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Authors are the souls of any journal, and deserve much respect. To publish a journal manuscripts are needed from authors. Authors have a great responsibility for producing facts of their work in terms of number and results truthfully and an individual honesty is expected from authors in this regards. Both ways its true "No authors-No manuscripts-No journals" and "No journals–No manuscripts–No authors". Reviewing a manuscript is also a very responsible and important task of any peer-reviewed journal and to be taken seriously. It needs knowledge on the subject, sincerity, honesty and determination. Although the process of reviewing a manuscript is a time consuming task butit is expected to give one's best remarks within the time frame of the journal.
Salient features of the JCDR: It is a biomedical, multidisciplinary (including all medical and dental specialities), e-journal, with wide scope and extensive author support. At the same time, a free text of manuscript is available in HTML and PDF format. There is fast growing authorship and readership with JCDR as this can be judged by the number of articles published in it i e; in Feb 2007 of its first issue, it contained 5 articles only, and now in its recent volume published in April 2011, it contained 67 manuscripts. This e-journal is fulfilling the commitments and objectives sincerely, (as stated by Editor-in-chief in his preface to first edition) i e; to encourage physicians through the internet, especially from the developing countries who witness a spectrum of disease and acquire a wealth of knowledge to publish their experiences to benefit the medical community in patients care. I also feel that many of us have work of substance, newer ideas, adequate clinical materials but poor in medical writing and hesitation to submit the work and need help. JCDR provides authors help in this regards.
Timely publication of journal: Publication of manuscripts and bringing out the issue in time is one of the positive aspects of JCDR and is possible with strong support team in terms of peer reviewers, proof reading, language check, computer operators, etc. This is one of the great reasons for authors to submit their work with JCDR. Another best part of JCDR is "Online first Publications" facilities available for the authors. This facility not only provides the prompt publications of the manuscripts but at the same time also early availability of the manuscripts for the readers.
Indexation and online availability: Indexation transforms the journal in some sense from its local ownership to the worldwide professional community and to the public.JCDR is indexed with Embase & EMbiology, Google Scholar, Index Copernicus, Chemical Abstracts Service, Journal seek Database, Indian Science Abstracts, to name few of them. Manuscriptspublished in JCDR are available on major search engines ie; google, yahoo, msn.
In the era of fast growing newer technologies, and in computer and internet friendly environment the manuscripts preparation, submission, review, revision, etc and all can be done and checked with a click from all corer of the world, at any time. Of course there is always a scope for improvement in every field and none is perfect. To progress, one needs to identify the areas of one's weakness and to strengthen them.
It is well said that "happy beginning is half done" and it fits perfectly with JCDR. It has grown considerably and I feel it has already grown up from its infancy to adolescence, achieving the status of standard online e-journal form Indian continent since its inception in Feb 2007. This had been made possible due to the efforts and the hard work put in it. The way the JCDR is improving with every new volume, with good quality original manuscripts, makes it a quality journal for readers. I must thank and congratulate Dr Hemant Jain, Editor-in-Chief JCDR and his team for their sincere efforts, dedication, and determination for making JCDR a fast growing journal.
Every one of us: authors, reviewers, editors, and publisher are responsible for enhancing the stature of the journal. I wish for a great success for JCDR."
Thanking you
With sincere regards
Dr. Rajendra Kumar Ghritlaharey, M.S., M. Ch., FAIS
Associate Professor,
Department of Paediatric Surgery, Gandhi Medical College & Associated
Kamla Nehru & Hamidia Hospitals Bhopal, Madhya Pradesh 462 001 (India)
E-mail: drrajendrak1@rediffmail.com
On May 11,2011
Dr. Shankar P.R.
"On looking back through my Gmail archives after being requested by the journal to write a short editorial about my experiences of publishing with the Journal of Clinical and Diagnostic Research (JCDR), I came across an e-mail from Dr. Hemant Jain, Editor, in March 2007, which introduced the new electronic journal. The main features of the journal which were outlined in the e-mail were extensive author support, cash rewards, the peer review process, and other salient features of the journal.
Over a span of over four years, we (I and my colleagues) have published around 25 articles in the journal. In this editorial, I plan to briefly discuss my experiences of publishing with JCDR and the strengths of the journal and to finally address the areas for improvement.
My experiences of publishing with JCDR: Overall, my experiences of publishing withJCDR have been positive. The best point about the journal is that it responds to queries from the author. This may seem to be simple and not too much to ask for, but unfortunately, many journals in the subcontinent and from many developing countries do not respond or they respond with a long delay to the queries from the authors 1. The reasons could be many, including lack of optimal secretarial and other support. Another problem with many journals is the slowness of the review process. Editorial processing and peer review can take anywhere between a year to two years with some journals. Also, some journals do not keep the contributors informed about the progress of the review process. Due to the long review process, the articles can lose their relevance and topicality. A major benefit with JCDR is the timeliness and promptness of its response. In Dr Jain's e-mail which was sent to me in 2007, before the introduction of the Pre-publishing system, he had stated that he had received my submission and that he would get back to me within seven days and he did!
Most of the manuscripts are published within 3 to 4 months of their submission if they are found to be suitable after the review process. JCDR is published bimonthly and the accepted articles were usually published in the next issue. Recently, due to the increased volume of the submissions, the review process has become slower and it ?? Section can take from 4 to 6 months for the articles to be reviewed. The journal has an extensive author support system and it has recently introduced a paid expedited review process. The journal also mentions the average time for processing the manuscript under different submission systems - regular submission and expedited review.
Strengths of the journal: The journal has an online first facility in which the accepted manuscripts may be published on the website before being included in a regular issue of the journal. This cuts down the time between their acceptance and the publication. The journal is indexed in many databases, though not in PubMed. The editorial board should now take steps to index the journal in PubMed. The journal has a system of notifying readers through e-mail when a new issue is released. Also, the articles are available in both the HTML and the PDF formats. I especially like the new and colorful page format of the journal. Also, the access statistics of the articles are available. The prepublication and the manuscript tracking system are also helpful for the authors.
Areas for improvement: In certain cases, I felt that the peer review process of the manuscripts was not up to international standards and that it should be strengthened. Also, the number of manuscripts in an issue is high and it may be difficult for readers to go through all of them. The journal can consider tightening of the peer review process and increasing the quality standards for the acceptance of the manuscripts. I faced occasional problems with the online manuscript submission (Pre-publishing) system, which have to be addressed.
Overall, the publishing process with JCDR has been smooth, quick and relatively hassle free and I can recommend other authors to consider the journal as an outlet for their work."
Dr. P. Ravi Shankar
KIST Medical College, P.O. Box 14142, Kathmandu, Nepal.
E-mail: ravi.dr.shankar@gmail.com
On April 2011 Anuradha
Dear team JCDR, I would like to thank you for the very professional and polite service provided by everyone at JCDR. While i have been in the field of writing and editing for sometime, this has been my first attempt in publishing a scientific paper.Thank you for hand-holding me through the process.
Dr. Anuradha
E-mail: anuradha2nittur@gmail.com
On Jan 2020
Original article / research
Full Version
Role of Sperm DNA Fragmentation Index in Semen Analysis in Couples with Unexplained Recurrent Pregnancy Loss: A Case-control Study
Correspondence Address :
Dr. Ruchi Hooda,
Senior Resident, Department of Reproductive Medicine, CIMAR, Edappal Hospital, Edappal-679576, Kerala, India.
E-mail: hoodaruchi@gmail.com
Introduction: Infertility has been on the rise, and male factor infertility has gained attention in cases of Recurrent Pregnancy Loss (RPL). Traditional Semen Analysis (SA) has been the main diagnostic tool, but with advancements in Assisted Reproductive Technology (ART) and the need for more accurate testing, there is a demand for improved diagnostic tests that correlate with reproductive outcomes. Sperm DNA Fragmentation Index (DFI) is a parameter used to assess the degree of sperm DNA damage and is considered crucial in evaluating semen quality.
Aim: To investigate the role of Sperm DNA Fragmentation (SDF) in patients presenting with RPL compared to a control group.
Materials and Methods: The study was conducted over a two-year period from January 2021 to December 2022 at the Centre for Infertility Management and Assisted Reproduction (CIMAR), Edappal Hospital, Edappal, Kerala, India. The control group (Group A; n=31) comprised males aged between 21-45 years whose partners had no history of recurrent abortions. The case group (Group B; n=31) included males with two or more pregnancy losses. Sperm DFI analysis and routine SA were performed in both groups to assess semen parameters such as volume, concentration, progressive and non progessive motility, morphology, and DFI. Statistical analysis was performed using the Independent student t-test.
Results: In the present study the mean age of males in Group A was 37.03±5.416 years and in Group B was 35.44±4.552 years. There were no significant differences observed between the case and control groups in terms of sperm volume (p=0.301), concentration (p=0.155), progressive motility (p=0.207), non progessive motility (p=0.178), and morphology (p=0.362). However, a statistically significant difference was found between the RPL and control groups for DFI (p<0.001), with a mean value of ±8.15 in the control group and ±19.35 in the case group.
Conclusion: The present study demonstrates that SDF is an important factor in RPL, with couples experiencing RPL showing a higher incidence of SDF. Therefore, incorporating SDF analysis alongside routine SA should be considered, particularly in patients with a history of RPL.
Assisted reproductive treatment, Deoxyribonuleic acid, Sperm chromatin dispersion
The definition of RPL is not uniform. The American Society of Reproductive Medicine defines RPL as two or more pregnancy failures (1). The Royal College of Obstetricians and Gynaecologists defines RPL as three or more consecutive losses with the same sexual partner, occurring before 24 weeks of pregnancy (2). The European Society of Human Reproduction and Embryology (ESHRE) guideline group defines RPL as the occurrence of three or more consecutive pregnancy losses before 20 weeks of gestation (3). The aetiology of RPL is unknown in about 50% of cases (4). The causes of RPL are heterogeneous and include endocrine dysfunction, autoimmune disorders, maternal and paternal age, genetic abnormalities, infectious diseases, congenital or structural uterine anomalies, and exposure to environmental toxins (4).
Infertility is on the rise, with an incidence of 10-15% in couples of reproductive age (5). Among these cases, 50% involve male factors, and 20% exclusively have “male factor” infertility (6). Recently, the evaluation of male factors has also gained importance in cases of RPL. The diagnosis of male factor infertility is heavily reliant on traditional SA. However, traditional SA is found to be normal in only about 15% of male patients experiencing infertility (7). This highlights the limitation of SA in making a diagnosis of male fertility, as it merely serves as a surrogate marker and provides limited information on sperm concentration, motility, and morphology, without revealing sperm functional competence and reproductive potential.
With advancements in ART and the increasing needs of at-risk couples, traditional SA alone has become insufficient. There is a demand for more advanced diagnostic tests that can correlate with reproductive outcomes. Sperm DFI is used to assess DNA damage, which is considered an important parameter in evaluating semen quality. It has been observed that sperm DNA fragmentation can affect fertilisation, embryonic development, and the transmission of paternal genetic information during both spontaneous and ART pregnancies [1,8].
The aetiology of sperm DNA damage can include abnormal chromatin packing, apoptosis, and elevated levels of Reactive Oxygen Species (ROS) (9). Several tests are available to assess sperm DNA integrity by evaluating strand breaks In-situ (10). Methods such as the single-cell gel electrophoresis assay (comet assay), Terminal Deoxynucleotidyl Transferase d-UTP Nick End Labelling (TUNEL) assay, and Sperm Chromatin Structure Assay (SCSA) are commonly utilised (11). Although, sperm DFI is not a part of the basic semen analysis, it is useful in cases of unexplained infertility with normal semen parameters. The evaluation of the male factor has been given less importance in couples with RPL, and a study was conducted to assess the role of the male factor and the role of DFI in semen analysis in association with RPL. The study reinforces that sperm DNA fragmentation testing could be used to provide explanations in couples with unexplained RPL (12). There is evidence supporting an independent association between RPL and sperm DNA fragmentation, regardless of female factors, according to the ESHRE guidelines. However, the guidelines highlight the need for further investigation to understand the impact of sperm DNA fragmentation on RPL (13).
Therefore, studies are needed to examine other aspects such as sperm DNA maturity and condition, as well as focusing on molecular factors implicated in male fertility, such as oxidative stress molecules, sperm DNA fragmentation, and Sperm Chromatin Density (SCD) (14). The aim of the present study was to investigate the role of sperm DNA fragmentation in patients presenting with RPL compared to a control group.
The present case-control study was conducted over a period of two years, from January 2021 to December 2022, at CIMAR, Edappal Hospital, Edappal, Kerala, India. The study was approved by the Research Ethics Board of Edappal Hospital Pvt., Ltd., Edappal, Kerala, India (IEC-13/19). The control group (Group A; n=31) consisted of males aged between 21-45 years whose partners had no history of recurrent abortions. The case group (Group B; n=31) included males who had two or more pregnancy losses. Sperm DFI analysis and routine SA were performed in both groups to assess semen parameters such as volume, concentration, progressive and non progessive motility, morphology, and DFI of the sperm.
Sample size calculation: Sample size was calculated based on a prevalence (P) of unexplained RPL of 1%, a significance level of 5%, a power (1-β) of 90%, and an effect size of 0.5, with a 10% allowance for loss to follow-up, using G power 3.1 software.
Inclusion criteria: males aged between 21-45 years whose partners had two or more pregnancy losses and in the control group, couples with unexplained infertility were included.
Exclusion criteria: Male partners with female partners having uterine anomalies, known antiphospholipid syndrome, inherited thrombophilias, endocrine causes, or balanced translocations were excluded from the cases. Patients with any known cause for male and female infertility leading to recurrent losses were excluded from the study.
Study Procedure
Basal and demographic data of the recruited patients were collected, including the age and BMI of the partners. Male partners underwent SA and DFI testing after an abstinence period of 2-7 days. Morphological abnormalities were assessed using Papanicolaou stain and categorised based on the location of the defect involving the head, neck, or tail. The Sperm Chromatin Dispersion (SCD) test was used to assess the DFI. After assessing the concentration, the semen sample was diluted with a culture medium to a concentration of 5-10 million/mL. Agarose was melted at 90o Celsius by placing it in the sperm chroma warmer 1 for five minutes. The melted agarose was then transferred to the sperm chroma warmer 2, which was maintained at 37o Celsius, and allowed to equilibrate for five minutes. Approximately 25 microliters of the diluted semen sample was added to the agarose and mixed thoroughly. The sperm cell suspension was immediately placed onto pretreated slides, and a cover slip was carefully placed, taking precautions to avoid the formation of air bubbles. The slide was then kept at 4o Celsius for five minutes, and the cover slip was gently removed by sliding it off. Throughout the procedure, the slide was maintained in a horizontal position.
The Sperm Chroma Kit from Cryolab International, which included four solutions (A, B, C, and D), was used following the protocol provided by the company. The slides were incubated horizontally in solution A for seven minutes, followed by lysis solution for 25 minutes. Afterward, the slides were immersed in distilled water for five minutes, 70% ethanol for two minutes, 90% ethanol for two minutes, and finally, 100% ethanol for two minutes. The slides were allowed to dry at room temperature. A mixture of solutions C and D in a 1:1 ratio was prepared and a layer of the stain was deposited 8horizontally on the slides, which were then left for 15-20 minutes. The stain was decanted, and the slides were gently washed with distilled water and dried at room temperature. The slides were visualised under a bright field microscope using a 20X or 40X objective. At least 300-500 sperm were counted to calculate the DFI. Sperm without DNA fragmentation exhibited a large 2 halo or medium halo, while sperm with DNA fragmentation appeared small, without a halo, or degraded.
Statistical Analysis
Statistical analysis was performed using the Statistical Package for the Social Sciences (SPSS) software, version 20.0. Continuous variables were presented as mean value±Standard Deviation (SD), and comparisons between groups were determined using Independent t-tests. The Shapiro-Wilk test or Kolmogorov-Smirnov test was used to assess normality. All tests were two-sided, and a p-value <0.05 was considered statistically significant. Kurtosis was used to describe the degree to which scores cluster in the tails or the peak of a frequency distribution.
The study included 31 women in the age range of 21-45 years (mean: 32.1±5.51 years) who had experienced RPL, and 31 women (mean: 29.74±4.69 years) as controls. There were no statistically significant differences in the ages of the women (p=0.078) (Table/Fig 1).
A statistically significant difference was observed between the RPL group and the control group only for the DFI (Table/Fig 2).
Sperm DNA integrity plays a crucial role in determining male reproductive potential and outcomes. The inclusion of sperm DNA Fragmentation (SDF) testing can be used for predictive purposes and to plan individualised treatment strategies. Unfortunately, many ART centres still overlook the evaluation of male partners facing fertility issues with normal SA or who have spermatozoa available for intracytoplasmic sperm injection without considering the potential impact of DNA fragmentation on RPL. This oversight can lead to multiple failed ART cycles, adding to the emotional distress experienced by the couple and the challenges faced by the treating physician. SDF testing in men with Oligoasthenoteratozoospermia (OAT) on routine SA could be considered, as recommended by the European Academy of Andrology (EAA) (15). The European Association of Urology (EAU) supports SDF testing in couples with unexplained infertility or after RPL (16). The European Society of Human Reproduction and Embryology (ESHRE) recognises the potential role of SDF testing in explaining the association with RPL (17). Recently, the American Urological Association (AUA) and American Society for Reproductive Medicine (ASRM) published guidelines on male infertility, recommending against SDF testing in the initial evaluation of fertility but acknowledging its importance in couples experiencing RPL (18).
In the present study, sperm from men in the RPL group exhibited a higher percentage of DNA fragmentation (31.22%) compared to sperm from men in the control group (18.5%) p<0.001. These findings are consistent with other studies that have reported significantly higher levels of abnormal DNA fragmentation in the recurrent spontaneous abortion group compared to the control group [19-21]. This further supports the association between increased DNA fragmentation and RPL (p<0.001). Leach et al., conducted a study involving 108 couples with a history of RPL and found significantly higher levels of SDF using the Sperm Chromatin Structure Assay (SCSA) (22). Similarly, Kamkar N et al., used a similar method to assess DNA fragmentation and found higher DFI in the RPL group compared to the control group (23). Regarding other semen parameters, the present study did not find any significant differences between the two groups in relation to an increased risk of RPL.
A meta-analysis conducted by Yifu P et al., included seven studies using the SCSA, nine studies using the SCD test, and eight studies using TUNEL assay. The results showed significant differences supporting an association between sperm DFI and RPL (24). Another meta-analysis that assessed the relationship among traditional semen parameters, SDF, and unexplained recurrent miscarriage included a total of 1182 couples with unexplained recurrent miscarriage and 1231 couples without recurrent miscarriage. The results showed significantly increased levels of SDF in unexplained recurrent miscarriage and significantly decreased levels of total motility and progressive motility compared to couples without recurrent miscarriage (25).
In contrast, De Geyter et al., suggested that sperm selection techniques used in ART to decrease SDF rates do not necessarily lead to improvements in pregnancy rates (26). Moreover, van den Berg et al., confirmed that even a few double-strand DNA breaks are sufficient to delay cell cycle progression (27). Another study by Gil-Villa AM et al., reported no significant correlation between DNA fragmentation and RPL using the SCSA and concluded that DFI was not an important cause or predictive factor for RPL (28). This may be due to the fact that oxidant agents have an impact on sperm morphology, motility, and concentration, but not on the sperm nucleus, which provides genetic material to the future embryo. Under these conditions, sperm can still fertilise the oocyte.
The correlation between DFI and RPL remains highly controversial. These differences in opinion among studies may be due to the limited number of cases with RPL and the use of different evaluation methods with varying sensitivities and specificities. The most commonly used methods are SCSA, TUNEL, and SCD (29). Although authors concluded, that routine semen analysis cannot solely be relied upon to assess the role of the male factor in the incidence of idiopathic recurrent early pregnancy loss, further additional sperm tests, such as SDF testing, are needed to search for a better diagnostic tool.
Limitation(s)
The small sample size was a limitation of present study. Additionally, the study was conducted in a single centre, so the findings cannot be generalised.
Sperm DFI is recognised as an important cause of RPL, and couples with a history of RPL have been shown to have a higher incidence of SDF. It has been observed that higher levels of sperm DFI are associated with a higher risk of RPL. Therefore, SDF testing has potential predictive value in diagnosing DNA fragmentation and assessing couples with RPL. It is necessary to perform an SDF test in couples facing RPL.
Authors’ contributions: RH contributed to the conceptual design of the article, as well as the acquisition and analysis of the data. GDG made a substantial contribution to the acquisition, analysis, and interpretation of data for the article.
The authors are thankful for the efforts received from the patients and hospital staff.
DOI: 10.7860/JCDR/2023/63044.18430
Date of Submission: Jan 22, 2023
Date of Peer Review: Apr 05, 2023
Date of Acceptance: Jun 03, 2023
Date of Publishing: Sep 01, 2023
AUTHOR DECLARATION:
• Financial or Other Competing Interests: None
• Was Ethics Committee Approval obtained for this study? Yes
• Was informed consent obtained from the subjects involved in the study? Yes
• For any images presented appropriate consent has been obtained from the subjects. Yes
PLAGIARISM CHECKING METHODS:
• Plagiarism X-checker: Feb 08, 2023
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ETYMOLOGY: Author Origin
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