| dc.description.abstract | Summary
In the past five decades anaerobic rumen fungi (ARF) have been explored more but research and information pertaining to the full extent of their capabilities is still limited compared to rumen bacteria. Anaerobic microbes work in synergy and therefore complement each other in the digestion of plant material for provision of host energy and nutrition. The rhizoidal nature of anaerobic rumen fungi allows them to embed themselves in the substrate cell wall making it difficult to extract them with commonly used extraction methods. The most common extraction method in proteomics is bead beating in the presence of a detergent, and this is effective for protozoa and prokaryotes but might be oblivious of anerobic fungi. Hence the proposal of a pretreatment by freeze grinding (FG) the plant material in liquid nitrogen prior to bead beating (BB). The hypothesis that the pretreatment would extract more fungal proteins was satisfied by the intensity of the proteins recovered. However, for more conclusive research a larger sample size is recommended. In this thesis samples six cannulated cows (from an animal feeding experiment conducted as a part of a research project named Seacow (Funded by the Norwegian Research Council, project number: 302639) ) were explored and their biological replicates were used with the two time points (2 hours and 6 hours after feeding)and two diet conditions (control and control+ Asparagopsis taxiformis (AT)) and for the two extraction methods bead beating versus freeze grinding. The dominating fungal family by protein count was Neocallimastix, with Neocallimastix sp. GF-Ma3-1 as the most detected genus and it had a higher count in freeze grinding (97) than bead beating (81). LFQ intensities showed a clear difference between bead beating and freeze grinding, where the fungal proteins had a significantly higher expression in the freeze grinding samples. The CAZyme annotation resulted in a total of eight protein groups being annotated as CAZymes. Of these, 5 CAZymes being glycoside hydrolases (GH), 2 of them were glycosyl transferases (GT)and 1 being a polysaccharide lyase (PL). These CAZymes were related to Piromyces MAGS (1), Piromyces sp. UH31-1 (2), Pecoramyces sp. F1 (1), Neocallimastix sp. WI3-B (1), and Neocallimastix sp. GF-Ma3-1 (6).I concluded that a pre-treatment step with freeze-grinding is required for sufficient extraction of anaerobic fungi from rumen samples and a larger sample size will possibly provide satisfactory elucidation of the anaerobic rumen fungi. | |
