ISSN 1062-3604, Russian Journal of Developmental Biology, 2008, Vol. 39, No. 1, pp. 42–51. © Pleiades Publishing, Inc., 2008.
Original Russian Text © E.N. Bocharova, L.F. Kurilo, L.V. Shileiko, E.E. Bragina, Yu.B. Yurov, S.G. Vorsanova, I.Yu. Iourov, R.R. Klimova, A.A. Kushch, 2008, published in
Ontogenez, 2008, Vol. 39, No. 1, pp. 47–57.
The factors affecting male fertility include sexually
transmitted infections (STIs) and genital herpes, in par-
ticular, which is caused by herpes simplex virus (HSV).
HSV is one of the most common pathogens among both
animals and humans (Halioua and Malkin, 1999).
The effect of HSV on the structure and life cycle of
mammalian cells is largely studied using in vitro cell
models. The study of human ﬁbroblasts infected by
HSV demonstrated that this virus blocks mitosis and
induces polyploidy and endoreduplication (Chenet-
Monte et al., 1986). Studies on cell cultures have also
demonstrated the genotoxic effect of HSV including
chromosomal aberrations in the form of chromosome
breaks. In addition, centromeric decondensation and
changes in chromatin structure have been reported.
Chromosome 1 is most commonly affected
(Mincheva et al., 1984; Peat and Stanley, 1986).
Recently, Fortunato and Spector (2003) have demon-
strated a nonrandom pattern of damage induced by
HSV, which largely affects chromosome 1 at loci
. They claim that the observed chromosome
breaks take place in cells in S-phase when they are
infected by HSV. Little data are available on the effect
of HSV on meiosis. At the same time, highly differen-
tiated cells such as sperm can contain HSV proteins,
DNA, viral capsids, and infectious viruses (Kotro-
nias and Kapranos, 1998; Kapranos et al., 2003; Bocha-
rova et al., 2003, 2004, 2006).
Attempts to simulate herpes infection of mature
germ cells by incubation of sperm with HSV in vitro
were not successful: the virions attached to the sperm
surface but no intragamete infection was observed (Pal-
lier et al., 2002; Courtot et al., 2004). However, sper-
matogenic epithelial cells in guinea pig proved sensi-
tive to HSV in a model of experimental herpetic orchitis
(Bragina et al., 2002). These data suggested that HSV
Analysis of Germ Cell Populations in Ejaculate
of Men Infected with Herpes Simplex Virus
E. N. Bocharova
, L. F. Kurilo
, L. V. Shileiko
, E. E. Bragina
, Yu. B. Yurov
, S. G. Vorsanova
I. Yu. Iourov
, R. R. Klimova
, and A. A. Kushch
Ivanovsky Institute of Virology, Russian Academy of Medical Sciences, ul. Gamalei 16, Moscow, 123098 Russia
Research Center for Medical Genetics, Russian Academy of Medical Sciences, ul. Moskvorech’e 1, Moscow, 115478 Russia
Belozerskii Institute of Physicochemical Biology, Moscow State University, Vorob’evy gory, Moscow, 119992 Russia
Research Center of Mental Health, Russian Academy of Medical Sciences, Zagorodnoe sh. 2, Moscow, 119152 Russia
Moscow Research Institute of Pediatrics and Pediatric Surgery, ul. Taldomskaya 2, Moscow, 127412 Russia
Received March 2, 2007; in ﬁnal form, July 1, 2007
—Cytological and molecular genetics methods were used to study sperm from patients with sperm
infected with herpes simplex virus (HSV) as indicated by virological and immunocytochemical tests. The fol-
lowing methods were used: (1) sperm analysis to evaluate the morphology and functional properties of sperm;
(2) ﬂuorescence in situ hybridization (FISH) with DNA probes speciﬁc for chromosomes 1, X, and Y to evaluate
nondisjunction frequencies of these chromosomes in sperm; and (3) quantitative analysis of immature germ
cells in the ejaculate to identify spermatogenic abnormalities. The total sperm count and the count of sperm
with normal motility proved similar to the norm. FISH analysis demonstrated no difference in the nondisjunc-
tion frequency of chromosomes 1, X, and Y between infertile patients with HSV-infected sperm and fertile
donors. Comparative quantitative analysis of immature germ cells from the ejaculate has demonstrated a sig-
niﬁcant and considerable (threefold) increase in the number of spermatocytes I in pachytene and diplotene, at
the prepachytene stages of prophase I (preleptotene, leptotene, and zygotene) in HSV patients compared to normal
donors. At the same time, HSV patients demonstrated a signiﬁcant decrease in the number of spermatocytes I, in
pachytene and diplotene, decrease in the proportion of spermatocytes II and spermatids, and a twofold increase
in the number of unidentiﬁable immature germ cells. The data obtained indicate a partial spermatogenic arrest
at the early stages of meiotic prophase I in HSV patients, which prompts further research into the cellular mech-
anisms of abnormal spermatogenesis after viral infection in humans.
permatogenesis, immature sperm cells, herpes simplex virus, chromosome identiﬁcation by ﬂuo-
rescence in situ hybridization (FISH), DNA probes.