2009: What's known about selected sperm abnormalities in the bull? What’s known about selected sperm abnormalities in the bull? 1 2 J. D. Smith 3 4 Department of Pathobiology and Population Medicine, College of Veterinary Medicine, 5 Mississippi State University, Mississippi State, MS, USA 6 7 Abstract 8 The morphological evaluation of spermatozoa is an important and often 9 overlooked aspect of the bull breeding soundness examination and it is imperative that 10 practitioners not only be able to recognize defective spermatozoa but also to understand 11 the origins of observed defects and provide information related to their effects on the 12 bull’s fertility. This review describes the common causes of altered spermatogenesis. It 13 details many of the morphological abnormalities frequently encountered during routine 14 bull breeding soundness exams. Insight is given into their causes as well as the potential 15 impact of each defect on the fertilization process. 16 Keywords: Bull, spermatozoa, acrosome, proximal droplet; breeding soundness 17 examination 18 Introduction 19 The testicle of a bull is a very sensitive organ which is capable of responding to 20 an assortment of insults including heat, hypoxia, radiation, toxicity, and stress as well as 21 the influence of genetic disorders. The response of the testis to these insults often leads 22 to the formation of spermatozoa with observable defects that can be seen during a routine 23 spermiogram. Disturbances in spermatogenesis result in the formation of a wide range of 24 defective spermatozoa which can be observed clinically. Attempts to make both an 25 etiologic diagnosis as well as provide a prognosis for the bull’s potential recovery of 26 normal spermatogenesis is one of the goals of a complete breeding soundness 27 examination when excessive numbers of abnormal spermatozoa are observed. This paper 28 337 will review a select population of sperm defects which could be encountered in veterinary 29 practice and provide their common causes as well as the prognosis for fertility when 30 known. 31 Causes of altered spermatogenesis 32 The two most common insults which affect bulls on pasture appear to be heat and 33 stress.1 The affects of trauma, elevated ambient temperature, fever, excessive deposition 34 of fat in the scrotal neck and scrotal frostbite can all lead to increased heat in the scrotum. 35 The impact of the excessive heat in the scrotum leads to altered spermatogenesis and the 36 eventual appearance of increased numbers of morphologically abnormal spermatozoa in 37 the ejaculate. The temperature of the scrotal contents in normal bulls generally varies 38 between 33.0 and 34.5 ºC with a temperature gradient of 6 ºC from that of core body 39 temperature.2 Numerous studies have shown that even very small increases in testicular 40 temperature can have dramatic impact on spermatogenesis leading to the subsequent 41 appearance of abnormal spermatozoa in the ejaculate.1,3,4 The testicles of bulls appear to 42 function in a nearly hypoxic state which is in part due to the actions of the pampiniform 43 plexus which essentially eliminates the pulse pressure and slows the blood entering the 44 testicle.2 When scrotal temperatures increase, the metabolic demands of the testicles 45 increase as well, but there is no corresponding increase in testicular blood flow ultimately 46 leading to testicular hypoxia and subsequent alterations in spermatogenesis. 1 47 Stress in the form of illness, pain, herd or social interactions, transportation, and 48 weather conditions can be experienced by bulls. There appear to be disturbances in the 49 endocrine pathways associated with normal spermatogenesis in bulls under stress.1 The 50 affect of stress has been measured by examining the relationship between cortisol, 51 338 luteinizing hormone (LH) and testosterone. High cortisol levels in bulls have been 52 associated with reduced levels of LH and testosterone when compared to bulls with 53 normal cortisol; the elevated cortisol levels may interfere with normal spermatogenesis.5 54 The toxic effects of gossypol on spermatogenesis are well-recognized and the 55 characterization of the of abnormal spermatozoa has been the subject of several studies.6,7 56 The spermatoxic effects of gossypol appear to be related to both the dose and duration of 57 consumption of the phenolic compound produced by the cotton plant.8 The incidence of 58 gossypol-induced altered spermatogenesis in bulls is likely low due to the rumen’s ability 59 to detoxify the gossypol. However, high intake of free gossypol, albeit rare, can 60 overwhelm this system and create toxicosis.7 61 Genetic causes of sperm abnormalities are not as common as environmental 62 causes but are becoming more recognized because of improved diagnostic tools.9 These 63 defects have been shown to either consistently or occasionally have a genetic mode of 64 transmission. With the ability to scrutinize large numbers of AI sires and their progeny a 65 number of sperm defects have been classified as genetic in nature. 66 Distal midpiece reflex 67 The distal midpiece reflex (DMR) is the most common tail abnormality 68 encountered when evaluating the morphology of bull sperm.10 The typical appearance of 69 a DMR is that of a distinct hairpin bend of the tail at the location of the distal midpiece, 70 however there can be varying bending patterns noted in the tail giving the affected 71 spermatozoa several different appearances. A consistent finding is the presence of a 72 cytoplasmic droplet noted within the bend of the tail.10 When this defect is observed in 73 339 live samples, the spermatozoa will appear to be swimming in a reverse motion often in a 74 circular pattern. 10 75 The DMR defect can be induced experimentally when spermatozoa are exposed 76 to hypotonic solutions or when cooled very rapidly.10 This is critically important when 77 preparing a slide for staining. Many of the morphology stains used today (ie. eosin 78 nigrosin) are hypoosmotic and will create a similar defect if spermatozoa are exposed for 79 an extended time.10 It is critical that the slide be prepared properly and dried quickly so 80 as to reduce the chance of creating this defect iatrogenically. One striking difference in 81 the defects which occur naturally and those that are caused in vitro is the presence of a 82 cytoplasmic droplet within the reflex of the tail. When a large number of DMR defects 83 are noted without the concurrent presence of a cytoplasmic droplet the possibility of an 84 artificial cause should be investigated.10 85 The DMR defect is produced in the corpus and cauda epididymis where the 86 spermatozoa still have a distal cytoplamic droplet which is present in the bending of the 87 tail.10 The evidence which supports the epididymis as the origin of this defect is based on 88 appearance of this defect after known testicular insults. Semen evaluation of 606 bulls 89 prior to a severe snowstorm showed the percentage of bulls with greater than 15% DMR 90 to be only 10.9% whereas of the 117 bulls examined 3-4 days after the snowstorm 45.3% 91 had greater than 15% DMR defects.10 A significant increase in the DMR defect was 92 observed 6-12 days after a brief period of scrotal insulation.10 93 Other recognized causes of this defect include treatment with estrogens, induced 94 hypothyroidism and fever. All of these conditions have been shown to reduce 95 340 testosterone levels, which appears to adversely affect the epididymal environment leading 96 to the formation of the DMR defect.10 97 The fertility of ejaculates containing high numbers of spermatozoa with DMR 98 defects has not been critically evaluated however it has been observed that the defect 99 could be found in normal fertile bulls with a prevalence up to 25%.10 Owing to the fact 100 that this defect is of epididymal origin, it would seem that any effect on fertility would be 101 short-lived provided the inciting insult was removed. Since affected spermatozoa would 102 swim in a reverse fashion it is unlikely that they would be able to participate in 103 fertilization thereby could be compensated for by additional spermatozoa. There is no 104 evidence that spermatozoa with the DMR defect are capable of regaining normal 105 function.10 Semen evaluation in affected bulls should be performed often to detect 106 changes in prevalence of the DMR defect that could help identify an inciting cause. 107 Knobbed acrosome 108 The knobbed acrosome defect has been described as a refractile or dark-staining 109 area or eccentric thickening often giving a beaded appearance to the apex of affected 110 spermatozoa.10 More commonly however, it appears with the apex of the spermatozoa 111 having a flattened or indented acrosome.10 With the availability of electron microscopy, 112 studies have shown that with both the beaded and indented forms the abnormal acrosome 113 folds back on the sperm apex and a few affected spermatozoa may show a bead-like 114 protrusion from the apex of the spermatozoa. The folding back of the acrosome may also 115 result in a bending back of the apex of the nucleus which causes the apex to appear to 116 have an indentation.10,11 117 341 The knobbed acrosome defect results from altered spermatogenesis caused by 118 either environmental or genetic factors leading to abnormal development of the fine 119 structure of the plasma membrane making it more susceptible to structural and functional 120 changes.9,12 It has been shown that spermatozoa with the knobbed acrosome defect lack 121 membrane integrity which can lead to premature capacitation and subsequent acrosome 122 reaction however the exact mechanisms involved remain elusive.11,12 123 It is generally accepted that bulls whose semen contains a high percentage of 124 knobbed acrosome defects will have poor fertility.10 The exact mechanism by which this 125 defect causes reduced fertility has yet to be elucidated, however it has been theorized that 126 spermatozoa with abnormally shaped heads including the knobbed acrosome defect may 127 have altered motility characteristics which could impair the passage of spermatozoa 128 through the female reproductive tract.13 Spermatozoa containing the knobbed acrosome 129 defect could also have altered sperm-oocyte binding and zona penetration.13 In vitro 130 fertilization (IVF) models have shown that knobbed acrosome affected spermatozoa have 131 a reduced ability to bind the zona pellucida and could not penetrate the zona pellucida.11 132 Thus the effect of the knobbed acrosome defect on fertility appears to be related to both 133 altered passage as well as impaired plasma membrane function.12 In vitro studies also 134 indicate there could be compromised fertility associated with apparently normal 135 spermatozoa from males with many knobbed acrosome defects.14 These normal 136 appearing spermatozoa have been shown to undergo premature capacitation and have 137 spontaneous acrosome reaction as well as evidence of chromatin condensation.12 138 This sperm defect has been associated with infertility in a number of species 139 including bulls, boars, rams and stallions.11 In cattle this defect was first reported in the 140 342 Friesian breed in the 1940’s however it has since been seen in Charolais, Simmental, 141 Maine Anjou, Salers, Horned Hereford, Angus and Normande.9,15 This defect is 142 associated with an autosomal sex-linked mode of transmission in the Friesian breed of 143 cattle and in boars.9 There appears to be evidence to support a genetic cause of this 144 defect in both the Charolais breed and Angus cattle in North America. 9,15 145 Cytoplasmic droplet 146 The presence of a cytoplasmic droplet is common on a small number of 147 spermatozoa in the ejaculate of fertile bulls. The cytoplasmic droplet is a spherical mass 148 of cytoplasm that is typically found in one of two locations on the spermatozoa. It is 149 considered a proximal droplet when located in the proximal midpiece and a distal droplet 150 when surrounding the midpiece just proximal to the annulus.10 Droplets are rarely 151 observed in an intermediate location due to the rapid migration of the proximal droplet to 152 the distal location prior to shedding. 153 Cytoplasmic droplets are formed during spermiogenesis when the spermatid 154 changes from its round shape to an elongated shape and cytoplasm is pulled from the 155 head region towards the tail.10 During this transition the Sertoli cell molds this cytoplasm 156 into a lobule called the residual body.10 At the time of spermiation the stalk connecting 157 this residual body to the spermatid is severed and leaving the droplet of cytoplasm in the 158 proximal neck region of the spermatozoa.10 159 As spermatozoa enter the caput epididymis almost 85 percent will have a 160 proximal droplet. As spermatozoa move through the epididymis and maturation occurs 161 the proximal droplet moves to a distal location and is eventually shed. By the time the 162 spermatozoa reach the caudal epididymis over 60 percent of the spermatozoa have a 163 343 distal droplet.10 The loss of the cytoplasmic droplet appears to be associated with the 164 gaining of motility in the epididymis. 165 It is very common to find a high incidence of proximal droplets in bulls that are 166 approaching puberty, however with repeated collections over the following months the 167 percentage of affected spermatozoa generally drops substantially. In yearling bulls a 168 major cause of failure to pass a breeding soundness examination is often the presence of a 169 large percentage of proximal droplets. In data collection from Colorado, 12-26% of 170 yearling bulls failed to pass an initial breeding soundness examination with 6.3% of these 171 failures attributable to proximal droplets.16 As bulls mature however, the incidence of 172 cytoplasmic droplets tends to drop dramatically. When over 1500 bulls of various ages 173 and breeds were examined the percentage of bulls whose ejaculates contained proximal 174 droplets was 67 percent, with the number of affected spermatozoa averaging only 2.7 175 percent.10 In older bulls a high incidence of spermatozoa with proximal droplets points 176 towards abnormal spermiogenesis likely due to a degenerative process of the 177 seminiferous epithelium.10 178 The prognosis for bulls with a high percentage of spermatozoa with proximal 179 droplets varies depending on the underlying cause and the presence of other defects. 180 Recovery has been seen in bulls with profound disturbances of spermatogenesis leading 181 to ejaculates which contain greater than 50 percent of spermatozoa with proximal 182 droplets, however recovery often requires months.10 In vitro fertilization was used to 183 evaluate the fertilizing potential of semen from young bulls with a high incidence of 184 proximal droplets. It was concluded that fertility was severely compromised but as the 185 bulls matured the incidence of proximal droplets decreased and fertility increased.17 This 186 344 study also showed the fertilizing potential of a bull whose semen contained ≥30% 187 spermatozoa with proximal droplets will be low until the incidence of proximal droplets 188 decreases. 17 189 Distal droplets are not considered to be a major problem and often indicate 190 insufficient maturation in the epididymis. It has been noted that ejaculates containing a 191 high number of spermatozoa with distal droplets when allowed to incubate for several 192 minutes will almost be totally cleared of the defect.10 193 Crater or diadem defect 194 The crater or diadem defect is seen in spermatozoa which have a nuclear vacuole 195 or invagination of the nuclear membrane into the nucleoplasm which occurs during 196 spermiogenesis.18,19 This defect can appear as a “string of pearls” at the acrosome-197 postacrosomal sheath or as round to elongated white spots which often appear to 198 sparkle.10 The defect often appears as a surface oriented crater and can range from 1 to 199 over 20 in number.10 This particular defect is often overlooked with light microscopy on 200 routine eosin-nigrosin stained spermiograms, whereas phase-contrast microscopy and 201 differential interference microscopy allow one to more easily detect this defect. The use 202 of a nucleus stain such as the Feulgen stain should also allow excellent visualization of 203 the nucleus and associated vacuoles.10 204 The incidence of the crater or diadem defect in bulls is generally very high, often 205 approaching 100%; however the percentage of affected spermatozoa within an ejaculate 206 can vary greatly. In a Czechoslovakian study of young and older bulls, nuclear vacuoles 207 were observed in all bulls with the incidence of affected spermatozoa ranging from 3-208 345 26%.10 In one study of bulls at an AI center, all bulls had some defects and 28% of the 209 bulls examined had an incidence of crater defects in excess of 20%.19 210 The precise pathogenesis of this defect has not been elucidated however there are 211 several theories put forth in the literature. Coulter suggested a possible viral etiology 212 based on the presence of viral-like particles within the vacuoles with the thought that the 213 virus may attack the developing spermatozoa.10 Others have shown that certain 214 insecticides when administered to bulls resulted in an increased incidence of nuclear 215 vacuoles.10 The most common and accepted theory involves the impact of stress on the 216 process of spermatogenesis. It has clearly been shown that administration of 217 dexamethasone to bulls to mimic a stressful event will significantly reduce their LH and 218 testosterone levels and is thought to impair spermatogenesis and lead to the formation of 219 defective sperm. Studies of the sequential appearance of sperm defects after 220 administration of dexamethasone to bulls clearly show there is an increase in the presence 221 of nuclear vacuoles with the peak incidence occurring around 21 days after 222 dexamethasone treatment.1 Whether the effects of dexamethasone on spermatogenesis 223 are direct or indirect have yet to be determined. When bulls were studied over long 224 periods of time, fluctuations were seen in the incidence of affected spermatozoa 225 indicating a nonheritable etiology. This is supported by the absence of the defect in 226 significant numbers in the ejaculates of bulls which are sons of a known affected bull.10 227 However there is reason to believe that there may be bulls which have a heritable 228 predisposition to produce spermatozoa with the crater defect in response to stress.10 229 It is clear that ejaculates containing high numbers of spermatozoa with nuclear 230 vacuoles are a cause of infertility in the bull. Breeding trials have shown that 231 346 spermatozoa with multiple nuclear vacuoles have reduced fertilization characteristics 232 both in vivo and in vitro.18,20 It appears that affected spermatozoa have a reduced 233 capacity to penetrate the zona pellucida as well as reduced ability to form a male 234 pronucleus.18 With this knowledge, bulls whose semen contains high numbers of 235 spermatozoa with nuclear vacuoles should be monitored with successive semen 236 evaluations as it has been reported that occasionally bulls recover and regain normal 237 fertility.10 238 Dag defect 239 The Dag defect traces back to a Jersey bull with this name in which this unique 240 defect was first discovered and reported. This heritable defect which occurs during late 241 spermiogenesis is characterized by severe coiling of the tail with fracture of the distal part 242 of the midpiece.10 A distal cytoplasmic droplet can also be seen associated with this 243 defect.9 Other features which may be seen include a roughened appearance to the 244 mitochondrial sheath and fracturing of the axonal elements leading to disruption of the 245 mitochondrial arrangement.10 This defect has been reported as a cause of infertility in 246 several breeds including the Jersey, Polled and Horned Hereford, and the Swedish Red 247 and White. In these particular infertile bulls the incidence of the Dag defect was 248 generally greater than 50%. This defect can be found in the ejaculates of bulls with 249 normal fertility however the incidence of affected spermatozoa rarely exceeds 5%.10 250 When the incidence of this defect approaches 50% there appears to be a profound impact 251 on fertility. 10 252 A genetic basis for this defect was proven in the Danish Jersey breed through 253 selected breeding of a suspected carrier to 120 of his daughters which produced 6 bulls 254 347 with the typical defective spermatoazoa.10 There may be breed specific differences in the 255 exact pathogenesis of this defect because investigators have noted distinct differences in 256 the microanatomy of affected spermatozoa from various breeds. 10 257 Pyriform or tapered heads 258 The pyriform defect is the most common defect of head shape and it appears as a 259 pear-shaped head with a pronounced narrowing of the postacrosomal area.10 There are 260 many variations of the pyriform defect that range from almost imperceptible to those with 261 severe narrowing all of which can occur in the same ejaculate. The tapered head shape is 262 slightly different than the pyriform head shape in that the entire nucleus is narrow and the 263 head appears elongated.10 Both of these head defects can be found in the same ejaculate 264 and affected spermatozoa always appear smaller than their normal counterparts. These 265 two defects appear closely related in origin and may be categorized together.10 266 The incidence of the pyriform or tapered head defect occurring in more than 15% 267 of spermatozoa in an evaluation of over 1300 range beef bulls was determined to be 268 8.7%. This was was lower than the 16.4% incidence found in another study of 216 dairy 269 and beef AI sires.10 The presence of this defect in low numbers however is a fairly 270 common finding in the ejaculate of many fertile bulls. 271 The pathogenesis of the pyriform or tapered head defect has not been proven but 272 appears to occur secondary to some disturbance of thermoregulation or an endocrine 273 aberration leading to impaired testicular function.10 Over-conditioned bulls which have 274 excess fat in their inguinal and scrotal areas commonly have these defects. The 275 appearance of pyriform head defects in experimentally-induced scrotal insulation peaked 276 348 at 22 days post insulation and at 24 days in dexamethasone-treated bulls suggesting the 277 damaging effects occurred during nucleus condensation and shaping.1 278 The effects of spermatozoa with pyriform or tapered heads on fertility appear to 279 be related to their reduced ability to bind and penetrate the zona pellucida however there 280 does not appear to be an increased incidence of embryo or fetal loss provided fertilization 281 occurs.10 Work by Saacke investigating the accessory sperm population has also shown 282 that the severity of the head defect may dictate the accessibility of spermatozoa to the 283 ovum thereby limiting the possibility of zona binding.21,22 284 The prognosis for bulls exhibiting large numbers of pyriform or tapered heads in 285 their ejaculates varies depending on the underlying cause. When a cause such as altered 286 testicular thermoregulation can be identified and corrected, the prognosis is generally 287 good provided enough time is allowed to resume normal spermatogenesis. However in 288 bulls which have high numbers of pyriform or tapered heads in which there is no 289 apparent reason for the altered spermatogenesis the prognosis is generally poor for 290 recovery.10 The percentage of affected spermatozoa can generally be used as a 291 prognostic indicator. 292 Detatched heads 293 The presence of detached heads on routine evaluation of a bull’s spermiogram is 294 relatively common. This defect is easy to identify during evaluation of a sperm 295 morphology slide. The incidence of this defect in bulls of normal fertility and various 296 ages appears to be around 5 percent.10 A number of conditions have been associated with 297 the presence of increased numbers of detached heads. In one study of eight bulls with 298 testicular hypoplasia the incidence of detached heads ranged between 39 and 93 percent. 299 349 However, of these eight bulls, seven were traced back to a common ancestor and the 300 possibility that the high incidence of detached heads was inherited could not be 301 overlooked.10 The relationship between detached heads and testicular hypoplasia is not 302 consistent among all bulls with this condition. In a study of 141 bulls with testicular 303 hypoplasia the percentage of detached heads was only 6.9 percent.10 304 There appears to be a condition in bulls that mimics the clinical appearance of 305 “plugged ampullae” in stallions. The incidence of this condition was found to be 1.1% 306 when over 1300 bulls were evaluated. The characteristics of the ejaculate consist of a 307 very large volume of highly concentrated semen which contains a high proportion of 308 dead spermatozoa as well as 15-45 percent detached heads.10 These bulls appeared to 309 improve with frequent repeated collections however the incidence of this defect would be 310 higher after periods of sexual rest. The most common theory is that these bulls have a 311 failure of normal sperm transport with accumulation of spermatozoa within the 312 epididymis and ampullae.10 313 Other conditions that have been shown to cause a transient increase in the number 314 of detached heads include: seminal vesiculitis, epididymitis, or any condition which leads 315 to failure of normal testicular thermoregulation. Lameness which causes the bull to lie 316 down for long periods of time with resultant failure of normal thermoregulation is known 317 to increase the number of detached heads.10 318 Abaxial attachment of the tail 319 Abaxial attachment of the tail is an infrequently encountered defect of bull 320 spermatozoa. In a retrospective analysis of semen analysis from 1049 range and AI bulls 321 the percentage of bulls found to have the defect was 10.5% and only 0.48% of bulls had 322 350 semen with greater than 50% abaxial attachment of the head.23 It is considered a normal 323 finding in the boar and stallion however its significance in the bull remains unclear. 324 There have been reports of sterile bulls that had an increased number of spermatozoa with 325 abaxial attachment of the head however there were no controlled breeding trials 326 performed.23 In three controlled experiments comparing the fertility of bulls with high 327 numbers of abaxial tail attachement to that of known fertile bulls there was no difference 328 in all fertility parameters measured between bulls.23 The combined results of these 329 experiments indicate that abaxial attachment of the tail does not impact fertility and 330 should be considered a normal morphological variation of bovine spermatozoa.23 331 Conclusion 332 There has been a strong correlation between morphologically abnormal sperm and 333 some degree of infertility for many years. It is important to not only be able to recognize 334 morphological defects of spermatozoa but also to provide the client with possible 335 etiologic causes of the defects observed and make suggestions as to their impact on 336 fertility. With the use of routine staining procedures most defects can be observed when 337 examined at 1000X with light microscopy. Electron microscopy may more accurately 338 identify the exact defect present in affected spermatozoa allowing a more in-depth 339 understanding of the sperm abnormalities. 340 References 341 342 1. Barth AD, Bowman PA: The sequential appearance of sperm abnormalities after 343 scrotal insulation or dexamethasone treatment in bulls. Can Vety J 1994;34:93-344 101. 345 351 2. Senger PL: Pathways to pregnancy and parturition. 2nd ed. Pullman, WA: Current 346 Conceptions; 2003. 347 3. Kastelic JP, Cook RB, Coulter GH, et al: Insulating the scrotal neck affects semen 348 quality and scrotal/testicular temperatures in the bull. Theriogenology 349 1996;45:935-942. 350 4. Vogler CJ, Bame JH, DeJarnette JM, et al: Effects of elevated testicular 351 temperature on morphology characteristics of ejaculatated spermatozoa in the 352 bovine. Theriogenology 1993;40:1207-1219. 353 5. Welsh TH, Johnson BH: Stress induced alterations in secretion of corticosteroids, 354 progesterone, lutenizing hormone and testosterone in bulls. Endocrinology 355 1981;109:185-190. 356 6. Chenoweth PJ, Chase CC, Risco CA, et al: Characterization of gossypol-induced 357 sperm abnormalities in bulls. Theriogenology 2000;53:1193-1203. 358 7. Chenoweth PJ, Risco CA, Larsen RE, et al: Effects of dietary gossypol on aspects 359 of semen quality, sperm morphology and sperm production in young brahman 360 bulls. Theriogenology 1994;42:1-13. 361 8. Randel RD, Chase CC, Wyse SJ: Effects of gossypol and cottonseed products on 362 reproduction of mammals. J Anim Sci 1992;70:1628-1638. 363 9. Chenoweth PJ: Genetic sperm defects. Theriogenology 2005;64:457-468. 364 10. Barth AD, Oko RJ: Abnormal morphology of bovine spermatozoa. Ames, IA: 365 Iowa State Univ Press; 1989. 366 11. Thundathil J, Meyer R, Palasz AT, et al: Effect of the knobbed acrosome defect in 367 bovine sperm on IVF and embryo production. Theriogenology 2000;54:921-934. 368 352 12. Thundathil J, Palasz AT, Barth AD, et al: Plasma membrane and acrosomal 369 integrity in bovine spermatozoa with knobbed acrosome defect. Theriogenology 370 2002;58:87-102. 371 13. Saacke RG, Nadir S, Nbel RL: Relationship of semen quality to sperm transport, 372 fertility and embryo quality in ruminants. Theriogenology 1994;41:45-50. 373 14. Soderquist L: Reduced fertility after artificial insemination in a ram with a high 374 incidence of knobbed acrosomes. Vet Rec 1998;143:227-228. 375 15. Barth AD: The knobbed acrosome defect in beef bulls. Can Vet J 1986;27:379-376 384. 377 16. Molnar E: Evaluation of breeding soundness exam in performance-tested beef 378 bulls: Colorado State University, 1989. 379 17. Amann RP, Seidel GE, Mortimer RG: Fertilizing potential in vitro of semen from 380 young beef bulls containing a high or low percentage of sperm with a proximal 381 droplet. Theriogenology 2000;54:1499-1515. 382 18. Pilip R, Campo MRD, Barth AD, et al: In vitro fertilization characteristics of 383 bovine spermatozoa with multiple nuclear vacuoles: a case study. Theriogenology 384 1996;46:1-12. 385 19. Coulter GH, Oko RJ, Costerton JW: Incidence and ultrastructure of "crater" defect 386 of bovine spermatozoa. Theriogenology 1978;9:165-171. 387 20. Miller BM, Hrudka F, Cates WF, et al: Infertility in a bull with a nuclear sperm 388 defect: a case report. Theriogenology 1982;17:611-619. 389 353 21. Saacke RG, DeJarnette JM, Barne JH, et al: Can spermatozoa with abnormal 390 heads gain access to the ovum in artificially inseminated super- and single-391 ovulating cattle? Theriogenology 1998;50:117-128. 392 22. Evenson DP: Loss of livestock breeding efficiency due to uncompensable sperm 393 nuclear defects. Reprod Fertil Dev 1999;11:1-15. 394 23. Barth AD: Abaxial tail attachment of bovine spermatozoa and its effect on 395 fertility. 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