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The Use of Ceftiofur Sodium in the Extension and Cooled Storage of Equine Semen: Its Effects on Motion Characteristics, pH, Osmolality, and Bacterial Growth

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Title: The Use of Ceftiofur Sodium in the Extension and Cooled Storage of Equine Semen: Its Effects on Motion Characteristics, pH, Osmolality, and Bacterial Growth
Author(s): Naughton, Katherine G.
Advisor(s): Clark, Sherrie G.
Department / Program: Vet Clinical Medicine
Discipline: VMS-Veterinary Clinical Medcne
Degree Granting Institution: University of Illinois at Urbana-Champaign
Degree: M.S.
Genre: Thesis
Subject(s): Spermatozoa equine antibiotics semen extender motion characteristics bacteriology pH level osmolality
Abstract: During semen collection in the stallion, contamination of the ejaculate occurs secondarily to the normal microflora present on the external genitalia. These bacteria and their metabolites can negatively affect spermatozoal motility and viability during cooled storage. Therefore, the addition of antibiotics to the semen extender is warranted to reduce bacterial load and prevent further growth. Naxcel® (ceftiofur sodium, Pfizer Inc., New York, NY) is a third generation cephalosporin antibiotic with high antibacterial activity and a broad resistance to β-lactamases. The purpose of the current study was to evaluate varying levels of ceftiofur sodium added to a skim milk-glucose based semen extender in comparison to control (antibiotic-free extender) and other commonly utilized antibiotics on spermatozoal motion characteristics, semen osmolality and pH, and bacterial growth during cooled storage for 48 hours. Semen was collected and pH measurement, osmolality measurement, and aerobic culture were undertaken for each ejaculate (total of thirteen from three stallions). Each ejaculate was then divided and extended in a skim milk-glucose based semen extender (Har-VetTM Semen Extender, Har-VetTM, Spring Valley, WI) without antibiotic (control) or with one of seven different antibiotic groups: ceftiofur sodium at 250µg/ml (CEFT250); ceftiofur sodium at 500µg/ml (CEFT500); ceftiofur sodium at 1,000µg/ml (CEFT1000); ceftiofur sodium at 2,500µg/ml (CEFT2500); combination of amikacin sulfate and potassium penicillin G at 1,000µg/ml and 1,000IU/ml, respectively (AMKPCN); gentamicin sulfate at 1,000µg/ml (GENT); and ticarcillin disodium at 1,000µg/ml (TICAR). Extended semen was cooled and stored in a semen-transport container (EquitainerTM, Hamilton Research Inc, South Hamilton, MA) at approximately 5°C. Motility measures using computer-assisted semen analysis, pH measurements, and osmolality measurements were performed at 0, 24, and 48 hours after collection. Aerobic culture was performed for the eight extended groups after 24 and 48 hours of storage. Motion characteristics measured included: total motility (MOT), progressive motility (PM), curvilinear distance (DCL), average path distance (DAP), straight line distance (DSL), curvilinear velocity (VCL), average path velocity (VAP), straight line velocity (VSL), linearity (LIN), straightness (STR), wobble (WOB), beat cross frequency (BCF), and amplitude of lateral head displacement (ALH). Ceftiofur sodium had a dose-dependent effect on DCL, VCL, and ALH, with higher concentrations (1,000µg/ml and 2,500µg/ml) yielding increased values in comparison to controls (F test, P<0.05; post-test comparison, P<0.0071). There was also a significant effect of extender on STR (F test, P<0.05) with higher concentrations of ceftiofur sodium tending to decrease values in comparison to controls (P<0.028). The changes in motion characteristics are consistent with a trend toward spermatozoal hyperactivation. Stepwise, multiple regression analysis revealed pH was the strongest indicator for the increased motion characteristics, but it was only a minor predictor. There was no significant effect of extender noted for MOT, PM, DAP, DSL, VAP, VSL, LIN, WOB, and BCF. There was an effect of time on MOT and PM with parameters decreasing significantly over time (F test, P<0.05). Time also had an effect on DAP, DSL, VAP, VSL, LIN, STR, and WOB (F test, P<0.05) with most values decreased significantly after 48 hours of storage. There was a significant effect of extender group on pH (F test, P<0.05). Overall, pH was significantly decreased for the CEFT2500, AMKPCN, and TICAR groups and significantly increased for the GENT group in comparison to control. Time also had a significant effect on pH, with values increasing during storage. Neither extender group nor time had a significant effect on osmolality. The most frequent isolates from the ejaculates were Corynebacterium spp., coagulase-negative Staphylococcus spp., and Pseudomonas spp. with each stallion tending to grow a consistent “normal flora”. All treatment groups grew bacteria at some point during the experiment except the amikacin/penicillin group after 48 hours of storage. Corynebacterium spp. was the most common isolate in the control groups at both 24 hours and 48 hours of storage. The AMKPCN extender group provided the most effective antimicrobial control with growth noted in two of the thirteen collections. The CEFT250 and TICAR extender groups provided the least effective antimicrobial control with growth noted in ten and eight of the thirteen collections, respectively. The CEFT1000 extender group provided acceptable antimicrobial control with growth noted in five of the thirteen collections. The most significant finding of the current study was the dose-dependent relationship between ceftiofur sodium and increases in motion characteristics associated with hyperactivation of spermatozoa. While pH was the strongest predictor for these changes of the measured variables, it was only a minor indicator. Furthermore, hyperactivation of spermatozoa is associated with an increase in intracellular pH and a decrease in solution pH was noted with high concentrations of ceftiofur in comparison to control in the current study. Further study is indicated for the evaluation of ceftiofur sodium in the extension and cooled storage of stallion semen and its relationship to hyperactivation of spermatozoa.
Issue Date: 2011-01-14
URI: http://hdl.handle.net/2142/18422
Rights Information: Copyright 2010 Katherine G. Naughton
Date Available in IDEALS: 2011-01-14
Date Deposited: December 2
 

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