1. Clinical Significance of Sperm Morphology Assessment
Aiswarya Lekshmi – 12106912
lekshmiaishwarya@gmail.com
07854065859
Clinical Significance of Sperm Morphology
Closely correlated with specific sperm functions
Regarded as the most consistent sperm variable that can be related to
IVF success
Configuration of acrosome is a good method to estimate sperm
fertilizing ability (Estherhuizen et al.)
Correlates significantly with the sperm cells ability to bind with zona
pellucida (Menkveld et al.)
Sperm with small acrosomes are more susceptible to cell death and
non-physiological acrosomal loss (Menkveld et al.)
Acrosome size reflects male fertility potential
Materials and methods
Search engines used: Pubmed, Google Scholar, Sciencedirect, Elsevier.
Keywords used: Sperm morphology; ICSI; male infertility; sperm selection; pregnancy outcomes, sperm morphological anomalies
and chromosomal aneuploidies, quality control, normal spermatozoa, sperm morphology defects, Diff-Quik, testsimplets, poster
presentation.
Visited histopathology department at The Royal Oldham Hospital to understand the clinical significance of semen analysis. In order
to improve the efficacy of sperm morphology analysis and to improve quality control, the lab now uses an automated analyzer. The
SQA-V Gold analyzer measures the 2 quantifiable attributes in semen (viz. motility and sperm concentration) to estimate the sperm
morphology. This has brought down the total time for analysis to 30 minutes. Semen analysis is usually done in post vasectomy and
reversal of sterility in males.
Strong effect on inducibility of acrosome reaction(Franken et al.)
Zona induced acrosome reaction(ZIAR) is an important indicator of
dysfunctional spermatozoa in men with normal semen quality.
Relationships between the % of normal morphological forms and
various fertility endpoints (time-to-pregnancy (TTP) is used for
prognosis of fertility
Sperm Morphology
Improved Neubauer Haemocytometer for
sperm count
Observations on morphologically normal spermatozoa recovered from postcoital endocervical mucus and from surface of zona
Test strip for Semen
pellucida. Papanicolaou, Shorr or DiffQuik stain is used for observing sperm morphology.
pH
Certain morphology patterns and sperm abnormalities have strong
prognostic value .eg. Sperm macrocephaly syndrome
Sperm Morphology and DNA Status (Franken, D. R.and Oehninger, S. (2012)
Sperm nucleus abnormalities with implications on reproductive
outcome includes:
DNA strand breaks
Numerical and structural chromosomal abnormalities
Y chromosome microdeletions
Alterations in the epigenic regulation of parental genome
Sperm DNA damage
•Single and double DNA strand
breaks
•Chemical modification of base.
eg.oxidation /alkylation
•Inter/intra strand cross-linkage
•DNA-protein cross-links
Tests to detect sperm DNA damage
•Direct method:
TUNEL
Comet @pH 7
•Indirect method (measurement after denaturation steps)
Sperm chromatin structure assay
Sperm chromatin dispersion
Comet @ acidic/ alkaline pH
Semen Analysis According to WHO manual
5th edition (Tygerberg criteria)
Copyright medical-dictionary.thefreedictionary.com
Semen has two major quantifiable attributes:
1.Total number of spermatozoa
2.The total fluid volume
Structure of normal sperm
Other Parameters for
sperm function
Nature of
spermatozoa
Morphology
Composition of
seminal fluid
Motility
Abnormal human spermatozoa
Normal human spermatozoa
Mid-piece(cross-section)
Categories of defects
•Head Defects -large or small, tapered,
pyriform, round, amorphous, vacuolated
(more than two vacuoles or >20% of the
head area occupied by unstained
vacuolar areas), vacuoles in the postacrosomal region, small or large
acrosomal areas (<40% or >70% of the
head area),double heads
•Neck and mid-piece defects:
asymmetrical insertion of the mid-piece
into the head, thick or irregular, sharply
bent, abnormally thin.
•Principal piece defects: short, multiple,
broken, smooth hairpin bends, sharply
angulated bends, of irregular width,
coiled
•Excess residual cytoplasm (ERC):
large amounts of irregular stained
cytoplasm, one third or more of the
sperm head size, often associated with
defective mid-pieces
Schematic drawings of some abnormal forms of human spermatozoa. (WHO, 2010)
Vacuolated Spermatozoa
Neck/ Midpiece defect
Acrosomal Defect
Principal piece defects
Two-headed Spermatozoa
Globozoospermia
Sterilizing defect
Tail(cross-section)
Vitality
[Progressive motility
(PR), Non-progressive
motility (NP),
Immotility (IM)]
Table 1: Cut-off values for semen variables as published in the fifth
World Health Organization (WHO) manual
Result And Discussion
The practical conducted on sperm morphology and
vitality on 11/02/2013 @ MMU –
Result - closer to the upper reference value
according to WHO 2010.
Reason – 1) lack of training 2) counting borderline
forms as normal.
WHO 2010 will assist in training technicians and
students to categorize spermatozoa consistently.
*Lower reference limit. Obtained from the lower fifth centile value.
Table 2: WHO criteria for morphological assessment of spermatozoa
Table 3: Result of sperm morphology & vitality practical
Franken, D. R., and Oehninger, S. (2012)
Box Whisker Plots recorded during pre- and posttraining workshops for sperm concentration
A prospective study on monitoring technologist reading skills in a sperm morphology quality control program (Daniel, R.F. et al.
2003) showed - 1) continuous quality control program can be initiated only after intensive training
2)baseline values at the onset of the quality control program serves as an internal reference value.
Study on influence of individual sperm morphology on fertilization, embryo
morphology, and pregnancy outcome of ICSI (De Vos, A et al. 2003) suggested that
normal sperm morphology correlated well with fertilization rate which followed live
births.
Current sperm selection techniques assume that if an ejaculated spermatozoon has cleared spermatogenesis with normal
morphological and/or membrane properties then it is normal. Sperm immobilized before ICSI undergo ultrastructural damage and
acrosomal disruption(Jose, M. et al. 2007).
Novel technologies for selecting the best sperm for IVF and ICSI are being researched (Sakkas, D. 2013) - most effective
techniques: electrophoretic apoptotic sperm deselection or microfluidic preparation followed by individual sperm selection for
ICSI using high magnification (IMSI) or by selecting sperm whose membranes possess hyaluronan receptor (HA binding).
A cohort study to analyse the advantage of IMSI over ICSI (Klememt et al, 2013), multivariate analysis showed an approximate
threefold increase existed for both pregnancy and delivery only in the case of couples failing an ICSI attempt who shifted to IMSI.
The results showed that sperm morphology has no prognostic value in IMSI and to promote the use of IMSI method only for
couples who failed ICSI cycle more than once. MSOME is an independent test proposed after repeated IVF or ICSI failures
(Delaroche, L et al. 2012).
Specimen requirement for semen analysis
Complete semen sample (including the first, sperm-rich
portion) collected in sterile container
Sample received in lab within 1 hour, analysed within 2
hours of production
Specimen produced after min 2 days and maximum 7 days
of sexual abstinence
Sample transported to the lab at body temperature.
Acknowledgment
I would like to thank Dr. Michael Carroll for giving the initial insight into Semen
analysis and providing us with the essential material and pictures on sperm
morphology. I would like to extent my gratitude to Ms. A Pearson and Mr. Zameer
Abbas, the staff in Histopathology at The Royal Oldham Hospital for their time and
effort in showing and explaining the whole process of semen analysis within their
busy schedule. I also thank my Senior Mrs. C Loy and colleagues Mr. M Sidorczuk
and Ms. T Mzumara for their critical feedback.
A study on a mild variant of SMS (Molinari, E. et al. 2013) characterised by large-headed, mono-tailed, mono-centriolar
spermatozoa with abnormal chromatin and swollen mitochondria but no mutations in AURKC gene. The sperm produced an
apparently normal fertilization and zygote development, followed by a pregnancy bound to be aborted for severe fetal genetic
aberrations. It was proposed that FISH analysis should be proposed to all macrocephalic sperm patients before allowing an ICSI.
Scientists are still in search of novel techniques to treat infertility and sperm morphology assessment along with other semen has a
very significant role to play.
* AURKC = aurora kinase C gene; HA = hyaluronic acid; ICSI = intracytoplasmic sperm injection, IMSI = intracytoplasmic morphologically selected
sperm injection; IVF= in vitro fertilization; MSOME = motile sperm organellar morphology examination; ROS = reactive oxygen species; SMS= Sperm
macrocephaly syndrome; TUNEL = terminal deoxynucleotidyl transferase mediated dUTP nick end labelling.
References:
De Vos, A., Velde, H.V.D., Joris,H.,Verheyen, G., Devroey, P., Steirteghem, A. V. (2003) ‘Influence of individual sperm morphology on fertilization, embryo morphology, and pregnancy outcome of intracytoplasmic sperm injection.’ Fertility and
Sterility 79(1), 43-48.
Franken, D. R., and Oehninger, S. (2012) ‘Semen analysis and sperm function testing.’ Asian Journal of Andrology, 14, 6–13
Klement, A. H., Koren-Morag, N., Itsykson, P., Berkovitz,A .(2013) ‘Intracytoplasmic morphologically selected sperm injection versus intracytoplasmic sperm injection: a step toward a clinical algorithm.’ Fertility and Sterility, [Online, 28
January 2013-03-19]
Menkveld , R. (2010) ‘Clinical significance of the low normal sperm morphology value as proposed in the fth edition of the WHO Laboratory Manual for the Examination and Processing of Human Semen.’ Asian Journal of Andrology, 12:,
47–58
Molinaria, E., Mirabellib, M., Raimondoc, S., Brussinoe, A., Gennarellif, G., Bongioannia, F.,Revelli, A.(2013) ‘Sperm macrocephaly syndrome in a patient without AURKC mutations and with a history of recurrent miscarriage.’ Reproductive
BioMedicine Online, 26(2), 148-156.
Sakkas, D.(2013) ‘Novel technologies for selecting the best sperm for in vitro fertilization and intracytoplasmic sperm injection.’ Fertility and Sterility, 99(4), 1023–1029
WHO laboratory manual for the examination and processing of human semen - 5th ed.