Technical Aspects and Clinical Limitations of Sperm DNA Fragmentation Testing in Male Infertility: A Global Survey, Current Guidelines, and Expert Recommendations.

PURPOSE: Sperm DNA fragmentation (SDF) is a functional sperm abnormality that can impact reproductive potential, for which four assays have been described in the recently published sixth edition of the WHO laboratory manual for the examination and processing of human semen. The purpose of this study was to examine the global practices related to the use of SDF assays and investigate the barriers and limitations that clinicians face in incorporating these tests into their practice. MATERIALS AND METHODS: Clinicians managing male infertility were invited to complete an online survey on practices related to SDF diagnostic and treatment approaches. Their responses related to the technical aspects of SDF testing, current professional society guidelines, and the literature were used to generate expert recommendations via the Delphi method. Finally, challenges related to SDF that the clinicians encounter in their daily practice were captured. RESULTS: The survey was completed by 436 reproductive clinicians. Overall, terminal deoxynucleotidyl transferase deoxyuridine triphosphate Nick-End Labeling (TUNEL) is the most commonly used assay chosen by 28.6%, followed by the sperm chromatin structure assay (24.1%), and the sperm chromatin dispersion (19.1%). The choice of the assay was largely influenced by availability (70% of respondents). A threshold of 30% was the most selected cut-off value for elevated SDF by 33.7% of clinicians. Of respondents, 53.6% recommend SDF testing after 3 to 5 days of abstinence. Although 75.3% believe SDF testing can provide an explanation for many unknown causes of infertility, the main limiting factors selected by respondents are a lack of professional society guideline recommendations (62.7%) and an absence of globally accepted references for SDF interpretation (50.3%). CONCLUSIONS: This study represents the largest global survey on the technical aspects of SDF testing as well as the barriers encountered by clinicians. Unified global recommendations regarding clinician implementation and standard laboratory interpretation of SDF testing are crucial.

Controversy and Consensus on Indications for Sperm DNA Fragmentation Testing in Male Infertility: A Global Survey, Current Guidelines, and Expert Recommendations.

PURPOSE: Sperm DNA fragmentation (SDF) testing was recently added to the sixth edition of the World Health Organization laboratory manual for the examination and processing of human semen. Many conditions and risk factors have been associated with elevated SDF; therefore, it is important to identify the population of infertile men who might benefit from this test. The purpose of this study was to investigate global practices related to indications for SDF testing, compare the relevant professional society guideline recommendations, and provide expert recommendations. MATERIALS AND METHODS: Clinicians managing male infertility were invited to take part in a global online survey on SDF clinical practices. This was conducted following the CHERRIES checklist criteria. The responses were compared to professional society guideline recommendations related to SDF and the appropriate available evidence. Expert recommendations on indications for SDF testing were then formulated, and the Delphi method was used to reach consensus. RESULTS: The survey was completed by 436 experts from 55 countries. Almost 75% of respondents test for SDF in all or some men with unexplained or idiopathic infertility, 39% order it routinely in the work-up of recurrent pregnancy loss (RPL), and 62.2% investigate SDF in smokers. While 47% of reproductive urologists test SDF to support the decision for varicocele repair surgery when conventional semen parameters are normal, significantly fewer general urologists (23%; p=0.008) do the same. Nearly 70% would assess SDF before assisted reproductive technologies (ART), either always or for certain conditions. Recurrent ART failure is a common indication for SDF testing. Very few society recommendations were found regarding SDF testing. CONCLUSIONS: This article presents the largest global survey on the indications for SDF testing in infertile men, and demonstrates diverse practices. Furthermore, it highlights the paucity of professional society guideline recommendations. Expert recommendations are proposed to help guide clinicians.

Controversy and Consensus on the Management of Elevated Sperm DNA Fragmentation in Male Infertility: A Global Survey, Current Guidelines, and Expert Recommendations.

PURPOSE: Sperm DNA fragmentation (SDF) has been associated with male infertility and poor outcomes of assisted reproductive technology (ART). The purpose of this study was to investigate global practices related to the management of elevated SDF in infertile men, summarize the relevant professional society recommendations, and provide expert recommendations for managing this condition. MATERIALS AND METHODS: An online global survey on clinical practices related to SDF was disseminated to reproductive clinicians, according to the CHERRIES checklist criteria. Management protocols for various conditions associated with SDF were captured and compared to the relevant recommendations in professional society guidelines and the appropriate available evidence. Expert recommendations and consensus on the management of infertile men with elevated SDF were then formulated and adapted using the Delphi method. RESULTS: A total of 436 experts from 55 different countries submitted responses. As an initial approach, 79.1% of reproductive experts recommend lifestyle modifications for infertile men with elevated SDF, and 76.9% prescribe empiric antioxidants. Regarding antioxidant duration, 39.3% recommend 4-6 months and 38.1% recommend 3 months. For men with unexplained or idiopathic infertility, and couples experiencing recurrent miscarriages associated with elevated SDF, most respondents refer to ART 6 months after failure of conservative and empiric medical management. Infertile men with clinical varicocele, normal conventional semen parameters, and elevated SDF are offered varicocele repair immediately after diagnosis by 31.4%, and after failure of antioxidants and conservative measures by 40.9%. Sperm selection techniques and testicular sperm extraction are also management options for couples undergoing ART. For most questions, heterogenous practices were demonstrated. CONCLUSIONS: This paper presents the results of a large global survey on the management of infertile men with elevated SDF and reveals a lack of consensus among clinicians. Furthermore, it demonstrates the scarcity of professional society guidelines in this regard and attempts to highlight the relevant evidence. Expert recommendations are proposed to help guide clinicians.

Sperm Chromatin Structure Assay (SCSA®) for Fertility Assessment.

The Sperm Chromatin Structure Assay (SCSA® ) is a federally registered protocol for simultaneous flow cytometric measures of sperm DNA integrity and chromatin structure. Fresh or frozen/thawed raw semen samples are diluted in buffer to a sperm concentration of ∼1-2×106 /ml and then treated with a pH 1.20 buffer for 30 s to open the DNA strands at sites of DNA strand breaks. The sperm are then stained with acridine orange (AO) that intercalates into double-strand DNA and fluoresces green (515-530 BP filter) and stacks on single-strand DNA that fluoresces red (630 LP filter) upon excitation from a 488 nm laser. The extent of single and double DNA strand breaks (DNA fragmentation index, %DFI) and level of excess nuclear histones (high DNA stainable sperm, %HDS) are simultaneously measured in individual sperm. From the time a fresh or frozen/thawed semen sample is received at the site of a flow cytometer (FCM) programmed for the SCSA protocol, data can be obtained within about 10 min on 5-10×103 sperm. The %DFI and %HDS can be determined by computer-gated regions on the green versus red cytogram. Alternatively, a determination is made by transforming the green versus red cytogram to a total DNA stainability (red + green fluorescence) versus red/red + green fluorescence cytogram from which a frequency histogram is produced and the %DFI calculated from it. The clinical threshold for human natural or IUI fertilization is 25% DFI at which point the ART lab should consider moving to ICSI fertilization. The clinical threshold for HDS is also 25%; values above this level may result in early embryo death due to abnormal gene readout caused by the abnormal tertiary structure of chromatin. Numerous lifestyle and environmental factors cause sperm DNA fragmentation. Reactive oxygen species (ROS) play a significant role in DNA breakage. © 2022 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Sperm Chromatin Structure Assay (SCSA®) Basic Protocol 2: SCSA data analysis: Calculations of %DFI and %HDS of semen samples by one of two methods Support Protocol 1: SCSA sample collection and shipping Support Protocol 2: Flow cytometer set up Support Protocol 3: Selection and use of reference samples.

A Comprehensive Guide to Sperm Recovery in Infertile Men with Retrograde Ejaculation.

Retrograde ejaculation (RE) is a condition defined as the backward flow of the semen during ejaculation, and when present can result in male infertility. RE may be partial or complete, resulting in either low seminal volume or complete absence of the ejaculate (dry ejaculate). RE can result from anatomic, neurological or pharmacological conditions. The treatment approaches outlined are determined by the cause. Alkalinizing urinary pH with oral medications or by adding sperm wash media into the bladder prior to ejaculation may preserve the viability of the sperm. This article provides a step-by-step guide to diagnose RE and the optimal techniques to retrieve sperm.

Sperm Morphology Assessment in the Era of Intracytoplasmic Sperm Injection: Reliable Results Require Focus on Standardization, Quality Control, and Training.

Semen analysis is the first, and frequently, the only step in the evaluation of male fertility. Although the laboratory procedures are conducted according to the World Health Organization (WHO) guidelines, semen analysis and especially sperm morphology assessment is very difficult to standardize and obtain reproducible results. This is mainly due to the highly subjective nature of their evaluation. ICSI is the choice of treatment when sperm morphology is severely abnormal (teratozoospermic). Hence, the standardization of laboratory protocols for sperm morphology evaluation represents a fundamental step to ensure reliable, accurate and consistent laboratory results that avoid misdiagnoses and inadequate treatment of the infertile patient. This article aims to promote standardized laboratory procedures for an accurate evaluation of sperm morphology, including the establishment of quality control and quality assurance policies. Additionally, the clinical importance of sperm morphology results in assisted reproductive outcomes is discussed, along with the clinical management of teratozoospermic patients.

Sperm DNA fragmentation testing: Summary evidence and clinical practice recommendations.

We herein summarise the evidence concerning the impact of sperm DNA fragmentation in various clinical infertility scenarios and the advances on sperm DNA fragmentation tests. The collected evidence was used to formulate 41 recommendations. Of these, 13 recommendations concern technical aspects of sperm DNA fragmentation testing, including pre-analytical information, clinical thresholds and interpretation of results. The remaining 28 recommendations relate to indications for sperm DNA fragmentation testing and clinical management. Clinical scenarios like varicocele, unexplained infertility, idiopathic infertility, recurrent pregnancy loss, intrauterine insemination, in vitro fertilisation/intracytoplasmic sperm injection, fertility counselling for men with infertility risk factors and sperm cryopreservation have been contemplated. The bulk evidence supporting the recommendations has increased in recent years, but it is still of moderate to low quality. This guideline provides clinicians with advice on best practices in sperm DNA fragmentation testing. Also, recommendations are provided on possible management strategies to overcome infertility related to sperm DNA fragmentation, based on the best available evidence. Lastly, we identified gaps in knowledge and opportunities for research and elaborated a list of recommendations to stimulate further investigation.

Relationships between the age of 25,445 men attending infertility clinics and sperm chromatin structure assay (SCSA®) defined sperm DNA and chromatin integrity.

OBJECTIVE: To determine relationships between age of men with potential male factor infertility and sperm chromatin structure assay (SCSA) measures of sperm DNA fragmentation (SDF) and high DNA stainable sperm (HDS), and to compare these data with those obtained from healthy donor men without reproductive issues. DESIGN: Retrospective study. SETTING: Infertility clinics and diagnostic laboratory. PATIENTS: A total of 25,445 men attending infertility clinics. Donors were 87 men working at Lawrence Livermore National Laboratory. INTERVENTION: None. MAIN OUTCOME MEASURES: SCSA measures (% DNA fragmentation index (DFI), X DFI, SD DFI, and %HDS) of men aged 21-80 years. RESULTS: In the study population, advancing paternal age was associated with increased sperm DNA fragmentation (SDF) scored as increased percentage of sperm in semen ejaculates with measurable DNA strand breaks (%DFI). The slope of increase in %DFI prior to age 41.6 years was 0.39, which increased after age 41.6 to more than double at a slope of 0.86. These changes in DNA/chromatin in more than 25,000 aging men attending infertility clinics are similar to those seen over the same age span (20-80 years) in 87 nonpatient, healthy men without reproductive issues. For the age group 20-50 years, there was no major significant difference in %DFI between patients and donor men. According to a logistic regression model, the estimated probability is that, for example, a 40-year-old and a 50-year-old man have a 20% and 40% chance, respectively, to have a pathological DFI ≥25% by age factor alone. The condensation of sperm chromatin in patients increased with age in a linear fashion, from a mean of 12.2 %HDS at age 20-25 to a mean of 7.9 %HDS at age 60-65. Patients had a greater %HDS than donors across all ages. CONCLUSIONS: The great heterogeneity of both DFI and HDS values at a specific age prevents the automatic translation of age into an index of DNA fragmentation. However, it reinforces the idea that both DFI and HDS evaluation can play a role in detecting potential male infertility in cases that are not resolved by routine testing and in cases of multiple miscarriages. DFI and HDS data can help clinicians to predict a man's fertility potential, to consider corrective therapeutic approaches, as well as to assess the risk to the offspring's health.

Melatonin and Caffeine Supplementation Used, Respectively, as Protective and Stimulating Agents in the Cryopreservation of Human Sperm Improves Survival, Viability, and Motility after Thawing compared to Traditional TEST-Yolk Buffer.

Cryopreservation processes can damage spermatozoa and impair structural and functional cell characteristics. Plasma, nuclear membranes, and cellular organelles can suffer from the freeze and thaw process. This study evaluates the protective and stimulant effect of melatonin and caffeine supplementation on the functional characteristics of human spermatozoa before and after freezing. Thirty seminal samples from normozoospermic men aged 19-45 years old collected between October 2012 and May 2017 were included. Semen samples were supplemented with either 2 mM melatonin (MEL) prior to cryopreservation, 2 mM caffeine (CAF) in postthaw, or CAF and MEL (CM) in precryopreservation and postthaw, respectively. Kinetics and seminal parameters, mitochondrial activity, DNA fragmentation, and reactive oxygen species (ROS) levels were analyzed before and after cryopreservation. A significant reduction in sperm concentration, total and progressive motility, sperm kinetics, and mitochondrial activity, as well as a significant increase in DNA fragmentation and ROS production in postthaw samples compared to fresh samples, was identified. After administration of a caffeine and/or melatonin supplement, there was a significant increase in progressive motility in the CAF (p = 0.005) and CM (p = 0.048) groups, as well as mitochondrial activity in the CM group (p < 0.05). Cryopreservation has negative effects on overall sperm quality and increases ROS production. A combination of caffeine and melatonin in prefreeze and postthaw sperm samples has proven to be a very effective and simple way to improve semen quality. This will be particularly useful for initial low-quality semen samples, those which suffer the most from the freezing/thawing process.

The Sperm Chromatin Structure Assay (SCSA(®)) and other sperm DNA fragmentation tests for evaluation of sperm nuclear DNA integrity as related to fertility.

Thirty-five years ago the pioneering paper in Science (240:1131) on the relationship between sperm DNA integrity and pregnancy outcome was featured as the cover issue showing a fluorescence photomicrograph of red and green stained sperm. The flow cytometry data showed a very significant difference in sperm DNA integrity between fertile and subfertile bulls and men. This study utilized heat (100°C, 5min) to denature DNA at sites of DNA strand breaks followed by staining with acridine orange (AO) and measurements of 5000 individual sperm of green double strand (ds) DNA and red single strand (ss) DNA fluorescence. Later, the heat protocol was changed to a low pH protocol to denature the DNA at sites of strand breaks; the heat and acid procedures produced the same results. SCSA data are very advantageously dual parameter with 1024 channels (degrees) of both red and green fluorescence. Hundreds of publications on the use of the SCSA test in animals and humans have validated the SCSA as a highly useful test for determining male breeding soundness. The SCSA test is a rapid, non-biased flow cytometer machine measurement providing robust statistical data with exceptional precision and repeatability. Many genotoxic experiments showed excellent dose response data with very low coefficient of variation that further validated the SCSA as being a highly powerful assay for sperm DNA integrity. Twelve years following the introduction of the SCSA test, the terminal deoxynucleotidyl transferase-mediated fluorescein-dUTP nick end labelling (TUNEL) test (1993) for sperm was introduced as the only other flow cytometric assay for sperm DNA fragmentation. However, the TUNEL test can also be done by light microscopy with much less statistical robustness. The COMET (1998) and Sperm Chromatin Dispersion (SCD; HALO) (2003) tests were introduced as light microscope tests that don't require a flow cytometer. Since these tests measure only 50-200 sperm per sample, they suffer from the lack of the statistical robustness of flow cytometric measurements. Only the SCSA test has an exact standardization of a fixed protocol. The many variations of the other tests make it very difficult to compare data and thresholds for risk of male factor infertility. Data from these four sperm DNA fragmentation tests plus the light microscope acridine orange test (AOT) are correlated to various degrees.

Luminal fluid of epididymis and vas deferens contributes to sperm chromatin fragmentation.

STUDY QUESTION: Do the luminal fluids of the epididymis and the vas deferens contribute to sperm chromatin fragmentation (SCF) in mice? SUMMARY ANSWER: The luminal fluids of both organs are required for activating SCF in mice, but the vas deferens luminal fluid does this more efficiently than that of the epididymis. WHAT IS KNOWN ALREADY: Mice sperm have the ability to degrade their DNA in an apoptotic-like fashion when treated with divalent cations in a process termed SCF. SCF has two steps: the induction of reversible double-strand DNA breaks at the nuclear matrix attachment sites, followed by the irreversible degradation of DNA by nuclease. Single stranded DNA breaks accompany SCF. STUDY DESIGN, SIZE, DURATION: Luminal fluids from two reproductive organs of the mouse (B6D2F1 strain), the epididymis and vas deferens, were extracted and tested for SCF activation with divalent cations using four different combinations of the sperm and the surrounding luminal fluids: (i) in situ--sperm were kept in their luminal fluid and activated directly; (ii) reconstituted--sperm were centrifuged and resuspended in their luminal fluid before SCF activation; (iii) mixed--sperm were centrifuged and resuspended in the luminal fluid of the other organ; (iv) no luminal fluid--sperm were centrifuged and reconstituted in buffer. All four experiments were performed without (controls) and with divalent cations (resulting in SCF). For each experimental condition, two different mice were used and the analyses averaged. PARTICIPANTS/MATERIALS, SETTING, METHODS: DNA damage by SCF was analyzed by three different methods, the sperm chromatin structure assay (SCSA), terminal deoxynucleotidyltransferase-mediated dUTP nick-end labeling (TUNEL) analysis and field inversion gel electrophoresis. MAIN RESULTS AND THE ROLE OF CHANCE: In all three assays that we used, the vas deferens luminal fluid was much more efficient in stimulating SCF in the sperm from either source than that of the epididymis (P < 0.0001). Vas deferens sperm were capable of initiating lower levels of SCF in the absence of luminal fluid (P < 0.0001). LIMITATIONS, REASONS FOR CAUTION: Analyses were performed in only one species, the mouse, but we used three separate assays in our analysis. WIDER IMPLICATIONS OF THE FINDINGS: The data suggest that the luminal fluid of the male reproductive tract interacts with sperm during their transit providing a mechanism to degrade the DNA. We hypothesize that this is part of an apoptotic-like mechanism that allows the reproductive tract to eliminate defective sperm. The SCF model also allowed us to identify differences in the types of DNA lesions that the three tests can identify, providing important background information for the use of these tests clinically.

The impact of sperm DNA damage in assisted conception and beyond: recent advances in diagnosis and treatment.

Sperm DNA damage is a useful biomarker for male infertility diagnosis and prediction of assisted reproduction outcomes. It is associated with reduced fertilization rates, embryo quality and pregnancy rates, and higher rates of spontaneous miscarriage and childhood diseases. This review provides a synopsis of the most recent studies from each of the authors, all of whom have major track records in the field of sperm DNA damage in the clinical setting. It explores current laboratory tests and the accumulating body of knowledge concerning the relationship between sperm DNA damage and clinical outcomes. The paper proceeds to discuss the strengths, weaknesses and clinical applicability of current sperm DNA tests. Next, the biological significance of DNA damage in the male germ line is considered. Finally, as sperm DNA damage is often the result of oxidative stress in the male reproductive tract, the potential contribution of antioxidant therapy in the clinical management of this condition is discussed. DNA damage in human spermatozoa is an important attribute of semen quality. It should be part of the clinical work up and properly controlled trials addressing the effectiveness of antioxidant therapy should be undertaken as a matter of urgency. Sperm DNA damage is a useful biomarker for male infertility diagnosis and prediction of assisted reproduction outcomes. It is associated with reduced fertilization rates, embryo quality and pregnancy rates, and higher rates of spontaneous miscarriage and childhood diseases. With all of these fertility check points, it shows more promise than conventional semen parameters from a diagnostic perspective. Despite this, few infertility clinics use it routinely. This review provides a synopsis of the most recent studies from each of the authors, all of whom have major track records in the field of sperm DNA damage in the clinical setting. It explores current laboratory tests and the accumulating body of knowledge concerning the relationship between sperm DNA damage and clinical outcomes. The paper proceeds to discuss the strengths and weaknesses and clinical applicability of current sperm DNA fragmentation tests. Next, the biological significance of DNA damage in the male germ line is considered. Finally, as sperm DNA damage is often the result of increased oxidative stress in the male reproductive tract, the potential contribution of antioxidant therapy in the clinical management of this condition is discussed. As those working in this field of clinical research, we conclude that DNA damage in human spermatozoa is an important attribute of semen quality which should be carefully assessed in the clinical work up of infertile couples and that properly controlled trials addressing the effectiveness of antioxidant therapy should be undertaken as a matter of urgency.

Sperm chromatin structure assay (SCSA®).

The SCSA(®) is the pioneering assay for the detection of damaged sperm DNA and altered proteins in sperm nuclei via flow cytometry of acridine orange (AO) stained sperm. The SCSA(®) is considered to be the most precise and repeatable test providing very unique, dual parameter data (red vs. green fluorescence) on a 1,024 × 1,024 channel scale, not only on DNA fragmentation but also on abnormal sperm characterized by lack of normal exchange of histones to protamines. Raw semen/sperm aliquots or purified sperm can be flash frozen, placed in a box with dry ice and shipped by overnight courier to an experienced SCSA(®) lab. The samples are individually thawed, prepared, and analyzed in ∼10 min. Of significance, data on 5,000 individual sperm are recorded on a 1,024 × 1,024 dot plot of green (native DNA) and red (broken DNA) fluorescence. Repeat measurements have virtually identical dot plot patterns demonstrating that the low pH treatment that opens up the DNA strands at the sites of breaks and staining by acridine orange (AO) are highly precise and repeatable (CVs of 1-3%) and the same between fresh and frozen samples. SCSAsoft(®) software transforms the X-Y data to total DNA stainability versus red/red + green fluoresence (DFI) providing a more accurate determination of % DFI as well as the more sensitive value of standard deviation of DFI (SD DFI) as demonstrated by animal fertility and dose-response toxicology studies. The current established clinical threshold is 25% DFI for placing a man into a statistical probability of the following: (a) longer time to natural pregnancy, (b) low odds of IUI pregnancy, (c) more miscarriages, or (d) no pregnancy. Changes in lifestyle as well as medical intervention can lower the %DFI to increase the probability of natural pregnancy. Couples of men with >25% DFI are counseled to try ICSI and when in the >50% range may consider TESE/ICSI. The SCSA(®) simultaneously determines the % of sperm with high DNA stainability (%HDS) related to retained nuclear histones consistent with immature sperm; high HDS values are predictive of pregnancy failure.The SCSA(®) is considered to be the most technician friendly, time- and cost-efficient, precise and repeatable DNA fragmentation assay, with the most data and the only fragmentation assay with an accepted clinical threshold for placing a man at risk for infertility. SCSA(®) data are more predictive of male factor infertility than classical semen analyses.

High levels of DNA fragmentation in spermatozoa are associated with inbreeding and poor sperm quality in endangered ungulates.

Inbreeding is known to cause deleterious effects upon reproduction and survival, but its effects upon sperm DNA integrity have not been examined. In the present study, we analyzed this relationship among three endangered ungulates: Gazella cuvieri, Gazella dama mhorr, and Gazella dorcas neglecta. In addition, we examined whether levels of sperm DNA fragmentation are associated with semen quality. The magnitude of sperm DNA damage in the two species with high levels of inbreeding (G. cuvieri and G. dama mhorr) was extremely high when compared to the species with low levels of inbreeding (G. dorcas neglecta) and to values previously reported for outbred populations. Levels of sperm DNA fragmentation significantly increased with inbreeding and age. Increased DNA damage in sperm was associated with increased sperm head abnormalities, lower percentage of sperm with an intact acrosome, and poor motility. Our findings suggest that the link between inbreeding and semen quality is mediated by the effects of inbreeding upon sperm DNA damage. The deleterious effects of inbreeding upon the paternal genome likely decrease male fertility and may cause genetic damage to future generations. Because inbreeding is common among endangered species, high levels of sperm DNA damage may have considerable impact upon the viability of their populations.

Paternal levels of DNA damage in spermatozoa and maternal parity influence offspring mortality in an endangered ungulate.

Understanding which factors influence offspring mortality rates is a major challenge since it influences population dynamics and may constrain the chances of recovery among endangered species. Most studies have focused on the effects of maternal and environmental factors, but little is known about paternal factors. Among most polygynous mammals, males only contribute the haploid genome to their offspring, but the possibility that sperm DNA integrity may influence offspring survival has not been explored. We examined several maternal, paternal and individual factors that may influence offspring survival in an endangered species (Gazella cuvieri). Levels of sperm DNA damage had the largest impact upon offspring mortality rates, followed by maternal parity. In addition, there was a significant interaction between these two variables, so that offspring born to primiparous mothers were more likely to die if their father had high levels of sperm DNA damage, but this was not the case among multiparous mothers. Thus, multiparous mothers seem to protect their offspring from the deleterious effects of sperm DNA damage. Since levels of sperm DNA damage seem to be higher among endangered species, more attention should be paid to the impact of this largely ignored factor among the viability of endangered species.

Boar fertility and sperm chromatin structure status: a retrospective report.

Little information exists about boar sperm chromatin quality and fertility within a commercial setting. The objective of this report is to provide information about boar sperm chromatin integrity and its relationship to fertility. The sperm chromatin structure assay (SCSA) was used retrospectively to characterize sperm from 18 sexually mature boars having fertility information. Boar fertility was defined by farrow rate (FR) and average total number of pigs born (ANB) per litter of gilts and sows mated to individual boars. Fertility data was compiled for 1867 matings across the 18 boars. The SCSA uses flow cytometry to evaluate the structural integrity of sperm nuclear DNA. The SCSA parameters measured in this retrospective analysis were the percentage DNA fragmentation index (%DFI) and standard deviation of the DNA fragmentation index (SD DFI). The %DFI and SD DFI showed the following significant negative correlations with FR and ANB; %DFI vs FR, r = -0.55, P < .01; SD DFI vs FR, r = -0.67, P < .002; %DFI vs ANB, r = -0.54, P < .01; and SD DFI vs ANB, r = -0.54, P < .02. Although more information is required to better understand the relationship between DFI and boar fertility, this report suggests that the SCSA assay may be an important assay for identification of boars having potential for lowered fertility.

Data analysis of two in vivo fertility studies using Sperm Chromatin Structure Assay-derived DNA fragmentation index vs. pregnancy outcome.

The 2006 American Society for Reproductive Medicine Compendium of Practice Report found no significant effects of elevated sperm DNA fragmentation by using a 30% DNA fragmentation index (DFI) threshold for natural fertilization and SCSA data (odds ratio, 1.07; 95% confidence interval, 0.39-2.93). In contrast, it is shown in this article that these two in vivo studies showed significant odds ratios of 6.54 (95% confidence interval, 1.71, 24.91) and 7.58 (95% confidence interval, 2.54, 22.67), which resulted in the conclusion that the pregnancy (PG) rates are statistically significantly higher for the group with DFI below the thresholds of 30% and 40%, respectively. In addition, all the chi(2) statistics used to test the null hypotheses of no association between the pregnancy status and DFI for natural (normal) fertilization confirmed this conclusion that the probability of pregnancy for the group with <30% or <40% DFI was statistically significantly higher than that for the group with >30% or >40% DFI.

Significant decrease in sperm deoxyribonucleic acid fragmentation after varicocelectomy.

OBJECTIVE: To measure sperm DNA integrity values before and after varicocelectomy in patients with elevated preoperative levels of sperm DNA fragmentation. DESIGN: Retrospective. SETTING: Private urology clinic. PATIENT(S): Eleven patients with grade 1, 2, or 3 varicocele. INTERVENTION(S): Varicocelectomy. MAIN OUTCOME MEASURE(S): Sperm DNA fragmentation values were assessed before and after varicocelectomy. RESULTS(S): Ninety percent of the patients showed a significant decrease in sperm DNA fragmentation levels. CONCLUSIONS(S): Although this study was small, 10 of the 11 patients with varicocele showed a significant decrease in sperm DNA fragmentation after varicocele repair. Elevated sperm DNA fragmentation has been shown to have a significant negative effect on pregnancy outcome with use of in vivo, IUI, routine IVF, and to a lesser extent intracytoplasmic sperm injection fertilization; therefore pregnancy outcome may improve after varicocelectomy.

Effect of vaginal lubricants on sperm motility and chromatin integrity: a prospective comparative study.

OBJECTIVE: To evaluate the effect of vaginal lubricants Pre-Seed, FemGlide, Astroglide, and Replens on human sperm motility and chromatin integrity. DESIGN: Prospective, comparative, in vitro study. SETTING: Andrology laboratory at tertiary care hospital. PATIENT(S): Thirteen normozoospermic donors. INTERVENTION(S): Semen samples from 13 subjects were incubated in human tubal fluid media (HTF) controls and 10% (vol/vol) of Pre-Seed, FemGlide, Astroglide, and Replens lubricants. After 30 minutes, progressive sperm motility was assessed by light microscopy. Semen samples of 12 patients were placed in positive control (HTF), negative control (10% K-Y Jelly lubricant), and 10% vol/vol Pre-Seed and FemGlide lubricants. After 4 hours culture, spermatozoa were analyzed for percent DNA fragmentation index with use of the acridine orange-based sperm chromatin structure assay. MAIN OUTCOME MEASURE(S): Sperm motility and percent DNA fragmentation index. RESULTS: Percent motility did not differ significantly between HTF controls and Pre-Seed, whereas FemGlide, Replens, and Astroglide lubricants demonstrated a significant decrease in motility. There was no significant difference in percent DNA fragmentation index between the HTF controls and Pre-Seed, but a significant decline in sperm chromatin quality occurred with FemGlide and K-Y Jelly. CONCLUSION: Pre-Seed does not cause a significant decrease in progressive sperm motility or chromatin integrity in contrast to other lubricants used by couples.