2.4. Experimental design
Before working on real contaminated semen samples,
the applicability of MALDI-MS to the bacterial screening
of bull sperm was tested. In this pilot experiment, Escherichia
coli ATCC 25922 and Staphylococcus aureus ATCC
29213 strains were cultured on BHI broth at 38 C for 24
hours. After growing, each bacterial pellet was diluted
in 100 mL of water, and 50 mL of the obtained solution
were added to 50 mL of bull semen. The spiked mix was
carefully submitted to inactivation and proteins extraction
as reported in the Materials and methods section. The
Escherichia coli and Staphylococcus aureus were properly
identified by Biotyper 3.0 with a high score (>2.3). This
proved that the semen proteins do not interfere with the
MS identification.
Thirty semen samples and 570 culture medium samples
(204, 172, and 194 for IVFm, IVCm, and IVMm, respectively)
collected at the commercial setting for bovine embryo IVP
were examined in this study.