title: Colloid centrifugation of fresh stallion semen before cryopreservation decreased microorganism load of frozen-thawed semen without affecting seminal kinetics.
abstract: Freezability of equine semen may be influenced by microorganism population of semen. The objective of this study was to verify the effect of single-layer density gradient centrifugation (SLC) of fresh semen before cryopreservation on semen's microbial load (ML) and sperm cells kinetics after freezing-thawing. For that, one ejaculate was collected from 20 healthy stallions and split into control (C) samples (cryopreserved without previous SLC) and SLC samples (subjected to SLC). Semen cryopreservation was performed according to the same protocol in both groups. Microbial load of each microorganism species and total microbial load (TML) expressed in colony-forming units (CFU/mL) as well as frozen-thawed sperm kinetics were assessed in both groups. Additional analysis of the TML was performed, subdividing the frozen-thawed samples in "suitable" (total motility ≥ 30%) and "unsuitable" (total motility < 30%) semen for freezing programs, and comparing the C and SLC groups within these subpopulations. After thawing, SLC samples had less (P < 0.05) TML (88.65 × 10(2) ± 83.8 × 10(2) CFU/mL) than C samples (155.69 × 10(2) ± 48.85 × 10(2) CFU/mL), mainly due to a reduction of Enterococcus spp. and Bacillus spp. A relationship between post-thaw motility and SLC effect on ML was noted, as only in samples with more than 30% total motility was ML reduced (P < 0.05) by SLC (from 51.33 × 10(2) ± 33.26 × 10(2) CFU/mL to 26.68 × 10(2) ± 12.39 × 10(2) CFU/mL in "suitable" frozen-thawed semen vs. 240.90 × 10(2) ± 498.20 × 10(2) to 139.30 × 10(2) ± 290.30 × 10(2) CFU/mL in "unsuitable" frozen-thawed semen). The effect of SLC on kinetics of frozen-thawed sperm cells was negligible.
authors.affiliation: Instituto de Ciências Biomédicas Abel Salazar, University of Porto, Portugal; Centro de Estudos de Ciência Animal (CECA/ICETA), University of Porto, Porto, Portugal.
authors.forename: T
authors.initials: T
authors.lastname: Guimarães
authors.affiliation: Instituto de Ciências Biomédicas Abel Salazar, University of Porto, Portugal; Centro de Estudos de Ciência Animal (CECA/ICETA), University of Porto, Porto, Portugal.
authors.forename: G
authors.initials: G
authors.lastname: Lopes
authors.affiliation: Instituto de Ciências Biomédicas Abel Salazar, University of Porto, Portugal.
authors.forename: M
authors.initials: M
authors.lastname: Pinto
authors.affiliation: Instituto de Ciências Biomédicas Abel Salazar, University of Porto, Portugal; Centro de Investigação em Biodiversidade e Recursos Genéticos (CIBIO), University of Porto, Porto, Portugal.
authors.forename: E
authors.initials: E
authors.lastname: Silva
authors.affiliation: Instituto de Ciências Biomédicas Abel Salazar, University of Porto, Portugal.
authors.forename: C
authors.initials: C
authors.lastname: Miranda
authors.affiliation: Fundação Alter Real, Alter do Chão, Portugal.
authors.forename: M J
authors.initials: MJ
authors.lastname: Correia
authors.affiliation: Private Practitioner, Évora, Portugal.
authors.forename: L
authors.initials: L
authors.lastname: Damásio
authors.affiliation: Instituto de Ciências Biomédicas Abel Salazar, University of Porto, Portugal; Centro de Investigação em Biodiversidade e Recursos Genéticos (CIBIO), University of Porto, Porto, Portugal.
authors.forename: G
authors.initials: G
authors.lastname: Thompson
authors.affiliation: Instituto de Ciências Biomédicas Abel Salazar, University of Porto, Portugal; Centro de Estudos de Ciência Animal (CECA/ICETA), University of Porto, Porto, Portugal. Electronic address: arocha@mail.icav.up.pt.
authors.forename: A
authors.initials: A
authors.lastname: Rocha
mesh_terms: D000818:Animals; D001419:Bacteria / Q000302:isolation & purification; D002499:Centrifugation, Density Gradient / Q000379:methods / Q000662:veterinary*; D003102:Colloids; D015169:Colony Count, Microbial; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary*; D006736:Horses; D007700:Kinetics; D008297:Male; D012661:Semen / Q000382:microbiology*; D012662:Semen Preservation / Q000379:methods / Q000662:veterinary*; D013081:Sperm Motility / Q000502:physiology
chemical_list: D003102:Colloids
keywords: Androcoll-E; Bacteria; Cryopreservation; Semen; Stallion