designed to improve results to artificial insemination should be tested by breeding early in estrus where sperm viability is most limiting and embryo quality is best. q 2000 Elsevier Science B.V.
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Keywords: Semen quality; Fertility; Embryo quality; Accessory sperm
1. Introduction
There is a wealth of published research showing positive relationships of seminal traits, as measured in the laboratory, to fertility. It is clear, however, that the modest
correlations obtained leave little doubt that we still lack sufficient knowledge to predict the outcome of matings based upon seminal quality as currently judged. Sources of the
Ž .
problem include repeatability of our laboratory tests Graham, 1978 , our ability to Ž
. accurately measure fertility Oltenacu and Foote, 1976 as well as clearly establish the
Ž .
existence of a variance in fertility among semen samples tested Linford et al., 1976 . Another aspect of the problem is our tendency to oversimplify the biological nature of
subfertility due to the male or inseminate, often assuming that a single test or pair of tests will embrace the majority of factors important to fertility. Therefore, I would like
to orient this paper toward our growing understanding of the nature of reproductive failure due to the male or inseminate. If we understand the principles involved, we then
should be able to place laboratory evaluation of semen traits in proper perspective and improve our ability to reach the stage of predicting pregnancy rate.
2. Compensable and uncompensable seminal deficiencies
It is now recognized that seminal deficiencies reflected in reduced pregnancy rates can be categorized as compensable or uncompensable. Seminal differences among males
in numbers of sperm required to reach their respective maximum pregnancy rates are considered compensable. Once the maximum pregnancy rate of a male is met and
further sperm inseminated are without effect, differences in pregnancy rates due to the male are then considered uncompensable.
2.1. Compensable semen traits Ž
. Pace et al. 1981 showed viability traits of sperm that could be considered compens-
able included percent progressively motile sperm and percent of spermatozoa with intact cell or acrosomal membranes. Up to a threshold, they demonstrated that it was the
number, and not percent of such cells in the inseminate that was important to pregnancy Ž
. rate. Overstreet et al. 1978 demonstrated that the population of sperm accessing the
Ž site of fertilization under the sustained sperm transport phase
potential fertilizing .
Ž sperm was nearly 100 live. Immotile sperm Mattner, 1963; Overstreet et al., 1978;
. Mullins and Saacke, 1989
or sperm with tail defects or protoplasmic droplets Ž
. Koeford-Johnsen, 1972; Mitchell et al., 1985 are markedly impaired at barriers in the
female tract precluding their access to the site of fertilization in the oviducts. Consider- ing inseminated spermatozoa with misshapen heads, only those with the most subtle
Ž .
distortions can cross barriers such as the uterotubal junction Krzanowski, 1974 or Ž
. appear as accessory sperm Saacke et al., 1998a . Morphologically, spermatozoa appear
to be excluded from contact with the ovum in vivo, more by severity rather than type of head distortion.
Clearly, there are compensable seminal traits that cannot be explained by the conventional assessments mentioned above. In the bovine, males can differ 10-fold or
more in ability to access the ovum in vivo based upon fertility at low insemination dose Ž
. den Daas et al., 1998 , or based on accessory sperm numbers in 6-day-old recovered
Ž .
ova and embryos Nadir et al., 1993 . These differences have not been explained by conventional viability and morphology semen tests. Undoubtedly important are func-
tional changes of sperm in the oviducts involving motility patterns or molecular events on the surface of sperm associated with capacitation, spermregg recognition or the
acrosome reaction. For penetration of the viscoelastic zona pellucida, hyperactivated
Ž .
rather than progressively motile sperm are thought to be critical Suarez and Dai, 1992 . Potential fertilizing sperm which colonize the lower oviductal isthmus by binding to the
Ž .
mucosa, may also be quantitatively important Lefebvre et.al., 1997 . Similarly, molecu- Ž
lar changes at the sperm surface may involve seminal plasma proteins Killian et al., .
Ž 1993, 1996 or seminal plasma-related sperm surface binding properties Bellin et al.,
. 1994; Ax et al., 1996 that have been positively related to fertility in bulls. Numbers of
sperm reaching the site of fertilization are small in relation to the number inseminated Ž
. Hunter and Wilmut, 1984 . Governing this number are many factors involving the
physiology of the Fallopian tube and the adjacent ovary which appear to work in concert Ž
. regarding cueing of sperm to the ovum see Hunter, 1998 for concepts and review .
However, there is also a positive relationship between the number of sperm inseminated Ž
and the number of sperm reaching the oviduct Morton and Glover, 1974; Larsson and .
Larsson, 1986 as well as with the number of sperm at the ovum competing for Ž
. fertilization based upon quantification of accessory sperm
Nadir et al., 1993 or
Ž .
numbers of ova with accessory sperm Hawk et al., 1988 . The importance of compensable traits to artificial insemination is the fact that such
traits govern the minimal numbers of sperm required for an inseminate to reach maximum fertility.
2.2. Uncompensable semen traits Observations in a variety of species indicate that factors associated with lowered
sperm quality as measured by viability and morphology, result in low embryo quality or Ž
very early embryonic failure prior to maternal recognition of pregnancy Barth, 1992; Courot and Colas, 1986; DeJarnette et al., 1992; Orgebin-Crist and Jahad, 1977; Setchell
. et al., 1988, see Setchell, 1998 for review . Clearly, semen or sperm traits leading to this
reproductive wastage suggests spermatozoal incompetence after fertilization. Such a deficiency could not be eliminated by sperm dosage alone, thus, the suggestion that the
deficiency is uncompensable. Differences among bulls in embryonic development of their conceptuses have been reported at the time of routine recovery for embryo transfer
Ž .
Ž Miller et al., 1982 and after observation of embryo survival in recipients Coleman et
. al., 1987 . Bulls were also shown to differ in the development of their embryos
Ž following in vitro fertilization Eyestone and First, 1989; Hillery et al., 1990; Shi et al.,
. 1990; Eid et al., 1994 . In low fertility bulls, early cleavage rates were reduced and
Ž .
pronuclear formation was delayed Eid et al., 1994 . This represents strong evidence for the uncompensable component in the low fertility bulls.
Incompetent fertilizing sperm that can be labeled as uncompensable have not been identified using contemporary laboratory semen tests. However, sperm with subtly
Ž misshapen heads as well as those with nuclear vacuoles craters, diadem, or nuclear
. pouches, defined by Coulter et al., 1986 on otherwise normally shaped heads are known
Ž to access the ovum following artificial insemination as determined from accessory
. sperm, Saacke et al., 1998a . In addition, semen with sperm having these traits has been
shown to yield higher frequencies of low quality embryos and lower fertilization rates Ž
than controls where such traits are missing or minimized Miller et al., 1982; DeJarnette .
et al., 1992; Saacke et al., 1994 . Morphologically, abnormal sperm in semen of bulls has been associated with
Ž subfertility and sterility for many years Williams and Savage, 1925, 1927; Lagerlof,
. 1934 . We now recognize that sperm with classically misshapened heads, described by
these early workers using simple microscopes, do not traverse the female reproductive Ž
. tract or participate in fertilization Saacke et al., 1998a . In addition, the zona pellucida
itself appears to provide one of the most formidable barriers against sperm with Ž
. misshapen heads Howard et al., 1993 . On this basis, a majority of abnormal sperm
would be considered a compensable deficiency. However, males having disturbances in spermatogenesis resulting in ejaculation of abnormal sperm usually provide a broad
spectrum in severity of morphological forms dependent upon the stage of spermatogene-
Ž .
sis affected by the disturbance Vogler et al., 1991, 1993 . Thus, we believe that recognition of classically abnormal sperm in semen may represent the ‘‘tip of the
iceberg’’ where disturbances in spermatogenesis signified by these sperm extend to otherwise normal or near-normal appearing sperm in the same ejaculates. These normal
and near-normal appearing sperm that can access the ovum in vivo are the most likely candidates for the ‘‘uncompensable’’ traits causing pregnancy wastage through very
early embryonic death.
Finally, it should be recognized that sperm with microscopically normal morphology but with defective chromatin have been implicated in cases of male subfertility for some
Ž .
time Gleldhill, 1970 . The chromatin structure assay developed by Evenson et al. Ž
. 1980 revealed a strong positive association between heterospermic fertility in bulls
Ž .
based upon genetic markers at birth and stability of the sperm DNA to acid denatura- Ž
. Ž
. tion Ballachey et al., 1988 . Using this same assay, Karabinus et al. 1997 have shown
that sperm ejaculated before a mild thermal insult of the testis by scrotal insulation have more stable DNA than those ejaculated after scrotal insulation where abnormal sperm
are also evident. Flaws in packaging and condensation of sperm chromatin during
Ž .
spermiogenesis Sakkas et al., 1995 and 1996 has been speculated to be involved in the instability of DNA of subfertile semen. The instability of the DNA may be due to
Ž .
limitations in disulfide bonding within nuclear proteins Kosower et al., 1992 , a process Ž
that continues throughout spermiogenesis and epididymal maturation Bedford and
. Calvin, 1974 . Such flaws from a testicular perturbation such as elevated testis tempera-
ture early in spermiogenesis would be expected to influence sperm head shape; however, the same perturbation late in spermiogenesis or during epididymal maturation could
Ž impact DNA condensation without effect on head shape, which is already set Vogler et
. al., 1993 . Flaws in sperm DNA have also been incriminated in altered kinetics of
Ž nuclear decondensation associated with pronuclear formation following fertilization Qiu
. et al., 1995 . Regarding the embryological impact of an incompetent sperm, this timing
agrees quite well with the uncompensable component in bulls affecting kinetics of rate Ž
. of pronuclear formation and early cleavage in the bovine Eid et al., 1994 discussed
earlier. Potentially underlying normal condensation and decondensation of DNA in sperm formation, maturation, and subsequent fertilization are optimum disulfide–thiol
relationships within the nuclear protein. These relationships, in turn, are dependent upon
Ž the redox status of the sperm environment at each step for reviews of this concept, see
. Sakkas et al., 1999; Levine, 1996 . Clearly, chromatin status in the sperm head presents
a strong focal point for research designed to reveal the ‘‘uncompensable’’ component in semen causing early pregnancy wastage.
3. Ova r r
