This document summarizes a lecture on abnormal semen parameters and what doctors should know. It discusses epidemiological trends showing rising rates of male infertility. It reviews routine semen analysis and new WHO reference values, noting lower cut-off levels. The document cautions about over-interpreting results within the new guidelines. It also emphasizes that semen analysis alone is not enough to evaluate male fertility, and that sperm chromatin integrity testing provides additional clinically relevant information beyond standard semen analysis.

1. Sandro Esteves, MD., PhD.
Director, ANDROFERT
Andrology & Human Reproduction Clinic
Campinas, BRAZIL
Abnormal Semen
Parameters
What doctors should know
Reproductive Andrology Surgery Workshop 2014
Al Jahra Hospital, KUWAIT
ISO 9001:2008

2. Available at:
h"p://www.androfert.com.br/review
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 2
2014 APRIL
ANDROFERT
Abnormal semen parameters:
what doctors should know

3. Lecture
Outline
Learning objectives
Epidemiological trends and sperm
development
Routine semen analysis: where we
are today
Importance of sperm chromatin
integrity and its clinical implications
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 3
2014 APRIL
ANDROFERT

4. Epidemiological Trends
• 10-15% of couples are infertile and 1 in 8 men seeks
medical assistance for male infertility.
• Male factor infertility is responsible for up to 50% of cases
of infertility: 20% as the sole reason and 30% as
contributory.
• Increased incidence of male infertility may be attributed to
environmental factors and modern life habits such as
obesity, ageing, exposure to gonadotoxins and certain
endocrine disruptors.
• Noticeable epidemiological increase in the incidence of
testicular cancer and urogenital anomalies
Irvin S, et al 1996, Auger J et al, 1995, Irvine DS 1994, Jørgensen N et al 2001, Jørgensen N et al 2002,
Swan SH 2003, Feki NC et al, 2009
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 4
2014 APRIL
ANDROFERT

5. ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 5
2014 APRIL
ANDROFERT

6. ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 6
2014 APRIL
ANDROFERT

7. Abnormal Semen
Multiple causes, not exclusive
– Hormonal problems
– Genetic causes
– Varicocele
– Genital infection
– Chemotherapy, radiotherapy
– Cryptorchidism
– Idiopathic
– Gonadotoxin exposure
– Life-style factors
– Endocrine disruptors
– Etc.
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 7
2014 APRIL
ANDROFERT

8. Routine Semen Analysis
Functional
Status of
Reproductive
Tract
Seminal Fluid
and Sperm
Central
Laboratory
Investigation
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 8
2014 APRIL
ANDROFERT

9. • Standardization
• Quality control
• Quality assurance
• Certification
Semen Analysis: Andrology Lab
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 9
2014 APRIL
ANDROFERT

10. Standards
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 10
2014 APRIL
ANDROFERT

11. New WHO Reference Values
WHO 20101
1.5
15
39
32 (a+b)
58
4
<1.0
Semen Parameter
WHO 1999
Volume (mL)
≥ 2.0
Count (x106/mL)
≥ 20
Total sperm number/ejaculate
≥ 40
Motility (%)
≥ 50 (a + b)
Vitality (%)
≥ 75
Morphology (%)2
(14)
Leukocytes (x106/mL)
< 1.0
1Lower Limit (5% percentile), Recent fathers; 2Strict criteria
Grade a = rapid progressive motility; Grade b = slow/sluggish progressive motility
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 11
2014 APRIL
ANDROFERT

12. New WHO Reference Values
Caution to Interpret Results
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 12
2014 APRIL
ANDROFERT

13. New WHO standards
Reasons for Lower Cut-off Values
• Method for semen analysis (QC standards)
• Adoption of strict criterion for morphology
• Single specimen of each individual
Different way of generating data
• Recent fathers with known TTP (≤ 12months)
• Selection bias
Population studied
Esteves et al. Urology 2012
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 13
2014 APRIL
ANDROFERT

14. Critical Appraisal of the WHO New Reference Values for Human Semen and Impact on
Diagnosis and Treatment of Subfertile Men
Esteves, Zini, Aziz et al, Urology, in press
Columbia
USA
Melbourne
Australia
Turku
Finland
Oslo
Norway
Edinburgh
UK
Paris
France
Copenhagen
Denmark
2010 WHO Reference: 1,953 men
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 14
2014 APRIL
ANDROFERT

15.
Volume (mL)
1.5
Sperm count (x106/mL)
15.0
Total count (x106)
39.0
% Motile (total)
40
% Motile (progressive)
32
% Normal (strict criteria)
4
%Alive
58
Cooper et al. Hum Reprod Update 2010
WHO
2010:
Recent
fathers
TTP≤12
mo.
Percentiles
5% 50%*
95%
3.7
6.8
73.0
213.0
255.0
802.0
61
78
55
72
15
44
79
91
New WHO Standards
Reasons for Lower Cut-off Values
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 15
2014 APRIL
ANDROFERT

16. New WHO Standards: Implications
Do we need to recall previous semen analysis
reports?
Abnormal
results
WHO 1999
Reclassified
as “Normal”
WHO 2010
(38.7%)
Couples (N=987) with infertility
duration >12 months
Source: ANDROFERT, Brazil
Morphology by strict
criterion accounted for
53% of reclassification
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 16
2014 APRIL
ANDROFERT

17. Referral Deferment
Semen Parameters
Case
study
Volume (mL)
2.3
Count (106/mL)
16.5
Progressive motility (%)
40
Vitality (%)
65
Morphology (%)
9
Reference
1999
2010
≥ 2
≥ 1.5
≥ 20
≥ 15
≥ 50
≥ 32
≥ 75
≥ 58
(14)
≥ 4
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 17
2014 APRIL
ANDROFERT

18. Laboratories adopting the new standards should
determine the strategy to communicate clinical
significance of the reported results
WHO Standards Interpretation
Esteves SC. Int Braz J Urol 2014
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 18
2014 APRIL
ANDROFERT

19. Semen Analysis Results
Not accurate to discriminate fertile from
infertile men
Male infertility workup
goes far beyond a simple
semen analysis. History,
physical examination,
laboratory and sperm
function tests are minimum
standards
Esteves, et al 2011; 2012; 2014
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 19
2014 APRIL
ANDROFERT

20. Practical Points
• Routine semen analysis still central in
laboratory evaluation of male infertility
• WHO new reference limits are lowered
compared with previous references
• Caution to interpret new references
– Comparison with 90% percentile distribution is
advisable
– Results not accurate to discriminate infertile
from fertile males unless if at extreme levels
– Complete male infertility evaluation should be
undertaken
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 20
2014 APRIL
ANDROFERT

21. Lecture
Outline
Learning objectives
Epidemiological trends and sperm
development
Routine semen analysis: where we
are today
Importance of sperm chromatin
integrity and its clinical implications
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 21
2014 APRIL
ANDROFERT

22. Why semen analysis is not
enough
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 22
2014 APRIL
ANDROFERT

23. Defects in Sperm DNA Structure
single-strand
break mis-match
damaged base
double-strand
break inter-strand
crosslink
intra-strand
crosslink
Single-strand DNA break (ss-DB)
Double-strand DNA break (ds-DB)
Base deletion or modification
Inter or intra-strand cross linkage
Esteves et al 2013; Alvarez and Gosálbez 2011; Ward 2011
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 23
2014 APRIL
ANDROFERT

24. Biological mechanisms of SDF
Protamination Failure
Replacement of histone to protamines during
spermiogenesis
Oxidative Stress
Epididymis transit
Post-ejaculation: leukocytes, immature sperm,
abnormal levels seminal plasma antioxidants
Apoptosis
During sperm maturation (testis & epididymis)
Fernández et al. 2009; Alvarez and Sakkas 2010; Agarwal et al. 2013
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 24
2014 APRIL
ANDROFERT

25. DNA
Damage
Environmental factors
Phtalate exposure, radiation,
temperature
Diseases
Varicocele, GTI, fever
Life-style
Obesity, smoking
Aging
External factors leading to SDF
Kort et al. 2006; Rubes et al 2007; Viloria et al 2007; Esteves & Agarwal 2011

26. SDF and Male Infertility Etiologies
Gosálbez et al. 2013
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 26
2014 APRIL
ANDROFERT

27. SDF and Infertility: Why bother?
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 27
2014 APRIL
ANDROFERT

28. 19%
1.5%
Normal Elevated
Live Birth Rates with
Intrauterine Insemination
OR = 0.07
[95% CI: 0.01-0.48]
Bungum et al. Hum Reprod 2007
IUI Outcome and SDF
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 28
2014 APRIL
ANDROFERT

29. 26%
42%
IVF
ICSI
Pregnancy by Method in
Cases of Elevated Sperm
DNA Fragmentation
IVF Outcome and SDF
Robinson et al. Hum Reprod 2012
Meta-analysis of 16 studies
and 2,969 couples
Increased miscarriage in
couples undergoing IVF/ICSI
with high sperm DNA damage
Risk ratio (RR) = 2.16
95% CI: 1.54-3.03; p<0.00001
Bungum et al. Hum Reprod 2007
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 29
2014 APRIL
ANDROFERT

30. Frequency of elevated SDF in men
with normal semen analysis
Normal semen analysis
results (WHO)
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 30
2014 APRIL
ANDROFERT
Androfert; N=987

31. Practical Implications
• Successful pregnancies in IVF/ICSI cycles can be
obtained using semen samples with a high
proportion of DNA damage
• Sperm DNA damage is promutagenic and can give
rise to mutations after fertilization, as the oocyte
attempts to repair DNA damage before the initiation
of the first cleavage.
• Mutations occurring at this point will be fixed in the
germline and may be responsible for the induction of
infertility, childhood cancer in the offspring and for a
higher risk of imprinting diseases
PANG M. G. et al Hum Reprod, 20: 1688–1694, 2005.
Burrello et al Cytogenet Genome Res, 111:363–365, 2005.
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 31
2014 APRIL
ANDROFERT

32. Fertility and Sterility 2014; 101(1):58-63
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 32
2014 APRIL
ANDROFERT

33. Correlation between SCD and
TUNEL
SCD more sensitive than
TUNEL.
Important to distinguish
between the methods as
they differently evaluate
SDF.
20.6
11.5
% SDF
SCD
TUNEL
Feijo & Esteves
Fertil Steril 2014; 101(1):58-63
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 33
2014 APRIL
ANDROFERT

34. Management
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 34
2014 APRIL
ANDROFERT

35. Take-home Messages (1)
• Male infertility is a important health issue
• Despite being genetically-determined,
male fertility is modulated by external
factors
• Life-style modifications should be
considered in males seeking fatherhood
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 35
2014 APRIL
ANDROFERT

36. Take-home Messages (2)
• Normal semen analysis is not a
guarantee of fertility, and vice-versa
• Male infertility evaluation should go far
beyond routine semen analysis
• Minimal standards include history
taking, physical examination, semen
analysis, sperm functional tests, and
other tests as appropriate
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 36
2014 APRIL
ANDROFERT

37. Take-home Messages (3)
• Sperm chromatin integrity critical for a
viable pregnancy
• Sperm DNA fragmentation common in men
with unexplained infertility
• SDF testing provides information that is
different and of better prognostic value
than semen analysis
• Grading SDF may help in designing a cost
effective management
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 37
2014 APRIL
ANDROFERT

38. Thank you Obrigado
‫شكرا‬
ANDROFERT
androfert.com.br
ANDROLOGY AND HUMAN REPRODUCTION CLINIC - REFERRAL CENTER FOR MALE REPRODUCTION
S ESTEVES, 38
2014 APRIL
ANDROFERT