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Impact of monozygotic twinning on multiple births resulting from in vitro fertilization in the United States, 2006–2010
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REPRODUCTIVE ENDOCRINOLOGY AND INFERTILITY
Impact of monozygotic twinning on multiple births resulting from in vitro fertilization in the United States, 2006e2010 Rebekah E. Gee, MD; Richard P. Dickey, MD, PhD; Xu Xiong, MD, DrPH; LaToya S. Clark, MD; Gabriella Pridjian, MD OBJECTIVE: To determine the contribution of monozygotic twining to in vitro fertilization multiple births. STUDY DESIGN: We performed a retrospective analysis of the inci-
dence of monozygotic twining in multiple births resulting from fresh embryo transfers using 2006-2010 data from the Society for Reproductive Technology Clinic Outcome Reporting System. RESULTS: The number of embryos transferred were fewer than the number of births in 0.5% (223/40950) of twin, 29% (659/ 2289) of triplet, and 64% (43/67) of quadruplet births resulting
from transfer of fresh embryos from 2006 to 2010. In 2010, 37% of triplets and 100% of quadruplet births occurred when fewer than 3 and fewer than 4 embryos respectively were transferred. CONCLUSION: Monozygotic twinning plays a key role in the develop-
ment of triplet and quadruplet pregnancies achieved through in vitro fertilization. Key words: IVF, monozygotic twining, quadruplet birth, singleton birth, triplet birth, twin birth
Cite this article as: Gee RE, Dickey RP, Xiong X, et al. Impact of monozygotic twinning on multiple births resulting from in vitro fertilization in the United States, 2006e2010. Am J Obstet Gynecol 2014;210:468.e1-6.
M
onozygotic twins (MZT) carry a significantly higher risk of perinatal morbidity and mortality than singleton and dizygotic twins.1-5 They are associated with a greater risk of prematurity and increased risk of anomalies as well as complications from placental sharing such as twin-twin transfusion syndrome, selective intrauterine growth restriction, and reversed arterial perfusion sequence. These complications can progress to severe morbidity and fetal death. Complications because of placental sharing are not limited to twins
From the Louisiana State University Department of Obstetrics and Gynecology (Drs Dickey, Gee, and Clark), Louisiana State University School of Medicine, and School of Public Health, New Orleans, LA (Dr Gee); Tulane University Louisiana Department of Epidemiology, School of Public Health and Tropical Medicine, New Orleans, LA (Dr Xiong); Tulane University Department of Obstetrics and Gynecology, School of Medicine, New Orleans, LA (Dr Pridjian); and The Fertility Institute of New Orleans, Mandeville, LA (Dr Dickey). Received Sept. 21, 2013; revised Dec. 12, 2013; accepted Dec. 23, 2013. The authors report no conflict of interest. Reprints will not be available from the authors. 0002-9378/$36.00 ª 2014 Mosby, Inc. All rights reserved. http://dx.doi.org/10.1016/j.ajog.2013.12.034
and are higher in dichorionic triamniotic triplet pregnancies compared with trichorionic triplet pregnancies.6-8 Dichorionic/diamniotic twins occur with embryo division between fertilization and morula stage, 4 days postfertilization. Monochorionic/diamniotic twins occur with embryo division between days 4 and 8 postfertilization. Monochorionic/monoamniotic twins occur when embryo division happens between postfertilization days 8 and 12 and carry additional pregnancy risks because of shared placental mass and fetoplacental circulation. After day 13, embryo division results in conjoined twins. In natural conceptions, twothirds of monozygotic twins are monochorionic.9 In spontaneous pregnancies, MZT are an infrequent event with an incidence of 1 in 240 (0.41%) naturally conceived births.10-13 Monozygotic twining has been found in 24% to 44% of spontaneous triplet pregnancies.6-7,14 Monozygotic rates following assisted reproductive technology (ART) procedures are reported to be between 2 and 12 times higher than the natural occurrence of 0.4%.8,10,12,15-19 The number of quadruplet and higher births resulting from ARTreached a peak of 316 in 1997 when they contributed to 54% of quadruplet and higher order
468.e1 American Journal of Obstetrics & Gynecology MAY 2014
births in the United States.20 The number of triplet births resulting from ART peaked at 3498 in 1998, when they accounted for more than 50% of all triplet births in the United States.20 In response to these high numbers, the American Society of Reproductive Medicine and the Society for Assisted Reproductive Technology (SART) in 1998 and 1999 respectively published national guidelines limiting the number of fresh embryos transferred to 2 for patients age <35 and one for patients with the best prognosis and additional embryos to cryopreserve.22-23 Because of these recommendations, the number of triplet and quadruplet births because of ART has decreased markedly; however, the number of twins has not.21 We hypothesize that the failure of twin rates to decrease following in vitro fertilization (IVF), and triplet and higher order multiple rates to decrease further may in part be because of increased MZT twinning rates in the IVF population. To define the role of MZT in multiple births resulting from IVF, we conducted a retrospective analysis of the number of embryos transferred for all singleton and multiple births resulting from transfer of fresh nondonor embryos in the United States during the years
Reproductive Endocrinology and Infertility
www.AJOG.org 2006-2010 using the SART Clinic Online Reporting System (CORS). To remain consistent with the majority of past studies of MZT following assisted conception,12 we assumed MZT to have occurred when initial heart rates were greater than the number of embryos transferred.
M ATERIALS
AND
M ETHODS
Study population The records from the SART CORS dataset included 141,030 live births resulting from fresh nondonor embryo transfers for the years 2006-2010 (Figure). Data from more than 90% of clinics performing ART in the United States has been collected, verified by SART and reported to the Centers for Disease Control and Prevention since 1992 in compliance with the Fertility Clinic Success Rate and Certification Act of 1992 (Public Law 102-493, 24 October
Research
1992). Institutional review board approval for our study was obtained from SART, Louisiana State University School of Medicine, and Tulane University School of Medicine.
total of 140,547 births. This number included 4 triplet and 1 quadruplet birth reported following single embryo transfer.
Determination of zygosity Zygosity was determined by ultrasound during the first 6-7 weeks of gestation. Monozygotic twining was determined to have occurred when the maximum number of fetal heart rates exceeded the number of embryos transferred.
Statistical analysis c2 test and c2 test for trend were used to compare the percent of singleton and multiple births attributable to transfer of 1 to 5 or more embryos. Analysis was further stratified by year of birth. The proportion of births that were twin, triplet, and quadruplet following transfer of 1 to 5 or more embryos was calculated using WinPepi Describe version 2.51J. (available at: http://www. brixtonhealth.com/pepi4windows.html; accessed Feb. 13, 2014) with percent with 95% confidence interval. The CochraneArmitage test was used to determine trends. All P values were 2-tailed, at a significance of .05.
Inclusion and exclusion criteria All records were included for which the number of embryos transferred was recorded. Records were excluded from analysis if numbers of initial heart rates or the number of infants born were. After exclusions there were 97,236 singleton, 40,950 twin, 2293 triplet births, and 68 quadruplet births, for a
FIGURE
All live births resulting from IVF with fresh embryo transfer: 2006-2010
IVF, in vitro fertilization. Gee. Monozygotic twining contribution to multiple gestations in IVF. Am J Obstet Gynecol 2014.
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TABLE 1
Total births from fresh nondonor eggs: 2006-2010 Birthsa Year
n
Singleton
Twins
Triplets
Quadruplets
Ave (SD)
n (%)
n (%)
n (%)
n (%)
2006
26,035
2.50 (0.92)
18,054 (69.3)
7503 (28.8)
458 (1.76)
20 (0.08)
2007
27,415b
2.44 (0.93)
18,890 (68.9)
8038 (29.3)
476 (1.74)
11 (0.04)
2008
29,234
2.40 (0.90)
20,036 (68.5)
8670 (29.6)
512 (1.75)
16 (0.05)
2009
29,211
2.30 (0.85)
20,340 (69.6)
8413 (28.8)
441 (1.50)
17 (0.06)
2010
29,133
2.22 (0.82)
20,315 (69.7)
8399 (28.8)
415 (1.42)
4 (0.01)
Total
141,028
2.37 (0.89)
97,635 (69.2)
41,023 (29.1)
2302 (1.63)
68 (0.05)
< .001
.010
P valuec
.000
.829
.071
.509
Ave SD, average standard deviation. a
All births with number born reported, (includes cases with missing number of embryos transferred or missing initial number of heart beats); b One set quintuplets were stillborn; c P value CochraneArmitage test for trend.
Gee. Monozygotic twining contribution to multiple gestations in IVF. Am J Obstet Gynecol 2014.
R ESULTS
Table 2 presents the percent of singleton and multiple births following transfer of 1 through 5 or more embryos. The minimal proportion of twin triplet and quadruplet births attributable to monozygocity of 1 or more embryos following transfer is indicated. During the 5 years from 2006 through 2010, 1.91% of single embryo transfers resulted in birth of twins, triplets, or quadruplets. The 223 births because of MZT following single embryo transfer accounted for only 0.5% of all twin births. During the same period, 0.80% of double embryo transfers resulted in birth of triplets and quadruplets. However, the 673 births because of MZT following transfer of 2 embryos were responsible for 28.7% of triplet births
The maximum number of heart rates was greater than the number of embryos transferred in 1.19% of fresh embryo transfers (Figure). The number of heart rates exceeded the number of embryos in 2.16% of single embryo transfers, 1.52% of double embryo transfers, 0.43% of triple embryo transfers, 0.07% of quadruple embryo transfers, and 0.03% (1 case) following transfer of 5 or more embryos. Table 1 presents the average numbers of fresh nondonor embryos transferred and plurality of IVF births from 2006 to 2010. The overall percent of triplet and quadruplet births decreased significantly although the percent of singleton and twin births did not change. TABLE 2
Contribution of monozygotic twining to multiparty in 140,547 births conceived by IVF relationship of number of embryos transferred to number of infants born Plurality
Number of embryos transferred
Number born
1
1
11.8% 0.5%
a
3
0.2%
a
4
1.5%a,b
2
5
2
3
56.6%
21.9%
6.8%
2.8%
68.6%
22.2%
6.2%
2.3%
27.7%
53.3%
13.6%
4.3%
20.6%a
42.6%a
23.5%
11.8%
a
4
IVF, in vitro fertilization. a
Percent because of monozygotic twining; b One case, not verified.
Gee. Monozygotic twining contribution to multiple gestations in IVF. Am J Obstet Gynecol 2014.
468.e3 American Journal of Obstetrics & Gynecology MAY 2014
and 20.6% of quadruplet births. In all, 64.7% of quadruplet births resulted from transfer of less than 4 embryos. Table 3 presents the percent of singleton and multiple births following transfer of 1 through 5 or more embryos stratified by year. During a 5 year period, the percent of twin births because of MZT following single embryo transfer increased from 0.3% to 0.8%. The percent of twin births resulting from transfer of 3 or more embryos decreased from 38.6% to 24.0%. In 2010 all 4 quadruplet births resulted from the transfer of 2 or 3 embryos.
C OMMENT In this study, we show that MZT plays a significant role in the occurrence of triplet and quadruplet births following IVF but only a small role in twinning. Although the number of heart rates exceeded the number of embryos in 2.16% single embryo transfers, only 0.5% off total twin births were the result of MZT following single embryo transfers. The remaining 95% of twin births following IVF were the result of the transfer of 2 or more embryos. Our data does not indicate, however, how many of the multiple births following the transfer of 2 embryos were a result of MZT or both embryos implanting. Although MZT does not play a major role in overall twinning rates, MZT
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TABLE 3
Relationship of number of embryos transferred to multiple births because of IVF: 2006-2010a Variable
Total embryos
Number of embryos transferred (%) by number born
Year
n
Ave. SD
2006
17,973
2.47 0.87
1490 (8.3)
9604 (53.8)
4655 (25.9)
1611 (9.0)
613 (3.4)
2007
18,807
2.42 0.97
1843 (9.8)
10,341 (55.0)
4521 (24.0)
1429 (7.6)
673 (3.6)
2008
19,971
2.36 0.93
2096 (10.5)
11,395 (57.0)
4482 (22.4)
1392 (7.0)
606 (3.0)
2009
20,259
2.28 0.90
2632 (13.0)
11,806 (58.3)
4165 (20.6)
1172 (5.8)
485 (2.4)
2010
20,226
2.17 0.86
3383 (16.7)
11,901 (58.8)
3515 (17.4)
1022 (5.0)
405 (2.0)
Total
97,236
2.34 0.93
11,444 (11.8)
55,047 (56.6)
21,337 (21.9)
6626 (6.8)
2782 (2.8)
< .001
< .001
< .001
< .001
< .001
1
2
3
‡5
4
Singleton births
P valueb
< .001
.998
Twin births 2006
7489
2.54 0.61
25 (0.3)
4570 (61.0 )
2036 (27.2)
627 (8.4)
231 (3.1)
2007
8022
2.47 0.82
41 (0.5)
5232 (65.2)
1974 (24.6)
569 (7.1)
208 (2.6)
2008
8653
2.45 0.80
41 (0.5)
5832 (67.4)
1942 (22.4)
611 (7.0)
227(2.6)
2009
8403
2.35 0.71
49 (0.6)
6160 (73.3)
1648 (19.6)
386 (4.6)
160 (1.9)
2010
8381
2.31 0.73
67 (0.8)
6298 (75.1)
1513 (18.0)
374 (4.4)
129 (1.5)
40,950
2.42 0.78
223 (0.5)
28,092 (68.6)
9113 (22.2)
2567 (6.2)
955 (2.3)
< .001
< .001
< .001
< .001
< .001
92 (20.2)
266 (58.3)
71 (15.6)
27 ( 5.9)
Total P value
< .000
b
c
.668
Triplet births 2006
456
3.09 0.4
2007
472
3.02 0.84
2 (0.4)
116 (24.6)
254 (53.8)
79 (16.7)
23 ( 4.8)
2008
510
2.95 0.84
1 (0.2)
147 (28.8)
269 (52.7)
73 (14.3)
21 ( 4.1)
2009
437
2.85 0.75
149 (34.1)
225 (51.5)
48 (11.0)
15 ( 3.4)
2010
414
2.78 0.77
206 (49.8)
41 ( 9.9)
12 ( 2.9)
2293
2.94 0.82
1220 (53.2)
312 (13.6)
98 ( 4.3)
Total P value
b
.074
—
— 1 (0.2) c
.998
155 (37.4)
4 (0.2)
659 (28.7)
.914
< .001
c
.010
.001
.014
d
Quadruplet births 2006
20
3.50 0.90
1 (5.0)
8 (40.0)
8 (40.0)
2 (10.0)
2007
11
3.58 1.02
—
2 (18.2)
5 (45.4)
2 (18.2)
2 (18.2)
2008
16
3.19 0.80
—
4 (25.0)
6 (37.5)
5 (31.2)
1 ( 6.2)
2009
17
3.29 1.12
—
5 (29.4)
8 (47.0)
1 (5.9)
3 (17.6)
2010
4
2.50 0.58
—
0
0
68
3.33 1.06
Total P valueb
.038
.375
1 (5.0)
2 (50.0) c
1 (1.5)
14 (20.6)
> .99
2 (50.0) c
.017
29 (42.6) .693
c
16 (23.5)
8 (11.8)
.016
.986
Ave SD, average standard deviation; IVF, in vitro fertilization. a
Cases were excluded (n ¼ 483) if the number of embryos transferred, number of initial heart rates, or the number of infants born were missing; b Cochrane-Armitage test for the proportion of births that were twin, triplet, and quadruplet following transfer of 1 to 5 or more embryos. All P values were 2-tailed, at a significance of .05; c Presumed evidence of monozygotic twining; d In 2007, a quintuplet stillbirth occurred at 24 weeks after transfer of 5 embryos.
Gee. Monozygotic twining contribution to multiple gestations in IVF. Am J Obstet Gynecol 2014.
following transfer of 2 or 3 embryos was a significant cause of triplet and quadruplet births. The findings that
MZT following transfer of 2 embryos was responsible for 28.8% of triplet births, and that MZT following transfer
of 2 or 3 embryos was responsible for 64.7% of quadruplet births during the 5 year period of study has not been
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Reproductive Endocrinology and Infertility
reported in a series with such a magnitude of patients. Another important finding of this study was that the percent of twin, triplet, and quadruplet pregnancies because of MZT increased over the study period. This increase could be due to a decrease in transfers of more than 2 embryos, an incremental increase in fifth day transfer, changes in laboratory practices, or additional unexplained factors. It is also possible that pregnancies that began with 1 or 2 embryos but end as triplets and quadruplets have always occurred but the contribution of MZT was hidden by the fact that most high order pregnancies were the result of transfer of 4 or more embryos. The contribution of MZT may have become more apparent whereas the number of embryos being transferred trended downward in recent years. The important role for MZT as a cause of triplet and quadruplet pregnancy following IVF and possibly ovulation induction has been underappreciated. Surprisingly few series have reported the chorionicity of spontaneously conceived triplets and most are of cases referred for twin-twin transfusion syndrome or other complications because of monochorionic dizygotic and trizygotic twining. The percent of MZT in triplet births resulting from IVF may be less than in spontaneous triplets. Machin and Bamforth14 reviewed the literature in 1996 and found only 38 sets of presumably spontaneously conceived triplets including 15 of their own. Of this number, 9 (24%) were monochorionic, 17 (45%) were dichorionic, and 12 (32%) were trichorionic. More recently Adegbite et al7 reported the perinatal outcomes of 88 sets of spontaneously conceived triplets referred to a tertiary center of which 39 (44%) were dichorionic triaminiotic and none were monochorionic triamniotic. Chow et al16 all reported a much lower incidence of MZT in 76 triplets and 10 quadruplets conceived by IVF that were referred to their institution for ultrasound scanning. The 1.19% incidence of MZT found in this study using early ultrasound was twice as high as s reported in a 2009 metaanalysis.12 In the metaanalysis, more than half the cases were from a
single study24 that was not limited to pregnancies ending in live births. In 14 of 21 studies in that metaanalysis when MZT was diagnosed by early trimester ultrasound studies, the incidence of MZTwas greater than ours. Determining the incidence of MZT by early ultrasound may underestimate the rate of MZT because it does not identify embryos that are the result of an embryo splitting when the total number of heart rates are equal to or less than the number of embryos transferred. Except for cases in which a single embryo is transferred, the true incidence of MZT can only be determined by placental histology or DNA analysis. In studies of MZT in multiple births resulting from IVF and diagnosed by placental histology, the incidence of MZT has ranged from 2.0% to 7.2% (average 3.2%).17,25,26 In a study which investigated the incidence of MZT diagnosed by DNA and placental histology in multiple births conceived by ovulation induction with clomiphene and human menopausal gonadotropin alone or in combination, the incidence of MZT was 1.2% ovulation induction compared with 0.45% following spontaneous ovulation.11 In large populations, the rate of MZT is commonly determined by Weinberg’s differential method where the incidence of MZT is estimated to be equal to the number of same sex twins remaining after subtracting double the number of opposite sex twins from the total number of twins.27 In a national study of multiple births resulting from 9954 IVF and 2,181,698 spontaneous pregnancies, the incidence of MZT according to Weinberg’s method was 1.11% and 0.43% respectively.13 This strengths of this study are that it includes 90% of all live births resulting from IVF during the years 2006 to 2010, and has both early ultrasound and birth outcome from greater than 140,000 IVF cycles. Our large sample size allows for a comprehensive and statistically significant examination of the relationship between numbers of embryos transferred and the number of resulting multiple births. A limitation of our data is that it did not include the number of transfers that ended in spontaneous or
468.e5 American Journal of Obstetrics & Gynecology MAY 2014
www.AJOG.org elective abortion. We included all multiple births with at least 1 live born because our objective was to determine the role of MZT in multiple pregnancies regardless of outcome. However, the true rate of monozygotic twinning after ART is undoubtedly underreported because only pregnancies ending in live birth were included. It is known that 50 to 60% of spontaneous pregnancies that begin as twins diagnosed by ultrasound are born as singletons.28-30 Our study confirms an increased incidence of MZT in pregnancies conceived by IVF compared with naturally conceived pregnancies, and helps to clarify the role of MZT in twin and higher order multiple births. At present, most IVF twins are the result of the transfer of 2 or more embryos but the contribution of MZT to twinning rates may be expected to increase as the practice of IVF trends toward single embryo transfer. This trend can already be observed in the case of triplets and quadruplets where 37% and 76%, respectively, were the result of MZT in the last years of this study. We believe that more research is needed on both the cause and the contribution of MZT to IVF pregnancies. One important step in advancing the research on MZT would be to include the numbers of twin, triplet, and quadruplet births in which the number born is greater than the number of embryos transferred in the Centers for Disease Control and Prevention National Summary and Fertility Clinic Report. Our study reinforces the recommendation that single embryos should be the preferred method of transfer for all IVF patients, not just those considered high risk if they were to become pregnant with twins. Lastly, the authors believe that explaining the risk of triplet and quadruplet births following the transfer of 2 or more embryos should be part of pretransfer counseling. ACKNOWLEDGEMENTS SART wishes to thank all of its members for providing clinical information to the SART CORS database for use by patients and researchers. Without the efforts of our members, this research would not have been possible.
www.AJOG.org REFERENCES 1. Sebire NJ, Snijders RJ, Hughes K, et al. The hidden mortality of monochorionic twin pregnancies. Br J Obstet Gynaecol 1997;104: 1203. 2. Ortibus E, Lopriore E, Deprest J, et al. The pregnancy and long-term neurodevelopmental outcome of monochorionic diamniotic twin gestations: a multicenter prospective cohort study from the first trimester onward. Am J Obstet Gynecol 2009;200:494.e1-8. 3. McPherson JA, Odibo AO, Shanks AL, et al. Impact of chorionicity on risk and timing of intrauterine fetal demise in twin pregnancies. Am J Obstet Gynecol 2012;207:190.e1-6. 4. Glinianaia SV, Obeysekera MA, Sturgiss S, Bell R. Stillbirth and neonatal mortality in monochorionic and dichorionic twins: a populationbased study. Hum Reprod 2011;26:2549. 5. Bajoria R, Kingdom J. The case for routine determination of chorionicity and zygosity in multiple pregnancy. Prenat Diagn 1997;17: 1207-25. 6. Bajora R, Ward SB, Adegbite AL. Comparative study of perinatal outcome of dichorionic and trichorionic iatrogenic triplets. Am J Obstet Gynecol 2006;194:415-24. 7. Adegbite AL, Ward SB, Bajora R. Perinatal outcome of spontaneoulsy conceived triplet pregnancies in relation to chorionicity. Am J Obstet Gynecol 2005;193:1463-71. 8. Derom R, Derom C, Vlietwick R, vanden Berghe M. Twin pregnancies after medically assisted reproduction;epidemiology, comparative perinatal morbidity and mortality. J Perinatol Med 1991;19:229-33. 9. Newman R, Ramsey Unal E. Multiple gestations. In: Gabbe SG, Niebyl JR, Galan H, et al, eds. Obstetrics: normal and problem pregnancies, 6th ed. Philadelphia, PA: Saunders; 2012. 10. Aston KI, Peterson CM, Carrell DT. Monozygotic twinning associated with assisted reproductive techniques: a review. Reproduction 2008;136:377-86. 11. Derom C, Vlietinck R, Derom R, Van den Berghe H, Thiery M. Increased monozygotic
Reproductive Endocrinology and Infertility twining rate after ovulation induction. Lancet 1987;i:1236-8. 12. Vitthala S, Gelbaya TA, Brison DR, Fitzgerald CT, Nardo KG. The risk of monozygotic twins after assisted reproductive technology: a systamatic review and meta-analysis. Hum Reprod Update 2009;15:45-55. 13. Tandberg A, Bjørge T, Børdahl PE, Skjaerven R. Increasing twining rates in Norway, 1967-2004: the influence of maternal age and assisted reproductive technecoloty. Acta Obstet Gynecol Scand 2007;86:833-9. 14. Machin GA, Bamforth. Zygosity and placental anatomy in 15 consecutive sets of spontaneously conceived triplets. Am J Med Genetics 1996;61:247-52. 15. Skiadas CC, Missmer SA, Benson CB, Gee RE, Racowsky C. Risk factors associated with pregnancies containing a monochorionic pair following assisted reproductive technologies. Hum Reprod 2008;23:1366-71. 16. Chow JS, Benson CB, Racowsky C, Doubilet PM, Ginsburg E. Frequency of a monochorionic pair in multiple gestations in relationship to mode of conception. J Ultrasound Med 2001;20:757-60. 17. Wenstrom KD, Syrop CH, Hammit DG, Van Voorhis BJ. Increased risk of monochronic twinning associated with assisted reproduction. Fertil Steril 1993;60:510-4. 18. Alikani M, Cekleniak NA, Walters E, Cohen J. Monozygotic twinning following assisted conception: an analysis of 81consecutive cases. Hum Reprod 2003;18:1937-43. 19. Schieve LA, Meikle SF, Peterson HB, Jeng G, Burnett NM, Wikcox LS. Does assisted hatching pose a risk for monozygotic twinning in pregnancies conceived through in vitro fertilization? Fertil Steril 2000;74:288-94. 20. Centers for Disease Control and Prevention. 2010 Assisted Reproductive Technology success rates: National Summary and Fertility Clinic Reports. Atlanta, GA: CDC; 2013. 21. Dickey RP. The relative contribution of assisted reproductive technologies and ovulation
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REPRODUCTIVE ENDOCRINOLOGY AND INFERTILITY
Impact of monozygotic twinning on multiple births resulting from in vitro fertilization in the United States, 2006e2010 Rebekah E. Gee, MD; Richard P. Dickey, MD, PhD; Xu Xiong, MD, DrPH; LaToya S. Clark, MD; Gabriella Pridjian, MD OBJECTIVE: To determine the contribution of monozygotic twining to in vitro fertilization multiple births. STUDY DESIGN: We performed a retrospective analysis of the inci-
dence of monozygotic twining in multiple births resulting from fresh embryo transfers using 2006-2010 data from the Society for Reproductive Technology Clinic Outcome Reporting System. RESULTS: The number of embryos transferred were fewer than the number of births in 0.5% (223/40950) of twin, 29% (659/ 2289) of triplet, and 64% (43/67) of quadruplet births resulting
from transfer of fresh embryos from 2006 to 2010. In 2010, 37% of triplets and 100% of quadruplet births occurred when fewer than 3 and fewer than 4 embryos respectively were transferred. CONCLUSION: Monozygotic twinning plays a key role in the develop-
ment of triplet and quadruplet pregnancies achieved through in vitro fertilization. Key words: IVF, monozygotic twining, quadruplet birth, singleton birth, triplet birth, twin birth
Cite this article as: Gee RE, Dickey RP, Xiong X, et al. Impact of monozygotic twinning on multiple births resulting from in vitro fertilization in the United States, 2006e2010. Am J Obstet Gynecol 2014;210:468.e1-6.
M
onozygotic twins (MZT) carry a significantly higher risk of perinatal morbidity and mortality than singleton and dizygotic twins.1-5 They are associated with a greater risk of prematurity and increased risk of anomalies as well as complications from placental sharing such as twin-twin transfusion syndrome, selective intrauterine growth restriction, and reversed arterial perfusion sequence. These complications can progress to severe morbidity and fetal death. Complications because of placental sharing are not limited to twins
From the Louisiana State University Department of Obstetrics and Gynecology (Drs Dickey, Gee, and Clark), Louisiana State University School of Medicine, and School of Public Health, New Orleans, LA (Dr Gee); Tulane University Louisiana Department of Epidemiology, School of Public Health and Tropical Medicine, New Orleans, LA (Dr Xiong); Tulane University Department of Obstetrics and Gynecology, School of Medicine, New Orleans, LA (Dr Pridjian); and The Fertility Institute of New Orleans, Mandeville, LA (Dr Dickey). Received Sept. 21, 2013; revised Dec. 12, 2013; accepted Dec. 23, 2013. The authors report no conflict of interest. Reprints will not be available from the authors. 0002-9378/$36.00 ª 2014 Mosby, Inc. All rights reserved. http://dx.doi.org/10.1016/j.ajog.2013.12.034
and are higher in dichorionic triamniotic triplet pregnancies compared with trichorionic triplet pregnancies.6-8 Dichorionic/diamniotic twins occur with embryo division between fertilization and morula stage, 4 days postfertilization. Monochorionic/diamniotic twins occur with embryo division between days 4 and 8 postfertilization. Monochorionic/monoamniotic twins occur when embryo division happens between postfertilization days 8 and 12 and carry additional pregnancy risks because of shared placental mass and fetoplacental circulation. After day 13, embryo division results in conjoined twins. In natural conceptions, twothirds of monozygotic twins are monochorionic.9 In spontaneous pregnancies, MZT are an infrequent event with an incidence of 1 in 240 (0.41%) naturally conceived births.10-13 Monozygotic twining has been found in 24% to 44% of spontaneous triplet pregnancies.6-7,14 Monozygotic rates following assisted reproductive technology (ART) procedures are reported to be between 2 and 12 times higher than the natural occurrence of 0.4%.8,10,12,15-19 The number of quadruplet and higher births resulting from ARTreached a peak of 316 in 1997 when they contributed to 54% of quadruplet and higher order
468.e1 American Journal of Obstetrics & Gynecology MAY 2014
births in the United States.20 The number of triplet births resulting from ART peaked at 3498 in 1998, when they accounted for more than 50% of all triplet births in the United States.20 In response to these high numbers, the American Society of Reproductive Medicine and the Society for Assisted Reproductive Technology (SART) in 1998 and 1999 respectively published national guidelines limiting the number of fresh embryos transferred to 2 for patients age <35 and one for patients with the best prognosis and additional embryos to cryopreserve.22-23 Because of these recommendations, the number of triplet and quadruplet births because of ART has decreased markedly; however, the number of twins has not.21 We hypothesize that the failure of twin rates to decrease following in vitro fertilization (IVF), and triplet and higher order multiple rates to decrease further may in part be because of increased MZT twinning rates in the IVF population. To define the role of MZT in multiple births resulting from IVF, we conducted a retrospective analysis of the number of embryos transferred for all singleton and multiple births resulting from transfer of fresh nondonor embryos in the United States during the years
Reproductive Endocrinology and Infertility
www.AJOG.org 2006-2010 using the SART Clinic Online Reporting System (CORS). To remain consistent with the majority of past studies of MZT following assisted conception,12 we assumed MZT to have occurred when initial heart rates were greater than the number of embryos transferred.
M ATERIALS
AND
M ETHODS
Study population The records from the SART CORS dataset included 141,030 live births resulting from fresh nondonor embryo transfers for the years 2006-2010 (Figure). Data from more than 90% of clinics performing ART in the United States has been collected, verified by SART and reported to the Centers for Disease Control and Prevention since 1992 in compliance with the Fertility Clinic Success Rate and Certification Act of 1992 (Public Law 102-493, 24 October
Research
1992). Institutional review board approval for our study was obtained from SART, Louisiana State University School of Medicine, and Tulane University School of Medicine.
total of 140,547 births. This number included 4 triplet and 1 quadruplet birth reported following single embryo transfer.
Determination of zygosity Zygosity was determined by ultrasound during the first 6-7 weeks of gestation. Monozygotic twining was determined to have occurred when the maximum number of fetal heart rates exceeded the number of embryos transferred.
Statistical analysis c2 test and c2 test for trend were used to compare the percent of singleton and multiple births attributable to transfer of 1 to 5 or more embryos. Analysis was further stratified by year of birth. The proportion of births that were twin, triplet, and quadruplet following transfer of 1 to 5 or more embryos was calculated using WinPepi Describe version 2.51J. (available at: http://www. brixtonhealth.com/pepi4windows.html; accessed Feb. 13, 2014) with percent with 95% confidence interval. The CochraneArmitage test was used to determine trends. All P values were 2-tailed, at a significance of .05.
Inclusion and exclusion criteria All records were included for which the number of embryos transferred was recorded. Records were excluded from analysis if numbers of initial heart rates or the number of infants born were. After exclusions there were 97,236 singleton, 40,950 twin, 2293 triplet births, and 68 quadruplet births, for a
FIGURE
All live births resulting from IVF with fresh embryo transfer: 2006-2010
IVF, in vitro fertilization. Gee. Monozygotic twining contribution to multiple gestations in IVF. Am J Obstet Gynecol 2014.
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TABLE 1
Total births from fresh nondonor eggs: 2006-2010 Birthsa Year
n
Singleton
Twins
Triplets
Quadruplets
Ave (SD)
n (%)
n (%)
n (%)
n (%)
2006
26,035
2.50 (0.92)
18,054 (69.3)
7503 (28.8)
458 (1.76)
20 (0.08)
2007
27,415b
2.44 (0.93)
18,890 (68.9)
8038 (29.3)
476 (1.74)
11 (0.04)
2008
29,234
2.40 (0.90)
20,036 (68.5)
8670 (29.6)
512 (1.75)
16 (0.05)
2009
29,211
2.30 (0.85)
20,340 (69.6)
8413 (28.8)
441 (1.50)
17 (0.06)
2010
29,133
2.22 (0.82)
20,315 (69.7)
8399 (28.8)
415 (1.42)
4 (0.01)
Total
141,028
2.37 (0.89)
97,635 (69.2)
41,023 (29.1)
2302 (1.63)
68 (0.05)
< .001
.010
P valuec
.000
.829
.071
.509
Ave SD, average standard deviation. a
All births with number born reported, (includes cases with missing number of embryos transferred or missing initial number of heart beats); b One set quintuplets were stillborn; c P value CochraneArmitage test for trend.
Gee. Monozygotic twining contribution to multiple gestations in IVF. Am J Obstet Gynecol 2014.
R ESULTS
Table 2 presents the percent of singleton and multiple births following transfer of 1 through 5 or more embryos. The minimal proportion of twin triplet and quadruplet births attributable to monozygocity of 1 or more embryos following transfer is indicated. During the 5 years from 2006 through 2010, 1.91% of single embryo transfers resulted in birth of twins, triplets, or quadruplets. The 223 births because of MZT following single embryo transfer accounted for only 0.5% of all twin births. During the same period, 0.80% of double embryo transfers resulted in birth of triplets and quadruplets. However, the 673 births because of MZT following transfer of 2 embryos were responsible for 28.7% of triplet births
The maximum number of heart rates was greater than the number of embryos transferred in 1.19% of fresh embryo transfers (Figure). The number of heart rates exceeded the number of embryos in 2.16% of single embryo transfers, 1.52% of double embryo transfers, 0.43% of triple embryo transfers, 0.07% of quadruple embryo transfers, and 0.03% (1 case) following transfer of 5 or more embryos. Table 1 presents the average numbers of fresh nondonor embryos transferred and plurality of IVF births from 2006 to 2010. The overall percent of triplet and quadruplet births decreased significantly although the percent of singleton and twin births did not change. TABLE 2
Contribution of monozygotic twining to multiparty in 140,547 births conceived by IVF relationship of number of embryos transferred to number of infants born Plurality
Number of embryos transferred
Number born
1
1
11.8% 0.5%
a
3
0.2%
a
4
1.5%a,b
2
5
2
3
56.6%
21.9%
6.8%
2.8%
68.6%
22.2%
6.2%
2.3%
27.7%
53.3%
13.6%
4.3%
20.6%a
42.6%a
23.5%
11.8%
a
4
IVF, in vitro fertilization. a
Percent because of monozygotic twining; b One case, not verified.
Gee. Monozygotic twining contribution to multiple gestations in IVF. Am J Obstet Gynecol 2014.
468.e3 American Journal of Obstetrics & Gynecology MAY 2014
and 20.6% of quadruplet births. In all, 64.7% of quadruplet births resulted from transfer of less than 4 embryos. Table 3 presents the percent of singleton and multiple births following transfer of 1 through 5 or more embryos stratified by year. During a 5 year period, the percent of twin births because of MZT following single embryo transfer increased from 0.3% to 0.8%. The percent of twin births resulting from transfer of 3 or more embryos decreased from 38.6% to 24.0%. In 2010 all 4 quadruplet births resulted from the transfer of 2 or 3 embryos.
C OMMENT In this study, we show that MZT plays a significant role in the occurrence of triplet and quadruplet births following IVF but only a small role in twinning. Although the number of heart rates exceeded the number of embryos in 2.16% single embryo transfers, only 0.5% off total twin births were the result of MZT following single embryo transfers. The remaining 95% of twin births following IVF were the result of the transfer of 2 or more embryos. Our data does not indicate, however, how many of the multiple births following the transfer of 2 embryos were a result of MZT or both embryos implanting. Although MZT does not play a major role in overall twinning rates, MZT
Reproductive Endocrinology and Infertility
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TABLE 3
Relationship of number of embryos transferred to multiple births because of IVF: 2006-2010a Variable
Total embryos
Number of embryos transferred (%) by number born
Year
n
Ave. SD
2006
17,973
2.47 0.87
1490 (8.3)
9604 (53.8)
4655 (25.9)
1611 (9.0)
613 (3.4)
2007
18,807
2.42 0.97
1843 (9.8)
10,341 (55.0)
4521 (24.0)
1429 (7.6)
673 (3.6)
2008
19,971
2.36 0.93
2096 (10.5)
11,395 (57.0)
4482 (22.4)
1392 (7.0)
606 (3.0)
2009
20,259
2.28 0.90
2632 (13.0)
11,806 (58.3)
4165 (20.6)
1172 (5.8)
485 (2.4)
2010
20,226
2.17 0.86
3383 (16.7)
11,901 (58.8)
3515 (17.4)
1022 (5.0)
405 (2.0)
Total
97,236
2.34 0.93
11,444 (11.8)
55,047 (56.6)
21,337 (21.9)
6626 (6.8)
2782 (2.8)
< .001
< .001
< .001
< .001
< .001
1
2
3
‡5
4
Singleton births
P valueb
< .001
.998
Twin births 2006
7489
2.54 0.61
25 (0.3)
4570 (61.0 )
2036 (27.2)
627 (8.4)
231 (3.1)
2007
8022
2.47 0.82
41 (0.5)
5232 (65.2)
1974 (24.6)
569 (7.1)
208 (2.6)
2008
8653
2.45 0.80
41 (0.5)
5832 (67.4)
1942 (22.4)
611 (7.0)
227(2.6)
2009
8403
2.35 0.71
49 (0.6)
6160 (73.3)
1648 (19.6)
386 (4.6)
160 (1.9)
2010
8381
2.31 0.73
67 (0.8)
6298 (75.1)
1513 (18.0)
374 (4.4)
129 (1.5)
40,950
2.42 0.78
223 (0.5)
28,092 (68.6)
9113 (22.2)
2567 (6.2)
955 (2.3)
< .001
< .001
< .001
< .001
< .001
92 (20.2)
266 (58.3)
71 (15.6)
27 ( 5.9)
Total P value
< .000
b
c
.668
Triplet births 2006
456
3.09 0.4
2007
472
3.02 0.84
2 (0.4)
116 (24.6)
254 (53.8)
79 (16.7)
23 ( 4.8)
2008
510
2.95 0.84
1 (0.2)
147 (28.8)
269 (52.7)
73 (14.3)
21 ( 4.1)
2009
437
2.85 0.75
149 (34.1)
225 (51.5)
48 (11.0)
15 ( 3.4)
2010
414
2.78 0.77
206 (49.8)
41 ( 9.9)
12 ( 2.9)
2293
2.94 0.82
1220 (53.2)
312 (13.6)
98 ( 4.3)
Total P value
b
.074
—
— 1 (0.2) c
.998
155 (37.4)
4 (0.2)
659 (28.7)
.914
< .001
c
.010
.001
.014
d
Quadruplet births 2006
20
3.50 0.90
1 (5.0)
8 (40.0)
8 (40.0)
2 (10.0)
2007
11
3.58 1.02
—
2 (18.2)
5 (45.4)
2 (18.2)
2 (18.2)
2008
16
3.19 0.80
—
4 (25.0)
6 (37.5)
5 (31.2)
1 ( 6.2)
2009
17
3.29 1.12
—
5 (29.4)
8 (47.0)
1 (5.9)
3 (17.6)
2010
4
2.50 0.58
—
0
0
68
3.33 1.06
Total P valueb
.038
.375
1 (5.0)
2 (50.0) c
1 (1.5)
14 (20.6)
> .99
2 (50.0) c
.017
29 (42.6) .693
c
16 (23.5)
8 (11.8)
.016
.986
Ave SD, average standard deviation; IVF, in vitro fertilization. a
Cases were excluded (n ¼ 483) if the number of embryos transferred, number of initial heart rates, or the number of infants born were missing; b Cochrane-Armitage test for the proportion of births that were twin, triplet, and quadruplet following transfer of 1 to 5 or more embryos. All P values were 2-tailed, at a significance of .05; c Presumed evidence of monozygotic twining; d In 2007, a quintuplet stillbirth occurred at 24 weeks after transfer of 5 embryos.
Gee. Monozygotic twining contribution to multiple gestations in IVF. Am J Obstet Gynecol 2014.
following transfer of 2 or 3 embryos was a significant cause of triplet and quadruplet births. The findings that
MZT following transfer of 2 embryos was responsible for 28.8% of triplet births, and that MZT following transfer
of 2 or 3 embryos was responsible for 64.7% of quadruplet births during the 5 year period of study has not been
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Reproductive Endocrinology and Infertility
reported in a series with such a magnitude of patients. Another important finding of this study was that the percent of twin, triplet, and quadruplet pregnancies because of MZT increased over the study period. This increase could be due to a decrease in transfers of more than 2 embryos, an incremental increase in fifth day transfer, changes in laboratory practices, or additional unexplained factors. It is also possible that pregnancies that began with 1 or 2 embryos but end as triplets and quadruplets have always occurred but the contribution of MZT was hidden by the fact that most high order pregnancies were the result of transfer of 4 or more embryos. The contribution of MZT may have become more apparent whereas the number of embryos being transferred trended downward in recent years. The important role for MZT as a cause of triplet and quadruplet pregnancy following IVF and possibly ovulation induction has been underappreciated. Surprisingly few series have reported the chorionicity of spontaneously conceived triplets and most are of cases referred for twin-twin transfusion syndrome or other complications because of monochorionic dizygotic and trizygotic twining. The percent of MZT in triplet births resulting from IVF may be less than in spontaneous triplets. Machin and Bamforth14 reviewed the literature in 1996 and found only 38 sets of presumably spontaneously conceived triplets including 15 of their own. Of this number, 9 (24%) were monochorionic, 17 (45%) were dichorionic, and 12 (32%) were trichorionic. More recently Adegbite et al7 reported the perinatal outcomes of 88 sets of spontaneously conceived triplets referred to a tertiary center of which 39 (44%) were dichorionic triaminiotic and none were monochorionic triamniotic. Chow et al16 all reported a much lower incidence of MZT in 76 triplets and 10 quadruplets conceived by IVF that were referred to their institution for ultrasound scanning. The 1.19% incidence of MZT found in this study using early ultrasound was twice as high as s reported in a 2009 metaanalysis.12 In the metaanalysis, more than half the cases were from a
single study24 that was not limited to pregnancies ending in live births. In 14 of 21 studies in that metaanalysis when MZT was diagnosed by early trimester ultrasound studies, the incidence of MZTwas greater than ours. Determining the incidence of MZT by early ultrasound may underestimate the rate of MZT because it does not identify embryos that are the result of an embryo splitting when the total number of heart rates are equal to or less than the number of embryos transferred. Except for cases in which a single embryo is transferred, the true incidence of MZT can only be determined by placental histology or DNA analysis. In studies of MZT in multiple births resulting from IVF and diagnosed by placental histology, the incidence of MZT has ranged from 2.0% to 7.2% (average 3.2%).17,25,26 In a study which investigated the incidence of MZT diagnosed by DNA and placental histology in multiple births conceived by ovulation induction with clomiphene and human menopausal gonadotropin alone or in combination, the incidence of MZT was 1.2% ovulation induction compared with 0.45% following spontaneous ovulation.11 In large populations, the rate of MZT is commonly determined by Weinberg’s differential method where the incidence of MZT is estimated to be equal to the number of same sex twins remaining after subtracting double the number of opposite sex twins from the total number of twins.27 In a national study of multiple births resulting from 9954 IVF and 2,181,698 spontaneous pregnancies, the incidence of MZT according to Weinberg’s method was 1.11% and 0.43% respectively.13 This strengths of this study are that it includes 90% of all live births resulting from IVF during the years 2006 to 2010, and has both early ultrasound and birth outcome from greater than 140,000 IVF cycles. Our large sample size allows for a comprehensive and statistically significant examination of the relationship between numbers of embryos transferred and the number of resulting multiple births. A limitation of our data is that it did not include the number of transfers that ended in spontaneous or
468.e5 American Journal of Obstetrics & Gynecology MAY 2014
www.AJOG.org elective abortion. We included all multiple births with at least 1 live born because our objective was to determine the role of MZT in multiple pregnancies regardless of outcome. However, the true rate of monozygotic twinning after ART is undoubtedly underreported because only pregnancies ending in live birth were included. It is known that 50 to 60% of spontaneous pregnancies that begin as twins diagnosed by ultrasound are born as singletons.28-30 Our study confirms an increased incidence of MZT in pregnancies conceived by IVF compared with naturally conceived pregnancies, and helps to clarify the role of MZT in twin and higher order multiple births. At present, most IVF twins are the result of the transfer of 2 or more embryos but the contribution of MZT to twinning rates may be expected to increase as the practice of IVF trends toward single embryo transfer. This trend can already be observed in the case of triplets and quadruplets where 37% and 76%, respectively, were the result of MZT in the last years of this study. We believe that more research is needed on both the cause and the contribution of MZT to IVF pregnancies. One important step in advancing the research on MZT would be to include the numbers of twin, triplet, and quadruplet births in which the number born is greater than the number of embryos transferred in the Centers for Disease Control and Prevention National Summary and Fertility Clinic Report. Our study reinforces the recommendation that single embryos should be the preferred method of transfer for all IVF patients, not just those considered high risk if they were to become pregnant with twins. Lastly, the authors believe that explaining the risk of triplet and quadruplet births following the transfer of 2 or more embryos should be part of pretransfer counseling. ACKNOWLEDGEMENTS SART wishes to thank all of its members for providing clinical information to the SART CORS database for use by patients and researchers. Without the efforts of our members, this research would not have been possible.
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