Traditionally, twinning mechanisms are classified as dizygotic twinning (DZT) and monozygotic twinning (MZT). DZT results from the fertilization of two distinct ova and shares approximately 50% DNA sequence identity, whereas MZT occurs by the splitting of a single embryo in 1–14 days following fertilization and shares 100% DNA sequence identity. The embryonic division in MZT is called a ‘twinning event’; however, the mechanisms that trigger the twinning event remain unknown (Hall, Reference Hall2003). Sesquizygotic twinning (SZT) is considered an exceptional intermediate between MZT and DZT, which shares between 50% and 100% of genetic identity, and the idea of SZT was proposed decades earlier than the discovery of clinical cases. SZT may arise from the splitting of chimeric embryos containing one genetic contribution from one parent and two genetic contributions from the other parent. When monochorionic twins exhibit significantly different traits, such as discordant genders, the possibility of being SZT should be considered. However, it should be noted that monochorionic DZT and heterokaryotypic MZT can also lead to similar results. SZT was first reported in 2007, in which the authors declared that they identified a third twinning mechanism. The report described an unusual, previously unreported twinning phenomenon, in which one fetus was a 46XX/46XY chimera with a phenotypically normal male reproductive system, whereas the other was a 46XX/46XY chimera with ambiguous external genitalia and was identified as true hermaphroditism. The fetuses shared 100% of the maternal alleles and approximately 50% of the paternal alleles based on a DNA analysis of skin fibroblasts. Another report described a pair of sex discordant, monochorionic 46XX/46XY chimeric twins without sexual ambiguity, in which the individuals shared 100% of the maternal genome and 77.7% of the paternal genome. The XX/XY ratio in the chimeric gonad cells played a major role in the gender formation of the fetuses. The authors of this report defined SZT as a third twinning mechanism, which is different from MZT and DZT. ‘Dispermic fertilization’ and ‘parthenogenetic activation’ were proposed as mechanisms to explain the origin of chimeric embryos in both SZT studies. Dispermic fertilization, in which an ovum is fertilized by two sperms containing different sex chromosomes, results in the formation of a 69XXY triploid zygote, and a unique phenomenon known as post-zygotic diploidization may occur to randomly remove one paternal genome at the cleavage stage of the triploid zygote. This generates a 46XX/46XY chimeric embryo containing one single maternal and two different paternal genomes, which is a subsequent twinning event resulting in the formation of SZT. However, it is important to note that triploid zygote diploidization is considered one of the causes of uniparental disomy (UPD), which leads to genomic imprinting disorders. For example, UPD15 can result in Angelman syndrome, while UPD2 may lead to fetal intrauterine death. Therefore, for potential SZT fetuses, UPD should be given special consideration in prenatal diagnosis. Parthenogenetic activation triggers the cleavage of the ovum to produce two cells with the same maternal genetic information in one zona pellucida. If the two cells are fertilized by two sperms containing different sex chromosomes, it may result in the formation of a 46XX/46XY chimeric embryo (Figure 1) (Gabbett et al., Reference Gabbett, Laporte, Sekar, Nandini, Mcgrath, Sapkota, Jiang, Zhang, Burgess and Montgomery2019; Giltay et al., Reference Giltay, Brunt, Beemer, Wit, Van Amstel, Pearson and Wijmenga1998; Golubovsky, Reference Golubovsky2003; Souter et al., Reference Souter, Parisi, Nyholt, Kapur, Henders, Opheim, Gunther, Mitchell, Glass and Montgomery2007; Denkboy Ongen et al., Reference Denkboy Ongen, Parlak and Eren2024; Gringras and Chen, Reference Gringras and Chen2001; Pearson et al., Reference Pearson, Wilde, Heald, Royston and Oliver2019; Tan et al., Reference Tan, Liu, Yan, Wei, Qin, Zeng and Yuan2023). Thus, it may be speculated that during earlier decades of obstetric and embryonic research, phenotypically normal SZTs of the same gender were most likely identified as MZT or DZT, which means the number of SZT cases currently existing may be far greater than two.

Figure 1.

Possible mechanism for SZT in both studies. (A) Dispermic fertilization of one ovum generates a 69/XXY triploid zygote; post-zygotic diploidization of triploids produces a 46XX/46XY chimeric embryo. (B) Parthenogenetic activation of one ovum produces two identical cells fertilized by two sperms containing different sex chromosomes.

One point to consider is that the twinning mechanism may be the same for different twinning types. Although there are diverse atypical twinning forms with huge phenotypic differences, none of them are defined as a third mechanism of twinning. In fact, almost all of the atypical twinning forms essentially belong to MZT or DZT. For example, conjoined twins are formed by incomplete splitting of the embryo when an MZT event occurs at the primitive streak stage. Mirror-image twinning has been defined as MZT with asymmetrical phenotypic features, which probably results from late embryonic splitting at days 9–12 following fertilization, just before the formation of conjoined twins. Acardiac malformation is a unique complication of monochorionic twinning, which is also known as twin reversed arterial perfusion sequence. An acardiac monster usually lacks normal upper limbs and a head. It may also have deformed lower limbs, and invariably, all body systems are deformed. This twinning status represents one of the most severe malformation events in humans. Heterokaryotypic MZT may exhibit differences and mosaicism, but it is fundamentally distinct from SZT. Heterokaryotypic MZTs arise from errors in chromosome segregation during zygotic mitosis, characterized by a genome comprising only two genetic contributions, which can result in Turner syndrome, clinically characterized by short stature and gonadal failure. Each of the four atypical twin types has its own unique mechanism, which has nothing to do with the ‘twinning mechanism’. There is no doubt that they are all essentially MZT, or special MZT. There are also many special DZT types. Heteropaternal superfecundation refers to DZT having different biological fathers, which occurs when two follicles are fertilized by sperm from two distinct males. Superfetation refers to a second conception during pregnancy, which may result in the asynchronous development of fetuses with apparent growth discordance at birth. Monochorionic DZT may form by fusion of the trophoblasts of two separate blastocysts, in which individuals always exhibit chimerism. Additionally, spontaneous conception following assisted reproductive treatment can also serve as a mechanism leading to DZT. This typically occurs when sexual intercourse takes place during the same treatment cycle in which a single embryo transfer was performed (Aston et al., Reference Aston, Peterson and Carrell2008; McNamara et al., Reference Mcnamara, Kane, Craig, Short and Umstad2016; Tan and Sepulveda, Reference Tan and Sepulveda2003; Tan, Reference Tan2023; Kyono et al., Reference Kyono, Nakajo, Nishinaka, Araki, Doshida, Toya and Kanto2009; Takehara et al., Reference Takehara, Takahashi, Hara, Matsuo, Igarashi and Kurachi2014; Rehman et al., Reference Rehman, Shariff, Arif and Memon2024). It is clear that these four atypical twin types represent DZT. Therefore, is SZT a third twinning mechanism or a special MZT just like other atypical twins? It must be determined whether the chimeric embryos are associated with the subsequent formation of twins. Therefore, the following key issues need to be investigated in SZT: (1) Is the twinning event triggered by chimerism? (2) Does the distribution of cells carrying different genomes follow some mechanism when chimeric embryos divide? These two points are almost impossible to resolve because even in classic monozygotic twins, the mechanism of embryonic division is still unclear. Furthermore, studies of sesquizygotic twinning have focused on ‘sesquizygotic’ rather than ‘twinning’. Both studies have only explored the causes of chimerism and various physiological indicators of chimeric fetuses and did not address the subsequent twinning mechanisms. ‘Dispermic fertilization’ and ‘parthenogenetic activation’ only reveal the unique mechanisms for the formation of 46XX/46/XY chimeric embryos, and embryonic splitting is not involved. Therefore, chimeric embryos and embryo splitting are two independent events. The basis for SZT is that a special embryo undergoes a twinning event to form a pair of special twins. The uniqueness of SZT resides in the formation of chimeric embryos, but the mechanism of embryonic splitting is just a traditional twinning event. Therefore, SZT is a rare type of twin with the same twinning mechanism as MZT, not a third twinning mechanism.

Is there any twinning type that may be defined as a twin mechanism different from DZT and MZT? Polar body twins (PBTs) have a significant possibility of representing a third mechanism. It was hypothesized that the formation of PBT results from the fertilization of an ovum and the first or second polar body by two separate sperms. It may produce two blastocysts within a single zona pellucida. The twinning origin of PBT is different from that of MZT and DZT. One of the maternal genetic contributions in twins is from the polar body rather than the ovum; however, there have been no confirmed cases of PBT reported thus far (Bieber et al., Reference Bieber, Nance, Morton, Brown, Redwine, Jordan and Mohanakumar1981).