ESHRE basic semen analysis courses 1995–1999: immediate beneficial effects of standardized training

L. Björndahl1,2,5, C.L.R. Barratt2, L.R. Fraser3, U. Kvist1 and D. Mortimer4

1 Andrology Centre, Department of Women and Child Health, Karolinska Hospital, Stockholm, Sweden, 2 Assisted Conception Unit, Birmingham Women's Hospital, University of Birmingham, Birmingham, 3 Endocrinology and Reproduction Research Group, School of Biomedical Sciences, Kings College London, London, UK and 4 Oozoa Biomedical Inc., West Vancouver, BC, Canada


    Abstract
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
BACKGROUND: Many reports have shown problems with the high variability in results of semen analyses. The Special Interest Group in Andrology (SIGA) of the European Society of Human Reproduction and Embryology (ESHRE) implemented a standardized training course which has been run in different regions of the world on more than 20 occasions since 1994. The aim of the present analysis was to investigate to what extent training resulted in any immediate effects on the variability of assessments made by different observers. METHODS: The variability in participants' results from the beginning to the end of each course was analysed in eight courses given between 1995 and 1999. RESULTS: For assessments of sperm concentration, motility, vitality and morphology, substantial improvement was seen over the duration of the course. CONCLUSIONS: A comprehensive, structured training course does lead to substantial reductions in inter-observer variability in semen analysis. This supports our contention that providing a thorough theoretical background and repeated practical training, combined with daily feedback of results, is highly effective in raising the technical skills of laboratory personnel performing semen analysis.

Key words: effect of training/semen analysis/standardization/training courses/WHO


    Introduction
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
The problem of high variability in basic semen analysis results—and thereby doubts as to their clinical value—has been described in several reports that showed large variations in results both among different investigators and for individual investigators on different occasions (Jequier and Ukombe, 1983Go; Neuwinger et al., 1990Go; Helmenhorst et al., 1995Go; Matson, 1995Go; Jörgensen et al., 1997Go; Auger et al., 2000Go; Keel et al., 2000Go). This has highlighted the need for, and the value of, standardized methods as well as effective training techniques (Chong et al., 1983Go; Jörgensen et al., 1997Go; Auger et al., 2000Go; Keel et al., 2000Go).

There have been several efforts made to improve the quality of semen analysis by recommending suitable methods and organizing training courses or programmes (Eliasson, 1971Go; Jequier and Ukombe; 1983, Jörgensen et al., 1997Go; Franken et al., 2000Go), but the effects of at least the earliest activities have, in the long run, apparently been limited mainly to the centres involved in the initial projects. The general lack of standardization of assessment criteria, laboratory procedures, patient information, sample collection and training is likely to be the main reason for results of semen analysis being unreliable and poorly comparable between laboratories.

Several studies have emphasized the importance of standardization of laboratory methods and training of staff to achieve reliable results in routine semen analysis (Mortimer et al., 1986Go; Dunphy et al., 1989Go; Seaman et al., 1996Go; Jörgensen et al., 1997Go; Auger et al., 2000Go). As a long-term project to improve standards in semen analysis the Special Interest Group in Andrology (SIGA) of the European Society of Human Reproduction and Embryology (ESHRE) decided to organize standardized training courses in semen analysis, aimed primarily at several regions in Europe. The basic concept was to teach robust and internationally accepted methods for semen analysis to improve quality and decrease variability in results among and within individuals.

The aim of this study was to investigate whether there were any immediate effects of the theoretical and practical training provided during the ESHRE–SIGA Basic Semen Analysis courses on the variability of assessments made by different observers.


    Materials and methods
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
Development of courses
The schedule and syllabus for the course were developed to teach robust and reliable basic semen analysis methods as an evolution from the courses run in Nottingham, UK, by Anne Jequier in the 1980s (Jequier and Ukombe, 1983Go) and courses organized by David Mortimer in Calgary, Canada. A pilot course was run in 1994 in Sheffield, UK, as a joint course between ESHRE and the British Andrology Society (BAS). Since then more than 20 courses have been given in accordance with the schedule and structured syllabus decided by the Education Subcommittee of SIGA (Table IGo).


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Table I. Registered Basic Semen Analysis courses given according to the ESHRE–SIGA syllabus during the period 1994–1999
 
One guiding principle of the courses was to offer them in the native language of the participants within geographical regions of Europe. In order to facilitate global standardization, the courses complied with the WHO recommendations for semen analysis (World Health Organization, 1992Go). While it is accepted that there is more than one procedure to obtain reliable results for each component assessment, in order to maximize standardization it was decided that participants would be taught and trained in the performance of only one particular technique for each procedure. The lectures included explanations of why particular methods were recommended, and how their careful application would minimize the opportunities for extrinsic variations and errors. Furthermore, a detailed syllabus of the entire course content was developed and applied in all the courses to ensure consistency and coherence in the methodological aspects. To increase the likelihood that the knowledge gained by the participants would lead to improvements in their routine laboratories, the participation of both laboratory staff and (medical) supervisors from each individual laboratory was strongly encouraged. Practical training was always accompanied by comprehensive background theory, including an overview of the clinical use of results. Thus, clinicians learned the theory of laboratory science and got some practical experience of difficulties at the laboratory bench, and technologists received some background regarding the clinical use of semen analysis as well as the theoretical basis for the recommended methods, and extensive practical training, tutored by experienced laboratory staff. To focus the ambitions of course participants, there were two examinations: one written (theory) and one practical (assessment of unknown samples). In order to obtain a diploma from ESHRE, a participant had to pass both written and practical examinations.

A summary of course contents is given in Table IIGo. Each course began with an introductory session where the scope and organization of the course were presented. An additional benefit of this introductory session was to allow the participants to meet and get to know each other before the first formal lecture. On the first day there was an initial practical session, referred to in the Results as the pre-test. Here the participants were expected to assess two samples using their current expertise using the standard equipment provided for the course, but without instruction. The paedagogical value of this pre-test was twofold—firstly, all participants would have tried their best to use the equipment before we discussed the theoretical background and, secondly, a set of `real world' results was available as a basis for discussion on the variability of assessments. Subsequently, each component procedure (i.e. sperm concentration and motility on fresh semen, vitality and morphology assessments on ready made smears) was taught in a formal lecture followed by a practical session during which measurements were made on two fresh semen samples. The results of these assessments were analysed and discussed with the participants the following morning, before the first formal practical training session, referred to in the Results as training, took place. On the last day of the course, after the theory examination, there was a practical examination, referred to in the Results as examination, during which two assessments were made of sperm concentration, motility, vitality, and morphology. The examinations were not only marked, but the answers of the participants were also analysed and presented to the participants prior to announcing the outcome of the examinations. To pass the practical examination the participant needed to achieve at least half of maximum marks. For each variable assessed 50% of the marks depended on how close the result was to the expert value. With increasing difference between result and expert value there was a gradual decrease in marks. The other 50% of the marks were obtained based on data in the report handed in. Marks were given for correct handling of the original counting data and for calculations demonstrating that recommended methods were used and understood. The number of participants who passed the examinations in each course is given in Table IIIGo.


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Table II. Topics covered in an ESHRE Basic Semen Analysis course and their approximate time allocation
 

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Table III. Numbers of participants and of results obtained at the different courses, as well as results of the examinations
 
To accompany the basic course schedule, a detailed manual on the methods taught during the courses was developed by the Nordic Association for Andrology (NAFA) and used in Nordic courses from 1997. A revised version to comply with the present WHO (1999) recommendations is available at www.ki.se/org/nafa.

Course venues
There were 24 courses registered at the ESHRE Central Office between 1994 and 1999 (Table IGo). The eight courses included in the present analysis are listed in Table IIIGo, and were selected for the present analysis because at least one of the authors was involved in running and/or teaching each course. The course language was English in these eight courses, even in the Nordic countries (organized jointly by ESHRE and NAFA) in order to eliminate the differences between the Nordic languages and to allow participation from the Baltic countries. Simultaneous translations were given by lecturers and tutors in the respective native tongue where necessary, except in Kharkiv where interpreters from the University of Kharkiv provided translation.

Data
The results from pre-test, training and examination were compared with results obtained by highly experienced technologists and/or course faculty (referred to as `expert' values). The difference in sperm concentration was expressed as a percentage of the expert value, to allow comparisons between samples with widely different sperm concentrations. All other variables were already expressed as percentages (motility fractions, proportion with normal or abnormal morphology, proportion of live sperm) or as a ratio (teratozoospermia index, TZI), and the differences between the participants' and the experts' values were used for comparisons. The expert value was always subtracted from the participant's value, so that when the participant underestimated a variable, the difference was negative.

In general, each participant assessed two samples during each practical session. During the course in Kharkiv there were some difficulties with translation and the availability of a limited number of phase contrast microscopes meant that not all participants were able to complete their analysis of two specimens in each practical session.

Unfortunately, in some of the courses, occasional participants failed to deliver a result at a session because of a lack of time due to minimal laboratory experience. A colour-blind participant did not participate in vitality assessments, but with this exception all but two participants managed to assess all samples at the practical examinations (Table IIIGo).

Purpose of the analysis
The focus of interest was to investigate any decrease in variability of results within the participant groups over time, i.e. from pre-test to training and then to examination. Therefore, group-averaged results were not considered for statistical analysis.

Statistical methods
Distributions of results for each sample were examined using box-and-whiskers plots. Analysis of variance (ANOVA) was used to compare standard deviations over the three phases (i.e. standard deviations of the pre-test, training, and examination samples). Where differences in distribution between the three phases did not allow direct analysis, log-transformation was performed. Tests for linear trend were used to determine whether a trend line was significantly different from zero.

Comparisons of concordant participant results (i.e. as compared with the expert results) were performed using the {chi}2-test. The arbitrary limit of ±10% was employed according to established standards in routine clinical andrology (Mortimer, 1994bGo).

Calculations were performed using GraphPad PrismTM 3.02 and GraphPad InStatTM 3.05 (GraphPad Software, San Diego, CA, USA, www.graphpad.com; Motulsky, 1996Go). Production of figures was also performed using GraphPad PrismTM 3.02.


    Results
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
Figure 1Go shows the box-and-whisker plots for the sperm concentration, motility (rapid, progressive and motile proportion), vitality, and morphology (both normal forms and TZI) assessments of each sample analysed during the pre-test, training and examination phases of each of the eight courses. For each variable assessed, substantial improvements were seen in the agreement between the participants' values and the reference values from pre-test to training, and from training to examination in each course.



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Figure 1. Distribution of results from assessments of the 46 test samples (numbers 1–8 represent the different courses and letters A or B refer to the two different test samples at each of the three phases pre-test, training and examination) shown as box-and-whiskers plots (lower whisker: minimum; lower box line: 25th percentile; middle box line: median; upper box line: 75th percentile; upper whisker: maximum); pre-test (n = 16), training (n = 15), and examination (n = 15). {Delta}% concentration = participant difference from expert result as a percentage of the expert value. {Delta}% motile = participant difference from expert result for the proportion of motile sperm. {Delta}% progressive = participant difference from expert result for the proportion of progressively motile sperm. {Delta}% rapid = participant difference from expert result for the proportion of rapidly progressive sperm. {Delta}% live = participant difference from expert result for the proportion of live sperm. {Delta}% normal = participant difference from expert result for the proportion of morphologically normal forms. {Delta}TZI = participant difference from expert result for the teratozoospermia index.

 
Concentration
There was a significant linear trend for decreasing variability of sperm concentration assessments, as measured by changes in average standard deviations (Figure 2Go: slope = –0.1085, r2 = 0.2087, P < 0.01), and the difference in average standard deviation between the groups was significant (one-way ANOVA, P < 0.01). The proportion of results within ±10% of expert values (Table IVGo) increased from 24% (pre-test) to 63% (examination) ({chi}2-test, P < 0.001) with a significant linear trend (P < 0.001).



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Figure 2. Decrease in course participants' variability, as revealed by average standard deviation (SD) of results from assessmentsof sperm concentration in test samples, from the pre-test (n = 16), through training (n = 15) to examination (n = 15). Values are shown as the mean (±SD) for each phase, and the broken line indicates the linear trend (slope = –0.1085, r2 = 0.2087,P < 0.01). One-way analysis of variance (ANOVA) showed a significant difference between the phases (P < 0.01).

 

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Table IV. The proportion of concordant results (concordance defined as the proportion of results within ±10% of the expert result (TZI ±0.10)) at different stages of the courses. {chi}2 values are given for tests of independence (difference between sessions) and for linear trends (increase from pre-test to examination).
 
Motility
The variability of assessment of rapidly progressive sperm decreased significantly from pre-test to examination (Figure 3Go, one-way ANOVA, P < 0.05) with a significant linear trend (slope = –1.731, r2 = 0.1289, P < 0.05). The proportions of results within ±10% of expert values (Table IVGo) increased for progressive sperm from 42–58%, and for motile sperm from 49–66%. For rapid sperm the change (44–50%) was not significant.



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Figure 3. Decrease in course participants' variability, as revealed by average SD of results from assessments of rapidly progressive sperm (WHO grade `a') in test samples, from pre-test (n = 16), through training (n = 15) to examination (n = 15). Values are shown as the mean (±SD) for each phase, and the broken line indicates the linear trend (slope = –1.731, r2 = 0.1289, P < 0.05). One-way ANOVA showed a significant difference between the phases (P < 0.05).

 
Vitality
The variability in assessments of sperm vitality did not improve significantly from pre-test to examination as measured by change in average standard deviation (Figure 1Go; {Delta}% live: one-way ANOVA, P = 0.6313). However, the proportion of results within ±10% of expert values (Table IVGo) was already high at pre-test (79%), but did increase to 88% at the examination ({chi}2- test P < 0.05, linear trend P < 0.01).

Morphology
Normal forms
The variability of assessments of sperm morphology had already decreased at the training phase as compared with the pre-test as measured by average standard deviation (Figure 4Go: one-way ANOVA P < 0.001; linear trend slope = –5.221, r2 = 0.4296, P < 0.001, pre-test to examination). The proportion of results within ±10% of expert values (Table IVGo) increased from 37% at pre-test to the very high level of 88% at training and 90% at examination ({chi}2-test P < 0.001; linear trend P < 0.001).



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Figure 4. Decrease in course participants' variability, as revealed by average SD of results from assessments of proportion of morphologically normal sperm in test samples, from pre-test (n = 16), through training (n = 15) to examination (n = 15). Values are shown as the mean (±SD) for each phase, and the broken line indicates the linear trend (slope = –5.221, r2 = 0.4296, P < 0.001). One-way ANOVA showed a significant difference between the phases (P < 0.001).

 
TZI
The variability of the TZI, as measured by change in average standard deviation, did not change significantly from pre-test to examination. There were some courses with extreme deviations in the pre-test results, and during the training there were still some extreme outliers, but not at the examination phase. However, in most courses, the central 50% of participants' results were within ±0.50 from the expert values at all phases of the courses (Figure 1Go; {Delta}TZI). The proportion of results within ±0.10 of expert values had increased from 16–33% already at the training, and remained at this level at examination (Table IVGo).


    Discussion
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
It is clear from the results of this study that a comprehensive training course does lead to substantial reductions in inter-observer variability in all aspects of semen analysis (Figures 1–4GoGoGoGo). The observation that training can have an immediate effect upon concordance among different observers and decreased inter-observer variability supports our contention that providing both theoretical background and repeated training with feedback of results is highly effective in raising the technical skills of most participants.

The results of the pre-test evaluations confirm and extend the findings of Jequier and Ukombe showing large variations among results obtained by different individuals examining the same semen sample, while a training programme does reduce the inter-observer errors (Jequier and Ukombe, 1983Go). It is not suggested that a 4 day course constitutes a comprehensive training programme for andrology technologists, rather it represents the initial, baseline theoretical and practical education that must then be followed by a further iterative process comparing a trainee's performance against reliable, reference values. Regular analysis of such data will provide positive reinforcement as concordance increases (Mortimer, 1994aGo,bGo; Franken et al., 2000Go). This practice provides a natural foundation that should be completed by robust internal quality control procedures (Clements et al., 1995Go). Wider-scale concordance can then be achieved by participation in an external quality assurance program (EQAP), but only if the EQAP provides feedback of performance against reference values. Participation in a successful EQAP can also serve as the basis for ongoing proficiency testing programmes (Franken et al., 2000Go).

All participants were expected to be at least familiar with the WHO manual, and most had indicated prior to the course that they complied with WHO recommendations. Because the technology as such is unsophisticated, adaptation to, or familiarization with, the equipment used on the course should not have been a major factor. Certainly, working within a group of peers can always be expected to be beneficial in any professional setting, but while this situation may have led to a greater comfort for the participants it is difficult to see how it could improve someone's ability to perform differential motility and morphology counts or to use the haemocytometer. The simple fact that a participant's result is different from that of the `experts' does not intrinsically create a change in how s/he performs a technique; only proper training can do this. We believe that the strong positive reinforcement during the course, where participants see the progressive decrease in discrepancies while training, illustrates the benefit of this structured training course.

In addition to the present study, there is also documentation of long term effects of the ESHRE Basic Semen Analysis courses. In Belgium (Punjabi and Spiessens, 1998Go) the participants showed a notably increased awareness of the need for standardization of semen analysis as well as a decrease in variability during the courses, as in the present study. In the Netherlands (Vreeburg and Weber, 1998Go) a large group of participating laboratories had significantly changed their laboratory procedures to reduce errors by starting to use positive displacement pipettes, improved Neubauer haemocytometers, heated (37°C) microscope stages and by complying with WHO motility and morphology criteria within 6 months of the course.

There is a need for improved standards in semen analysis. Cost effective health care for the couple seeking help for infertility includes accurate semen analysis. Semen analysis performed with improper and therefore unreliable methods without internal and external quality control is a waste of resources and often provides the wrong diagnosis, which causes negative effects for both the couple and the health service provider. Robust and reliable methods for semen analysis are available—methods that have been standardized and successfully used for training. These methods should therefore be used in all semen laboratories. This analysis provides strong support for the ESHRE–SIGA initiative on standardized, comprehensive training courses in basic semen analysis, as well as for further development of the ESHRE–SIGA Quality Assurance Program.


    Acknowledgements
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
The authors are grateful to ESHRE for financial support and assistance from the Central Office for co-ordinating these courses. We acknowledge the enormous efforts made by the co-organizers of these courses: Mrs Inger Söderlund, Mrs Solveig Johansson, Mrs Eva Lilliehöök and Mrs Ingmarie Sundgren, all from Stockholm, have contributed to the Nordic courses; Dr Erik Ernst and his co-workers in Århus; Dr Kersti Lundin and her co-workers in Göteborg; Dr Trine Haugen and her co-workers in Oslo. Kharkiv: Professor Vladimir Lesovoy and Dr Ola King Aromire. Pretoria: Professor Riana Bornman, Dr Tiaan deJager, Mrs Natalie Aneck-Hahn and co-workers. We thank Olympus, Nikon, Zeiss and Leica for kindly lending modern microscopes to the courses. Special thanks go to Mr Norbert Junker at the Olympus office in Hamburg for his long-term commitment to this project, thereby also making the course in Ukraine possible. Our gratitude also goes to Hamilton–Thorne Research (Beverly, MA, USA) for the loan of highly transportable `MiniTherm' heated stages. A minor part of these results (from the first three Scandinavian courses) was published in a presentation given at Modern ART in the 2000s—Andrology in the Nineties (Björndahl and Kvist, 1998Go).


    Notes
 
5 To whom correspondence should be addressed at: Assisted Conception Unit, Birmingham Women's Hospital, University of Birmingham, Birmingham B15 2TG, UK. E-mail: l.bjorndahl{at}bham.ac.uk Back


    References
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
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Submitted on September 20, 2001; accepted on November 3, 2001.