ARTIFICIAL INSEMINATION IN FARM ANIMALS - Phenix-Vet

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148Artificial Insemination in Farm Animalsand 95.8% correctly identified as satisfactory according to the criteria of 200×10 6 motilesperm/mL or 180×10 6 motile, normal sperm/mL, respectively. In our quantitative test, themaximum overall accuracy of 92.9% confirmed the high discrimination power for boarsemen according to a criterion value of SI at 180×10 6 sperm/mL.3.3 Stability of butanol extracts in terms of A610After developing the assay, we wondered if it was possible to measure the absorbance at alater date, i.e. within a day or even a week of the assay. A satisfactory level of agreementwould indicate that the modification was successful, which in turn would greatly enhancethe usefulness of the assay as it could then be performed even if a spectrophotometer wasnot immediately available.We measured the A 610 of each butanol extract of 112 samples on days 0, 1 and 7 after storageat 4C. The differences were obtained between A 610 at day 0 (A0) and day 1 (A1) andbetween days 0 (A0) and 7 (A7).The limits of agreement were calculated as follows: limits = d 2sdiff , where d is themean of differences for all the samples, and sdiff is the standard deviation of the differences.2sdiff is also named the reproducibility coefficient. Differences between absorbances (A1 -A0) were plotted against their average value ((A1 + A0)/2) for each sample. Satisfactoryagreement is achieved when minimum 95% of the absolute differences are less than thereproducibility coefficients (Petrie & Watson, 1999).It is necessary to establish that a method is repeatable before comparing two measurementsfor reproducibility (Petrie & Watson, 1999). The within-run coefficient of variation,calculated as 7.79 4.06 %, confirmed satisfactory repeatability of the method, therefore thepairs of measurement of A610 were allowed to compare. The differences betweenmeasurements (A 610 ) immediately after centrifugation (day 0) and after 7 days were plottedagainst the average of these values. 95.54 % of differences lie within the limits of agreement(Fig. 13).4030difference in A 61020100-10-20upper limit of agreementaverage of differenceslower limit of agreement-300 100 200 300 400 500 600average A 610Fig. 13. A 610 values as a function of time of measurementMeasurements obtained on the day of performing the test and the measurements after 24hours also agree; 99.1 % of the differences lie within the limits of agreement (data notshown). The results obtained leading to the conclusion that we can measure A 610 of butanol
Evaluation of a New Method and Diagnostic Test in Semen Analysis 149extracts within 7 days from the day of test performing, confirming a great practical value ofthe method.In a diagram of the differences between absorbances plotted against their average, thescatter of the points is random (Fig. 13) indicating, that the size of the discrepancy betweenthe two absorbance is not related to the size of the absorbance. More than 95 % of absolutedifferences were less than the reproducibility coefficients in both cases of testing the stabilityof butanol extracts. This is a satisfactory agreement, therefore we can measure theabsorbance immediately after performing the test or within 7 days of that time. Thereforethe test is useful even if spectrophotometer is not available at the location of semenevaluation. The results obtained leading to the conclusion that we can measure A 610 ofbutanol extracts within 7 days from the day of test performing, confirming a great practicalvalue of the method.4. ConclusionsThe usefulness of sperm counting is greatly enhanced by the simplicity of determination byphotometer (Photometer SDM5, MiniTüb, Germany) in on-farm AI laboratories. The use ofphotometer for determining sperm concentration would, therefore, be of benefit also tolivestock producers in evaluating the quality of boar semen.The resazurin reduction assay was shown to be a reliable, easy-to-perform test that requiresno sophisticated equipment. It was demonstrated that the results of the assay can be used toselect semen samples with minimum requirements of sperm concentration, motility andnormal morphology, which are all combined in sperm index. Because reproductiveperformance depends on metabolic processes, the assessment of metabolic rates ofspermatozoa could provide even better or more complete information about semen qualitythan other tests. It allows the concentration of active spermatozoa to be determined, andmay provide a better means of evaluating semen quality than assessing the characteristics,mentioned above, independently. Expressing the latter in semen evaluation is complex,although fertility results from insemination with evaluated semen could provide a goldstandard of fertilizing capacity. Additional research is required for relevant and validinformation about replacing or updating the methodology of semen evaluation.5. AcknowledgementsThis work was supported by the Slovenian Ministry of Higher Education, Science andTechnology, programme group ''Endocrine, immune, nervous and enzyme responses inhealthy and sick animals'' (P4-0053).Special thanks go to author's collegues who contributed to the research work, presented inthis chapter: Janko Mrkun, DVM, PhD, Marjan Kosec, DVM, PhD, Janez Kunc, DVM, MSc,Maja Zakošek Pipan, DVM.6. ReferencesAnzar, M., Kroetsch, T., Buhr, M.M. (2009). Comparison of different method for assessmentof sperm concentration and membrane integrity with bull semen. J Androl, 30, 6, pp.(661-668)
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ARTIFICIAL INSEMINATION IN FARM ANIMALS - Phenix-Vet

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