Comparison of live birth rate and fetal outcomes between fresh embryo and frozen-thawed embryo transfers: a prospective study

Abstract

Background

The study aimed to compare the pregnancy, prenatal, and postnatal outcomes between fresh and frozen embryo transfer (ET) in intracytoplasmic sperm injection (ICSI) cycles at Al-Zahra Referral Women’s Hospital in northwest Iran.

Methods

A prospective study was conducted among all infertile women (N = 469) who underwent embryo transfer between 2018 and 23 at Al-Zahra referral infertility center. Patients in cycles with fresh embryo transfer and patients for whom frozen embryos were transferred were compared in terms of live birth rate and fetal outcomes. Multiple logistic regression analysis was used to estimate adjusted odds ratios (AORs) with 95% confidence intervals (CIs).

Results

The majority of the participants were primary infertility 83.3%. The rate of chemical and clinical pregnancy was (21.8% vs. 17.2%) and (19% vs. 13.4%) in the fresh embryo and the frozen embryo transfers, respectively. Likewise, the rate of live births was (14.1% vs. 9.1%), respectively. The number of retrieved oocytes was significantly higher in frozen ET compared to fresh ET (P = 0.001). In the final analysis, after adjusting for potential confounders, no significant associations were found for clinical pregnancy (AOR = 1.51; 95% CI: 0.90–2.5; P = 0.125) and chemical pregnancy (AOR = 1.31; 95% CI: 0.81–2.3; P = 0.238) rates between fresh and frozen ETs. Similarly, there were no significant differences in live birth rate (AOR = 1.6; 95% CI: 0.54–12.4), preterm birth (AOR = 0.62; 95% CI: 0.33–5.5), and primary infertility (AOR = 0.73; 95% CI: 0.34–1.6) between fresh and frozen ETs. The incidence of multiple pregnancies and spontaneous abortion was (5% vs. 13.8%) and (22.2% vs. 30.2%) in the fresh embryo and frozen embryo groups, respectively.

Conclusion

No significant differences in perinatal and postnatal outcomes were found between fresh and frozen embryo transfers.

Introduction

Infertility is a disease characterized by the failure to establish a clinical pregnancy after 12 months of regular and unprotected sexual intercourse [1, 2]. Although the prevalence of infertility varies among societies, its global prevalence is estimated at 10% [3]. Couple infertility can be linked to women in 20–35% of cases, men in 20–30% of cases, or both in 25–40% of cases [4, 5]. In this respect, assisted reproductive technology (ART) can be used, including standard techniques such as in-vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI). When the first successful pregnancy was achieved outside the human body, it was a considerable success that led to the use of ARTs worldwide. IVF treatment is a choice method for many infertility cases, but its efficacy is limited to female infertility and cannot apply to male infertility cases. In contrast, unlike the IVF method, ICSI has an up to 70% success rate for most male infertility cases [6].

During assisted reproductive methods, one of the basic steps is embryo transfer (ET), which is done in two ways, frozen and fresh. In fresh ET, the embryo transfer is performed often either three or five days after the retrieval. In frozen ET, the embryo has been previously created for many months and/or even years earlier and then will be placed into the uterus. With either transfer, the endometrium (the lining of the uterus) must be prepared to facilitate easier implantation of the embryo. A systematic review and meta-analysis indicated that a significant increase in live birth rates with frozen ET was observed solely in hyper-responders and in patients undergoing preimplantation genetic testing for aneuploidy. In terms of safety, frozen ET may reduce the risk of moderate and severe Ovarian Hyperstimulation Syndrome (OHSS), while potentially increasing the risk of preeclampsia [7]. Another review found that IVF outcomes may be improved by performing frozen ET compared with fresh ET [8]. While Wong et al. reported no significant differences between fresh and frozen ETs regarding pregnancy and live birth rates [9]. In cases of failure to conceive with the transfer of fresh embryos or the presence of contraindications for using this method, such as the occurrence of OHSS, frozen embryos are used for transfer. In this method, embryos that have been frozen outside the uterus after the fertilization cycle are first thawed and warmed, and then transferred to the uterus in a regular cycle or after endometrial hormonal preparation [8].

The increase of available embryos through freezing and pre-conception genetic tests, also the ability to synchronize embryo transfer with the endometrial cycle are among the other main reasons for the widespread use of frozen transfer [10–13]. Moreover, it can be said that the most important reasons for the development of freezing and frozen transfer methods is to reduce the risk of multiple pregnancies by transferring a limited number of new embryos because multiple pregnancies itself is one of the most critical risk factors for miscarriage and premature birth, so with this method, all new embryos are not forced to be transferred; so they can be used in the future because of freezing [14, 15].

However, conflicting evidence regarding the outcomes of frozen and fresh transfers requires further investigation. The results of recent studies of embryo transfer methods alone indicate the necessity of investigating their consequences [16]. For example, a prospective study demonstrated non-divergent growth trajectories in IVF/ICSI pregnancies after frozen-thawed or fresh blastocyst transfer from 19 weeks of gestation to birth, while there were a higher standardized estimated fetal weight and birth-weight Z-scores in the frozen-thawed in comparison with fresh blastocyst transfer cases [17]. Another study conducted by Cavoretto et al. showed that thawed blastocyst transfers after IVF/ICSI conceptions present greater fetal crown-rump length compared with fresh at 6–14 weeks, but both IVF/ICSI groups show smaller crown-rump length than the general population [18]. Limited evidence compares the wide range of pregnancy, prenatal, and postnatal outcomes of these two methods of fresh and frozen embryo transfers with a prospective design. Assisted reproduction methods aim to give birth to a healthy baby with the least risk for the mother, and this should be distinguished from trying to increase the pregnancy rate. ART leaves a lot of material and spiritual costs both for the couple and for the treatment system [19, 20]. Infertile couples can be fully informed before embarking on an emotionally difficult and costly process. Although the current techniques regarding the factors involved in frozen transfer have increased dramatically, conflicting results have been reported regarding different aspects of frozen transfer and its advantages and disadvantages compared to fresh transfer [21, 22]. In addition, different ART protocols in different centers are one of the reasons involved in the heterogeneity of results, which reminds us of the necessity of conducting studies in this field [23].

The infertility center at Al-Zahra Hospital serves as a referral center in northwestern Iran, where a significant number of ART procedures are conducted annually. The importance of evaluating the outcomes of both frozen and fresh embryo transfers at this center is twofold. Numerous female patients from neighboring provinces and across northwestern Iran seek health services from this hospital. However, there is no evidence of the efficacy and effectiveness of the above two ART in increasing fertility rates in this center. The results of this study can provide valid evidence for the best clinical decision-making, where many clinicians are actively trying to improve their patient’s probability of becoming fertile. This issue is of utmost importance for low-income settings forced to use expensive ART by couples. Therefore, this study aims to compare a wide range of fetal, pregnancy, prenatal, and postnatal outcomes between fresh and frozen embryo transfer in ICSI cycles at the infertility center of Al-Zahra Hospital in the Northwest of Iran.

Methods

Study design and participants

In this prospective study, 462 ICSI-ET cycles (142 fresh ET and 320 frozen ET) from 493 women undergoing ICSI treatment referred to Al-Zahra Infertility and Reproductive Health Research Center in Tabriz, Iran from 2018 to 2023 were analyzed. The study population was all women undergoing ICSI treatment referred to Al-Zahra Infertility Center. The study compared patients in cycles of fresh or frozen embryo transfer in the manner of live birth rate and fetal outcomes as the primary outcome. The study protocol with the code (IR.TBZMED.REC.1400.599) was approved by the Ethics Committee of Tabriz University of Medical Sciences, Tabriz, Iran.

Eligibility

Inclusion criteria were ICSI treatment using the long protocol (GnRH agonist), endometrial thickness more than 8 mm, having normal follicle-stimulating hormone (FSH) of the third day of the menstrual cycle, age under or equal to 42 years, absence of internal uterine anomalies (such as adenomyosis, uterine fibroids, uterine abnormalities, and clear endometrioma mass), and absence of systemic diseases. We excluded women having more than three cycles of ART, natural cycles, history of endocrine disorders (hypothyroidism and hyperthyroidism, diabetes, hyperprolactinemia), Asherman’s syndrome, oocyte donation, history of surgical removal of endometriosis, leiomyoma, uterine anomalies in hysterosalpingography (HSG), uterine septum, and hysteroscopy and gamete donation.

Outcome evaluation

Primary outcome was the live birth rate and fetal outcomes of fresh embryo transfer in comparison with frozen embryo transfer. Other comparisons included comparing the transfer of live and frozen embryos in a patient in two separate processes, as well as comparing the type of preparation of the uterus (in terms of medication) in the frozen embryo transfer process were also considered.

Concerning the ET timing, based on previous evidence progesterone intake started on the theoretical day of oocyte retrieval in hormone replacement therapy and to perform blastocyst transfer at hCG + 7 or LH + 6 in the modified or true natural cycle, respectively [24]. Embryos were transferred on Day 5 of culture. The embryo(s) that had the best morphology were transferred first. On Day 6 of culture, all embryos that were surplus were cryopreserved. Women with a fresh transfer had luteal support with 600 mg vaginal micronized utrogestan and continued this until the pregnancy test, 14 days after the fresh transfer. If not pregnant, frozen embryo transfer was scheduled in artificial cycles with oral estrogen and vaginal micronized progesterone supplementation. Women were given vaginal micronized progesterone and scheduled thawing and transfer if their endometrium reached 8 mm on vaginal ultrasound. Until 11 + 5 weeks gestation, women continued taking estrogen and progesterone supplements. If not pregnant, a subsequent artificial cycle was started [25].

According to the routine technique of assisted pregnancy, a serum beta-hCG titration test was performed 14 days after embryo transfer. Validation of successful implantation was done by detecting an increased β-HCG concentration (> 50 U/ml) 16 days post embryo transfer and was defined as positive biochemical pregnancy. About four weeks after embryo transfer, a gynecologist performs a transvaginal ultrasound, and if a gestational sac is observed, it is considered a clinical pregnancy. First, the rate of abortion less than 20 weeks in the embryo transfer groups was calculated separately, and then this rate was compared in the two groups. Live birth after 25 weeks was obtained from the records of the patients in two groups, and if it was not in the file, it was acquired by telephone call.

Secondary outcomes include chemical, clinical, and ectopic pregnancy rates, abortion rates, and other obstetric outcomes (such as preterm labor, preeclampsia, placenta accreta, etc.) were recorded according to oocyte owner age group and fetus owner age separately.

Chemical confirmation of pregnancy was defined as a serum beta-hCG level of more than 50 IU/L after fourteen days of embryo transfer. Clinical pregnancy was defined as observing fetal heart activity by transvaginal ultrasound five weeks after a positive beta-hCG result. After recording all the patient’s information, the incidence of each outcome was calculated to compare the consequence of two types of transmission on pregnancy outcomes. Then they were compared with the results of the occurrence of the outcome in the other group. Baseline, clinical, and paraclinical information, along with pregnancy and postnatal outcomes, were gathered from the medical records at Al-Zahra Infertility Center. For incomplete data, particularly regarding some postnatal outcomes, follow-up was conducted via telephone.

Statistical analysis

Statistical analysis was carried out using the SPSS (Statistical Package for the Social Sciences, version 21.0, SPSS Inc, Chicago, Illinois, USA). The results were demonstrated as mean ± standard deviation (SD) and frequency (percentage). First, the data normality was checked by the Kolmogorov-Smirnov test. Quantitative variables were compared by Independent T-test when normal distribution and homogeneity of variance were met and/or Mann-Whitey U was carried out for non-parametric variables. Chi-Square test was used for categorical variables when the value of the cell expected were 5 or more in at least 80% of the cells, and no cell had an expected of less than one, otherwise, Fisher’s Exact test was used.

Multiple logistic regression was used to estimate adjusted odds ratios (AORs) with 95% confidence intervals (CIs) and adjusted for the potential confounders. Variables with a p-value less than 0.25 included in multiple logistic regression analysis using Enter method. The model goodness of fit was assessed Hosmer and Lemeshow statistic [26].

Results

We included 493 candidates for ICSI treatment at the infertility center of Al-Zahra Hospital in Tabriz. Of these, 24 patients were excluded due to a lack of follow-up and missing clinical records information, and finally, 469 patients were evaluated. A total of 142 patients received fresh ET and 320 patients were received frozen ET during the study period. The mean age of the patients was 31.43 ± 4.49 years (ranging from 16 to 42 years), while the average age of the husband was 35.77 ± 6.30 years (ranging from 21 to61 years). The mean body mass index (BMI) was 26.57 ± 4.14 kg/m² (17–43 kg/m²). The mean duration of the infertility period was 20.6 ± 4.61 years (1–25 years). Three hundred ninety-one patients (83.4%) had primary infertility, and 78 (16.6%) had secondary infertility. The mean puncture attempts and transfusion cycles were 1.26 ± 0.61 and 1.82 ± 1.05, respectively. The live birth rate of embryo transfers was 10.6% (49/462). Also, in 5 patients, twins were born.

Table 1 shows the baseline characteristics of the participants based on fresh and frozen-thawed embryo transfer groups. No significant differences were found regarding the age of the women and their husbands, infertility period and types (primary and secondary), cycle puncture attempts, history of ART, and BMI (P > 0.05). However, there was a significant difference in the average number of transfer attempts (P = 0.009).

Table 1.

Baseline characteristics of the participants by the fresh and frozen-thawed embryo transfers groups

Variable		Fresh ET (n = 142)	Frozen ET (n = 320)	p-value
Age (years)		31.10 ± 6.23	31.78 ± 6.52	0.364
Husband age (years)		35.61 ± 6.20	35.79 ± 6.32	0.843
BMI (kg/m 2 )		26.67 ± 4.53	26.56 ± 4.10	0.882
Infertility period (years)		6.40 ± 4.99	6.18 ± 4.57	0.779
Infertility type	Primary	40 (81.6%)	351 (83.6%)	0.430
	Secondary	9 (18.4%)	69 (16.4%)
Cycle puncture attempts		1.21 ± 0.68	1.27 ± 0.60	0.563
Transfers attempts		1.48 ± 0.88	1.86 ± 1.06	0.009
History of ARTs		2 (4.08%)	4 (0.95%)	0.180

BMI: Body mass index; ART: Assistant reproductive treatment.

The comparison of ICSI cycle outcomes based on the embryo transfer method (fresh embryos and frozen embryos) is shown in Table 2. Out of 469 patients, 142 (30.3%) were fresh embryos transferred, and 320 (68.2%) were frozen embryos transferred, and in contrast, in 7 (1.5%) cycles, embryos were not transferred. The rate of chemical and clinical pregnancy was (21.8% vs. 17.2%; OR = 1.34 (0.82–2.20)) and (19% vs. 13.4%; OR = 1.51 (1.0-2.56)) in the fresh embryo and the frozen embryo cases, respectively. Likewise, the rate of live births was (14.1% vs. 9.1%), respectively. There were four cases of unsuccessful transfer, one case of a uterine anomaly, and two cases of no egg formation. One clinical abortion was performed due to a Down’s syndrome diagnosis before 20 weeks of gestation. The chemical and clinical pregnancy rates were 18.6% (86/462) and 15.2% (70/462), respectively. The Apgar score and birth weight were (8.4 vs. 8.5) and (2.9 vs. 3.02 kg) in fresh and frozen ETs, respectively, with no significant differences between the groups (P > 0.05). Likewise, the distribution of preeclampsia was comparable in both transfer methods (P > 0.05).

Table 2.

Comparison of intracytoplasmic sperm injection cycle outcomes based on the study groups

Variable	Fresh ET (n = 142)	Frozen ET (n = 320)	OR (95% CI)	p-value
Chemical pregnancy rate	31 (21.8%)	55 (17.2%)	1.34 (0.82–2.20)	0.146
Clinical pregnancy rate	27 (19%)	43 (13.4%)	1.51 (1.0-2.56)	0.048
Spontaneous abortion rate	6 (22.2%)	13 (30.2%)	1.04 (0.38–2.79)	0.327
Clinical abortion rate	1	1	1.64 (0.89–3.02)	0.999
Live birth rate	20 (14.1%)	29 (9.1%)	1.43 (96.0-2.46)	0.075
Preterm labor rate	3 (15%)	6 (20.7%)	1.12 (0.27–4.58)	0.455
Twin birth rate	1 (5%)	4 (13.8%)	0.56 (0.06–5.06)	0.311
Preeclampsia	4 (2.8%)	6 (1.9%)	1.1 (0.84–2.4)	0.385
Retrieved oocyte number	6.64 ± 4.24	13.35 ± 7.80	2.7 (1.75–6.57)	0.001
Apgar score	8.4 ± 2.7	8.5 ± 2.5	0.97 (0.67–1.23)	0.466
Birth weight (kg)	2.9 ± 1.7	3.02 ± 1.6	0.95 (0.59–1.5)	0.742

OR: Odds ratio; CI: Confidence interval. 42

Table 3 indicates the prenatal and postnatal outcomes between the frozen and fresh embryo transfers. The frequency of spontaneous abortion in the first trimester was 27.1% (19/70). There is no significant difference in comparing prenatal and postnatal outcomes between the frozen and fresh embryo transfers. Details of the prenatal and postnatal outcomes are presented in Table 3.

Table 3.

Comparison of prenatal and postnatal outcomes between the frozen and fresh embryo transfers

Variables		Fresh ET (n = 142)	Frozen ET (n = 320)	p-value
Amniocentesis		2 (8.7%)	5 (15.2%)	0.387
Normal		2	4
Abnormal		0	1
Birth age (week)		37.66 ± 2.02	37.34 ± 2.63	0.641
Delivery	NVD	1 (5%)	0 (0%)	0.408
	C/S	19 (95%)	29 (100%)
C/S etiology	Patient’s desire	10 (50%)	9 (31.1%)	0.125
	Complication	10 (50%)	20 (68.9%)
Birth weight (gram)		3196.25 ± 605.33	2967.14 ± 596.60	0.234
Congenital abnormality		0	0	NA
PPROM		2 (10%)	4 (13.8%)	0.527
IUGR		1 (5%)	2 (6.9%)	0.639
Gestational HTN		1 (5%)	1 (3.4%)	0.655
GDM		2 (10%)	2 (6.9%)	0.534

NVD: Natural vaginal delivery; C/S: Cesarean section; PPROM: Preterm premature rupture of the membranes; IUGR: Intrauterine growth restriction; HTN: Hypertension; GDM: Gestational diabetes mellitus, ET: embryo transfer

Table 4 presents the results of a multiple logistic regression analysis estimating the crude and adjusted AORs for prenatal and postnatal outcomes of fresh embryo transfer compared to frozen embryo transfer. After adjusting for potential confounders such as age, body mass index (BMI), history of ART, number of retrieved oocytes, and cesarean section; fresh transfer slightly increased the odds of clinical pregnancy by 1.5 times compared to frozen transfer, although this difference was not statistically significant (AOR = 1.51; 95% CI: 0.91–2.5; P = 0.125). Similarly, the chemical pregnancy rate for fresh transfer was higher than that for frozen transfer, but this association was also not significant (AOR = 1.31; 95% CI: 0.80–2.3; P = 0.238). The odds of live birth were 1.6 times greater in fresh embryo transfer than in frozen transfer, yet no statistically significant relationship was identified (AOR = 1.6; 95% CI: 0.54–12.4; P = 0.235). Furthermore, there were no significant differences in the type of infertility (AOR = 0.73; 95% CI: 0.34–1.6; P = 0.436) and preterm rates (AOR = 0.62; 95% CI: 0.33–5.5) between the groups.

Table 4.

) results of multiple logistic regression analysis to estimate crude and adjusted ORs and 95% CIs of pregnancy outcomes of fresh ET in comparison with frozen-thawed ET

Variables	Crude OR (95% CI)	P-value	Adjusted OR (95% CI)	P-value
Chemical pregnancy	1.34 (0.82–2.20)	0.146	1.31 (0.81–2.3)	0.238
Clinical pregnancy	1.51 (1.0-2.56)	0.048	1.51 (0.91–2.5)	0.125
Primary infertility	1.04 (0.38–2.79)	0.327	0.73 (0.34–1.6)	0.436
Live birth	1.43 (96.0-2.46)	0.075	1.6 (0.54–12.4)	0.235
Preterm labor	1.12 (0.27–4.58)	0.455	0.62 (0.33–5.5	0.514
Retrieved oocyte number	2.7 (1.75–6.57)	0.001	1.3 (1.16–1.75)	0.001

OR: Odds ratio; CI: Confidence interval

* Adjusted for age, BMI, history of ART, retrieved oocyte number, and cesarean section

Discussion

This study compared the pregnancy, prenatal, and postnatal outcomes between the frozen and fresh embryo transfers following intracytoplasmic sperm injection (ICSI) cycles at a referral infertility center in the northwest of Iran using a prospective design. The decision to choose the optimal ET strategy is often influenced by clinical features, proven indications, and patient satisfaction, making random allocation of patients challenging and, in some cases, unethical. In this study, to address potential biases (selection and information) and confounding variables such as age, prior ART history, BMI, oocyte quality and quantity, cesarean section, and infertility type, we conducted multiple logistic regression analysis to estimate adjusted ORs with 95% CIs. In the final analysis, although slight differences were observed between the two strategies, no significant differences were found regarding pregnancy and postnatal outcomes.

Infertility is one of the leading social health problems, with a prevalence of about 15% [27]. According to recent studies on the Iranian population, the prevalence of primary and secondary infertility was 79% and 21%, respectively [28]. In the current study, the frequency of primary infertility cases was 83.3%, and secondary infertility was 16.7%, which aligns with recent studies in Iranian society.

Today, the transfer of frozen or fresh embryos is widely used for infertility treatments worldwide, and the treatment results obtained in this respect are also different in several studies [29]. According to previous studies, the pregnancy rate after frozen embryo transfer cycles is lower than that in fresh embryos. After frozen embryo transfer cycles, newborns also have better birth weight and fewer perinatal complications than in fresh embryo transfer [30–34]. We also compared the live birth rate and fetal outcomes between fresh and frozen embryo transfer in ICSI cycles. Summarizing the results of the present study showed no difference between the two embryo transfer methods (frozen or fresh) in terms of pregnancy success rate. In line with our findings, Ku et al. reported no difference between the two methods [35]. The same findings were found in a review study conducted by Wong et al. [9].

In the current study, the chemical pregnancy rates in fresh and frozen embryo transfer groups were 21.8% and 17.2%, respectively. Similarly, Aflatoonian et al. reported 27% and 22.1% of the chemical pregnancy rates in fresh and frozen embryo groups, respectively [36]. In Iran, Basirat et al. also reported 23% and 18.8% of the chemical pregnancy rates in fresh and frozen embryo groups, respectively [37]. We found that the frequency of clinical pregnancy in the fresh and frozen embryo groups was 19% and 13.4%, respectively, and the rate of live births was 14.1% and 9.1%, respectively. In line with our results, Gu et al. reported clinical pregnancy rates of 37.5% for the frozen embryo group and 47.5% for the fresh embryo group, which aligns with our study [38]. Also, Wang et al. reported that the incidence of clinical pregnancy in all evaluated patients was 16.4%, which is comparable with our study [15]. Compared to other studies, the statistics of chemical and clinical pregnancy observed in the current study are lower, which seems to be due to the inclusion of patients at the age of 48 years and higher, which most of our patients are in the age group of 38 to 48 years. Some findings indicated that IVF outcomes may be improved by performing frozen ET compared with fresh ET [8].

Zhu et al. showed higher gestation and implantation rates in frozen blastocysts than in fresh transfer cycles. The clinical pregnancy rate of the fresh and frozen blastocyst transfer groups was 36.4% and 55.1%, respectively (p < 0.05), hile the implantation rates for the fresh and frozen group were 25.2% and 37.0% (p < 0.05) [39]. Kuc et al. showed that the clinical pregnancy rates for the verification and slow-freezing groups of day 5 or day 6 blastocysts were notably different. The clinical pregnancy rates of the slow-freezing and vitrification groups were 25.9% and 50.4% (p < 0.05), respectively [40]. Embryo transfer is a crucial stage in IVF, where the quality of the procedure influences the outcome. Recent evidence indicates that transvaginal ultrasound guidance during the transfer significantly enhances the percentage of pregnancies per transfer, both in the general population and in the reference population, compared to transfers conducted under transabdominal ultrasound guidance. For instance, this issue was emphasized in a review study and in research carried out by Larue et al. [41, 42]. Additionally, some evidence recommended that IVF outcomes may be improved by performing frozen ET compared to fresh ET [8]. In line with our study, a randomized controlled trial indicated that the ongoing pregnancy rate is lower in frozen ET compared to fresh (RR = 0.59; 0.36–0.98), while there is no benefit to a freeze-all strategy in cumulative ongoing pregnancy rates [25]. There are conflicting findings regarding various pregnancy and postnatal outcomes of fresh and frozen ETs. For instance, another study found that, compared to spontaneous pregnancies, fresh pregnancies—unlike frozen ones—were linked to lower birth weights and shorter gestational ages. Preterm birth was associated with both fresh and frozen transfers. The association of frozen embryos with advanced gestational age was notable. While pregnancy rates have improved in ART protocols, it remains essential to monitor adverse neonatal outcomes in these cases [43]. The reasons for such differences between studies, factors such as the study design and methods, age, indication parameters, ethnicity, sample size, and the history of ART can be the causes of the possible difference in the results.

Insogna et al., in comparing fresh and frozen embryo transfer, the rates of live births were 56.5% and 44%, respectively, and the rates of clinical pregnancies were 66.7% and 54.2%, respectively. The rate of spontaneous abortion was 9.3% and 9.4%, respectively. They reported a statistically significant difference between the two groups [44]. Acharya et al. reported that in high responders (number of oocytes received > 15) and moderate responders (number of oocytes received 6–14), live birth rates were better in frozen embryo transfer, and in contrast, in low responders (number of oocytes received 1–5), fresh embryo transfer has a better result [45].

Our results showed that the frequency of multiple pregnancies and births was 5% and 13.8% in the fresh and frozen embryo groups, respectively. Since one of the essential functions of frozen embryo transfer is to increase the chance of pregnancy by reducing the occurrence of multiple births as a risk factor of ART treatment failure, and even though the average number of transferred embryos is different in the two types of transfer methods, the results showed that the rate of twin pregnancy in fresh embryos is much lower than in frozen embryos; However, there is not a statistically significant difference. The 33.4% of frozen embryo transfers in Hajshafiha et al.‘s study resulted in multiple pregnancies, which is higher than ours and probably may be due to the different age distribution of the study and the average number of transferred embryos [46]. Mandelbaum et al., like our study, reported that the frequency of multiple pregnancies was 17.3% in the frozen embryo transfer [47]. In this study, we found no statistically significant difference between the two transfer methods. We recommend selecting the transfer method based on established indications, clinical characteristics, paraclinical results, and the patient’s condition and satisfaction [48]. We also suggest conducting other issues such as macrocosmic, large-for-gestational-age infants, and the impact of types of infertility, especially unexplained infertilities, in future studies.

Limitations and strengthens

In this prospective study, we compared pregnancy and fetal outcomes between fresh embryo and frozen-thawed embryo transfers. However, our study had some limitations. Although it was a cohort study conducted in a referral women’s hospital, potential confounders such as maternal age, BMI, obstetric characteristics, clinical features, infertility type, and previous ART cycles may distort the true associations between groups. To address this issue, we performed multiple logistic regression analyses to estimate adjusted ORs with 95% CIs. Randomized controlled trials using stratified randomization could randomly assign and balance the distribution of baseline and confounding variables in the study groups to obtain a true measure of associations. However, there were two barriers to conducting the randomized controlled trial, first, it is challenging to randomly assign patients for each of fresh and frozen ETs without considering clinical features and indication, and second, the representativeness and external validity of trial studies are diminished due to the limitations of inclusion criteria.

Conclusion

Although some pregnancy outcomes were slightly better in fresh embryo transfer than frozen, no significant statistical difference was found. Multicenter clinical trials and meta-analysis studies are recommended for better clinical decision-making. Our suggestion is to conduct future studies on other issues, like macrocosmic, large-for-gestational-age infants, and the impact of types of infertility, particularly unexplained infertility, in future studies.

Author contributions

RK and SFF participated in the protocol development, conducting, and supervision. PH, FT, MH, MA, NR, RV, and lS contributed to protocol development, data creation and collection, manuscript development, and review. HA analyzed, iterpreted, reviewed, and edited the manuscript. All authors read and approved the final manuscript.

Funding

This study was funded by Tabriz University of Medical Sciences, Iran.

Data availability

The datasets generated and/or analyzed during the current study are available from the corresponding author on reasonable request.

Declarations

Ethics approval and consent to participate

The study protocol was approved by the ethics committee of Tabriz University of Medical Sciences to number IR.TBZMED.REC.1400.599. Written informed consent was obtained before the study. The study was conducted in accordance with the Declaration of Helsinki.

Consent for publication

Not Applicable.

Competing interests

The authors declare no competing interests.