infection and healthy dogs can be detected with LC-MS method.

metabolomics ; Babesia canis ; chromatography ; mass spectrometry

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