Adapting lipidomic sample processing methods for boars housed in commercial settings

Authors: Mills, Kayla; MINTON, AMANDA - Acufast; FERREIRA, CHRISTINA - Purdue University

Submitted to: Translational Animal Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/13/2024
Publication Date: N/A
Citation: N/A

Interpretive Summary: Markers indicative of fertility status in commercial boars would be valuable to the swine industry as the inclusion of subfertile boars into breeding programs results in a loss of efficiency and causes negative economic consequences for the farm. Emergent technology in mass spectrometry known as multiple reaction monitoring (MRM) profiling can distinguish between different fertility phenotypes across species (men and female pigs) making it a valuable approach for fertility marker discovery. However, this technology has not yet been utilized in boars for fertility status prediction. In addition, samples for this type of analysis are transported in liquid nitrogen or dry ice and are not commonly available at commercial boar studs. Therefore, the objective of this study was to determine if lipid sample processing at room temperature using methanol could be used in lipid samples sourced from commercial boars. We observed that room temperature sample processing did not compromise the total number of lipids detected in the boar’s ejaculate using MRM profiling and results were comparable to other studies where semen samples were frozen in liquid nitrogen. We also noted that boars have a distinct ejaculate lipidome (cellular fat composition) profile based on major plasma membrane lipid classes. These results indicate that this method of sample processing for MRM profiling is effective and can be used in commercial settings for future fertility marker discovery.

Technical Abstract: Multiple reaction monitoring (MRM) profiling is a sensitive method of lipid screening that has the capability to distinguish between different fertility phenotypes in gilts. However, MRM profiling has not yet been utilized to evaluate fertility phenotypes in boars. Markers indicative of fertility status in boars would be valuable as inclusion of subfertile boars in breeding programs results in a loss of efficiency and negative economic consequences. In addition, semen samples for lipidomic analysis are transported in liquid nitrogen or on dry ice to suspend metabolic activity within the sperm cells, however, these cryopreservation techniques are not commonly available at commercial boar studs. Therefore, the objective of this study was to develop a method of sample processing for MRM profiling that suspends metabolic activity within semen without freezing the sample. Five, sexually mature boars of similar genetics enrolled in a commercial breeding program were collected for the study. Following collection, ejaculates were aliquoted into methanol to suspend metabolic activity and shipped to Purdue University overnight for lipid extraction. Lipids were extracted using the Bligh & Dyer method and MRM profiling was used for lipid screening. A total of 329 ion transitions (MRMs) related to lipids were detected with most lipids being characterized as plasma membrane lipids (74%) which were comprised of phosphatidylcholines (40%), ceramides (16%), phosphatidylethanolamines (11%), and phosphatidylserines (7%). acylcarnitines (AC) represented approximately 8% of the ejaculate lipidome. Hierarchical cluster and principal component analysis revealed that boars have a distinct ejaculate lipidome profile based on major plasma membrane lipid classes. In addition, we observed that one boar was unique in his abundance of AC which are related to progressive motility and sperm cell metabolism. These results indicate that this method of sample processing for MRM profiling is suitable to be used to evaluate the lipidome of ejaculates from commercial boars and has potential for broader applications across different livestock species in commercial environments.