Lation [5] and artificial vagina [7]. Having said that, high proportion of ejaculates obtained by way of rectal massage represented a sperm-rich fraction with minimal seminal SARS-CoV-2 3CLpro/3C-like protease plasma contribution and might not represent a `true ejaculate’. Good motility ejaculates have been characterized by bigger volume and lower sperm concentration, hence indicating greater quality ejaculates with larger percentages of sperm motility were diluted with bigger amounts of seminal plasma. As a result, even though the rectal massage strategy is practical and represents a somewhat safe choice for semen collection from captive managed bull elephants, an option method that mimics all-natural ejaculatory responses such as contributions from the accessory sex glands may possibly be warranted to enhance semen collection efficiencies in elephants. As observed herein and in a earlier study [3], a high proportion of ejaculates obtained through the rectal massage techniqueFigure two. One-dimensional gel electrophoresis of Asian elephant seminal plasma proteins. Every single column represents protein profiles from seminal plasma samples obtained from distinct ejaculates. E1, E2, and E3 denote person Asian elephant bulls. G1, G2, G3, G4, G5, and G6 denote seminal plasma samples obtained from ejaculates exhibiting good motility ( 65 tMOT). P1, P2, P3, P4, and P5 denote seminal plasma samples obtained from ejaculates exhibiting poor motility (#10 tMOT). MW: molecular weight marker. Band A was detected in 85 of seminal plasma samples from very good motility ejaculates (a representative gel is shown). Within this figure, protein bands have been visualized with Coomassie staining. doi:ten.1371/journal.pone.0071033.gPLOS One | plosone.orgLactotransferrin in Elephant Seminal PlasmaFigure 3. Protein CDCP1, Mouse (Biotinylated, HEK293, His-Avi) separation and peptide mass sequencing of Asian elephant seminal plasma. Seminal plasma proteins had been separated by two-dimensional gel electrophoresis. Selected spots had been excised and submitted for mass spectrometry (MALDI-TOF) for protein identification by de novo sequencing of MALDI-PSD tandem mass spectra. Panel A: 2D gel of seminal plasma proteins from an ejaculate exhibiting superior sperm motility ( 65 tMOT). The same sample from Panel A is also shown in Figure two, Lane G5. Cored spots from inside the train of this protein (circled; corresponding with band A in Figure two) were submitted for mass spectrometric analysis. Panel B: Example of a 2D gel of seminal plasma proteins from an ejaculate exhibiting poor sperm motility (#10 tMOT). Panel C: Mass spectrum results from MALDI-TOF. Panel D: Peptide mass sequencing identified the cored proteins as possessing homology to lactotransferrin (i = isoleucine or leucine). doi:10.1371/journal.pone.0071033.gPLOS One particular | plosone.orgLactotransferrin in Elephant Seminal PlasmaFigure four. Amino acid sequence of predicted African elephant (Loxodonta africana) lactotransferrin determined from the 29 Mammals sequencing project ( Alignment of tryptic peptide sequences determined by MALDI-PSD sequencing of SPITC derivitized peptides shown italicized. Residues marked as “i” represent either isoleucine (I) or leucine (L), which are isobaric. Asparagine 546 has probably been deamidated to its corresponding aspartic acid. doi:10.1371/journal.pone.0071033.gconsistently contained spermatozoa with poor motility. Nevertheless, premium quality ejaculates have also been obtained utilizing the same rectal massage method [3,4]. This suggests that probably other things, including seasonality and/or circulatin.