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Re-evaluation of sperm morphology assessment and results in light of new
WHO (2010) manual reference limits
Roelof Menkveld, PhD
Andrology Laboratory, Department of Obstetrics and Gynaecology, Tygerberg Academic Hospital and University
of Stellenbosch, Tygerberg, South Africa.
Pre-congress course: Cotemporary approaches in embryology laboratory – How can IVF success be raised?
III rd Congress of the Society of Reproductive Medicine
Cornelia Diamond Resort Belek, Antalya, Turkey
05 to 09 October 2011
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Historical background of sperm morphology evaluation methodology
Basically two different evaluation approaches or methodologies – Liberal (old WHO) approach– Strict (Tygerberg criteria) approach
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Early Liberal Approach
Normality for human spermatozoa• Based on approach in domestic animals with a
homogeneous sperm population– Using modal forms of fertile males animals
• In humans – heterogeneous picture– Thus not a feasible approach – Described abnormal spermatozoa based on consensus
decision – thus normal by elimination• Morphological forms depicted by schematic and
inaccurate drawings
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Liberal approach
• Theoretical disadvantages– No specific criteria for normal
• If not abnormal = Normal
– Normal population will consist of • Abnormal population • True normal population
– Therefore, can expect poor correlation with • Normal sperm function• Fertilisation and pregnancy rates
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Liberal approachDisadvantages according to literature
• Abnormal sperm morphology:– Is less sensitive for evaluation of ejaculate
• Van Duijn et al., 1972– Has no correlation with pregnancy
• Page and Holding, 1951– Of less importance compared to normal morphology
• Page and Holding, 1951• Hellinga, 1976
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Strict Approach
• Conceptualized – Late 1970’s early 1980’s
– Tygerberg Hospital (R Menkveld)
• Biological based concept for normality– Sperm selective capability of good cervical
mucus
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Strict (Tygerberg) criteria (1)
• Whole spermatozoon must be considered– Head
• Oval with smooth contours• Good distinction between acrosome and post
acrosome region• Homogeneous light blue staining of acrosome
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Strict (Tygerberg) criteria (2)
• Correct neck implantation
• No neck/midpiece abnormalities
• No tail abnormalities
• No cytoplasmic residues (>30% normal head)
NB - Borderline normal is abnormal
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Literature on origin of Strict Criteria
Menkveld (1987) – The influence of environmental factors on
spermatogenesis and semen parameters. PhD Dissertation. Faculty of Medicine, University of Stellenbosch, Tygerberg (Cape Town), South Africa.
Menkveld et al. (1990)– The evaluation of morphological characteristics
of human spermatozoa according to stricter criteria. Hum Reprod 5(5):586-92.
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Evolution of sperm morphology evaluation approaches in consecutive WHO manuals
• 1980– Basic liberal approach
– Very basic descriptions for normal spermatozoon
• 1987– Same basic approach
– Slightly more descriptive information
• 1992– Strict approach should be followed
– Borderline normal = Abnormal
• 1999 and 2010 (WHO-5)– Accept strict criteria - functionality based
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Consequences of introduction of strict criteria
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Overview of declining sperm morphology values over years
70
60
50
40
30
20
10
0
Year of semen analyses
Morp
holo
gy (
% n
orm
al)
1968
1970
1972
1974
1976
1978
1980
1982
1984
1986
1988
1990
1992
1994
1996
1998
2000
2002
2004
2006
2008
Menkveld etal., 1986; Menkveld, 2009
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Normal values from WHO manuals, editions 2- 4 and lower reference limits from new 5th WHO manual (2010)
Semen parameterWHO edition and year
2nd - 1987 3rd - 1992 4th - 1999 5th - 2010
Volume (ml) 2.0 2.0 2.0 1.5
Sperm concentration (106/ml) 20 20 20 15
Total sperm count (106) 40 40 40 39
Motility (% progressive) 50 50 50 28
Vitality (% live) 50 75 75 59
Morphology (% normal) 50 30 (15) 4
Cooper, 2007 (ESHRE campus meeting)
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Possible reasons for lower normal sperm morphology values
• Decline may be due to– Stricter application of (strict) sperm morphology
evaluation criteria– Negative environmental influences– Recognition of additional sperm morphology
abnormalities/parameters
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Possible solution for declining normal sperm morphology values
In WHO-5 abnormal morphology group (≤ 3%)
Identification of
Additional abnormal sperm morphology patterns
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Assessment of specific sperm morphology abnormalities
Four basic sperm abnormalities• Head abnormalities (Several classes)• Neck and midpiece abnormalities• Tail abnormalities• Presence of cytoplasmic residues
Teratozoospermia index (TZI – WHO-5)
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Head abnormalities can be used to determine abnormal sperm morphology patterns
Head abnormalities (Several classes)– Large– Small– Elongated (Tapering and pyriforms)– Acrosome abnormalities (Several classes)
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Acrosome morphology classes
• Differential classification of acrosomes– Normal– Staining defects– Too large – Too small– Other/Amorphous Total number of sperms with
normal acrosomes Sperm morphology patterns
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Are these specific abnormalities of any clinical significance?
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Large acrosomes – Spermac stain
• Spontaneous acrosome reaction
• No zona pellucida binding of spermatozoa
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Small acrosomes•Mostly non-viable•Can not undergo acrosome reaction•Can not bind to zona pellucida
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Acrosome reacted – Papanicolaou staining•Not able to bind to the zona pellucida
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Acrosome reacted – Spermac stain
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Acrosomes with staining defects• Beginning of acrosome reaction ?• Cysts and vacuoles ?• Membrane damage ?• Not able to bind to zona pellucida ?• DNA status (MSOME) ?
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Large headed spermatozoa• DNA status ?• Poor prognosis for normal in vitro fertilisation
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Elongated spermatozoa pattern
• DNA damage
• Ultrastructural nuclear defects
• Stress
• Chromosome aneuploidy (Prisant et al., 2007)
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Neck defects• Absence of centriole – no spindle formation in oocyte
Midpiece abnormalities• Mitochondrial defects (? Poor motility)
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Cytoplasmic residues
• ROS production
• Immaturity of spermatozoa
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Sperm morphology and fertilisation
Important aspects
• Need morphological normal spermatozoa for normal sperm functions throughout the whole fertilisation pathway
• Patients with clear abnormal sperm patterns– ? Need for sperm functional tests
• Patients with apparent high % normal morphology– ? Sperm functional tests
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Strict criteria still applicable?
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Normal morphology distributions in 2000 vs 2007 (76 and 112 couples, mean normal morphology 7.3% and 7.2%; P=0.5443 )
25
20
15
10
5
0
MORPH NormF
Rhemrev et al., Unpublished data
2000 200720002000 2007
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Normal morphology: 2007 comparison between infertile and fertile population (n = 40 and 112)
Norm
25
20
15
10
5
0
Diagnosis
0 1
>3.0Sens: 87.5Spec: 85.0
Infertile Fertile
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Strict criteria still applicable?
Yes - with world wide co-operation
Problem– Lack off standardisation between different
international QC schemes
• Better Quality Control– Inter- and Intra-laboratory
• Need international cooperation for – standardisation– Quality control
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Tygerberg Academic Hospital and University of Stellenbosch Medical school
Thank you for your attention