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Study Question: What is the prevalence of multiple pregnancy with zygotic splitting after single embryo transfer (SET)?Summary Answer: The prevalence of multiple pregnancy with zygotic splitting after SET was 1.36%.What Is Known Already: In 2008, the Japan Society of Obstetrics and Gynaecology (JSOG) recommended the adoption of SET to reduce multiple births. Since then, to improve the clinical pregnancy rate, elective SET using blastocyst transfer and frozen-warmed ET has increased. Blastocyst culture and zona pellucida manipulation, including ICSI and AH, have been widely reported as risk factors for monozygotic twinning. However, all these studies may have included cases with dizygotic pregnancies produced by a transferred embryo and a spontaneous conception.Study Design, Size, Duration: A retrospective observational study was performed, based on 937 848 SET cycles in registered ART data from the JSOG between 2007 and 2014. The study was approved by the Registration and Research Subcommittee of the JSOG and Juntendo University Ethics Committee.Participants/materials, Setting, Methods: To identify possible factors affecting the prevalence of zygotic splitting, we identified pregnancies, in which the number of foetuses exceeded the number of gestational sacs (GSs), to restrict our analysis to 'true' zygotic splitting. Multiple logistic regression analysis was performed using singleton pregnancy after SET, as control. P < 0.05 was considered statistically significant.Main Results and the Role Of Chance: Fresh and frozen-warmed SET produced 276 934 clinical pregnancies (29.5%/SET), including 4310 twins (1.56% of pregnancies) and 109 triplets (0.04% of pregnancies). Based on sex analysis of dichorionic twins after SET, the prevalence of multiple pregnancy with zygotic splitting was 1.36%. Statistical analysis revealed that compared to singleton pregnancies zygotic splitting pregnancies were associated with frozen-warmed ET cycles (odds ratio [OR] = 1.34; 95% CI: 1.16-1.55), blastocyst culture (OR = 1.79; 95% CI: 1.54-2.09) or AH (OR = 1.21; 95% CI: 1.08-1.35). In fresh ET cycles, the prevalence rate of zygotic splitting pregnancy after single blastocyst transfer was significantly higher than that after SET cycles with cleavage embryos (OR = 2.20; 95% CI: 1.83-2.66). However, no significant difference in ovarian stimulation and fertilization methods was recognized.Limitations, Reasons For Caution: In the current Japanese ART registry system, data regarding frozen-warmed ET do not include information about ovarian stimulation and fertilization methods. Registration for AH only began in 2010. There is no way of validating if data submitted by clinics is correct.Wider Implications Of the Findings: Clinicians should consider whether to counsel couples about the small increase in the risk of zygotic splitting associated with some embryo manipulations.Study Funding/competing Interest(s): No external funds were used for the study. The authors have no competing interests to declare.Trial Registration Number: None. [ABSTRACT FROM AUTHOR]

The dose response of embryos of the zebrafish, Danio rerio , irradiated at 5 h post fertilization (hpf) by 2-MeV neutrons with ≤100 mGy was determined. The neutron irradiations were made at the Neutron exposure Accelerator System for Biological Effect Experiments (NASBEE) facility in the National Institute of Radiological Sciences (NIRS), Chiba, Japan. A total of 10 neutron doses ranging from 0.6 to 100 mGy were employed (with a gamma-ray contribution of 14% to the total dose), and the biological effects were studied through quantification of apoptosis at 25 hpf. The responses for neutron doses of 10, 20, 25, and 50 mGy approximately fitted on a straight line, while those for neutron doses of 0.6, 1 and 2.5 mGy exhibited neutron hormetic effects. As such, hormetic responses were generically developed by different kinds of ionizing radiations with different linear energy transfer (LET) values. The responses for neutron doses of 70 and 100 mGy were significantly below the lower 95% confidence band of the best-fit line, which strongly suggested the presence of gamma-ray hormesis. [ABSTRACT FROM AUTHOR]

Exotic animals can cause genetic contamination of indigenous species if they escape and reproduce in wild populations. The loach Misgurnus anguillicaudatus and species related to it have been imported into Japan for commercial use. We collected live loach specimens from the central wholesale market in Tokyo. For the 451 specimens collected, ploidy status was examined by DNA content flow cytometry, and polyploid loaches with triploid, tetraploid, and other higher ploidy ranges were detected. Hypertriploid and hypertetraploid individuals were easily detected by flow cytometry using standard eudiploid, eutriploid, and eutetraploid controls, and the reproductive capacities of these hyperpolyploid males were then examined. Sperm of hypertriploid males did not exhibit active progressive motility, and major populations of spermatozoa or spermatozoon-like cells were detected in the triploid and hexaploid ranges. Motile haploid spermatozoa were very rare in sperm from hypertriploid males. Therefore, hypertriploid males were sterile, whereas hypertetraploid males produced fertile hyperdiploid spermatozoa with active progressive motility after contact with ambient water. Viable progeny occurred in crosses between normal wild-type diploid females and hypertetraploid males, but androgenotes induced by the fertilization of UV-irradiated eggs with the sperm of hypertetraploid males were inviable hyperdiploids. Cytogenetic analyses of those androgenotes indicated that hypertetraploid males should produce hyperdiploid spermatozoa with 2n = 54 (i.e., four supernumerary microchromosomes). [ABSTRACT FROM AUTHOR]

Morphological studies were undertaken on Gastroclonium pacificum ( E. Y. Dawson) C. F. Chang et B. M. Xia ( Champiaceae, Rhodymeniales) from Japan. We describe the details of male reproductive structures, the procarp and early post-fertilization stages. This species has a solid axis, with both tetrasporangia and polysporangia, spermatangia are cut off from spermatangial parent cells, and a procarp is composed of a 4-celled carpogonial branch and two 2-celled auxiliary cell branches. The mature cystocarp lacks a conspicuous ostiole, a characteristic of the genus Gastroclonium. The most distinctive characteristic of the species is the tela arachnoidea, which is lacking in other species of Gastroclonium. [ABSTRACT FROM AUTHOR]

Abstract  The reproductive biology of the fanray, Platyrhina sinensis, was examined in Ariake Bay, Japan, from May 2002 until December 2006. Females reached sexual maturity at a larger size than males [total length (TL) at 50% sexual maturity: male, 393 mm; female, 421 mm]. The present data support a distinct annual reproductive cycle for P. sinensis. The gonadosomatic index (GSI) for mature males showed a clear seasonal trend, declining from August to November. Histological observations showed that mature sperm in the testes occurred from August to November when monthly GSI declined. Concomitantly, pre-ovulatory ova were observed in females collected from August to November. These data indicate that mating, ovulation and fertilization occurred from August to November. Near-term embryos, neonates and recent post-partum females also occurred from August to November. Additionally, all post-partum females possessed large pre-ovulatory ova. Parturition occurred from August to November followed immediately by mating, ovulation and fertilization. Mature females become pregnant every year, and the gestation period is almost 1 year. Fertilized uterine eggs without macroscopic embryonic development were present throughout the annual reproductive cycle, indicating that P. sinensis utilizes embryonic diapause as its reproductive strategy. Both reproductive tracts of females were functional, and fecundity ranged from 1 to 12 with a mean of 6.0, increasing with TL. [ABSTRACT FROM AUTHOR]

Using a natural population of masu salmon (Oncorhynchus masou (Brevoort, 1856)) in a stream of Shikaribetsu Lake, Hokkaido, Japan, we combined behavioural observations with genetic parentage analysis to explore the factors affecting fertilization success achieved by alternative mating tactics (fighting by large migratory males and sneaking by small mature male parr). Larger males gained priority access to females; migrant males do this by holding a guarding position near the nesting female and mature male parr do this by adopting sneaker behaviour and attending spawning groups. Status of mature male parr was related to success of nest entry but not to timing of nest entry, although the timing of nest entry influenced fertilization success of sneakers and ejaculation simultaneous with pair spawning was needed for fertilization by sneakers. The relative body size of each male who successfully spawned with a female is also likely to determine the proportion of eggs he fertilized because larger males have larger ejaculate. These results provide insight into factors relating to variation in fertilization success, how body size dimorphisms may be related to fitness, and evolution of alternative mating tactics. [ABSTRACT FROM AUTHOR]

This study was performed to obtain information on the occurrence of multiple paternities in three species of viviparous Japanese surfperch using allelic markers of microsatellite DNA loci. Direct evidence for multiple fertilizations was established by reconstructing paternal genotypes from the progeny of gravid females. Multiple paternities were ascertained in five of 10 broods of Ditrema temmincki and in three of nine broods of Neoditrema ransonneti, but not in Ditrema viride. The number of patrilines detected in the progeny of D. temmincki and N. ransonneti females were two or three, respectively, as determined by the GERUD v2.0 algorithm for reconstructing parental genotypes from half-sib progeny arrays. [ABSTRACT FROM AUTHOR]

The effects of silvering state of wild female Japanese eels Anguilla japonica on the success of induced maturation and the following spawning were examined. Thirty-eight females, collected in Mikawa Bay, were divided into four stages based on their silvering state: yellow (Y1), late-yellow (Y2), silver (S1) and late silver eels (S2). Despite injections of salmon pituitary extract (SPE) through the standard technique, Y1 and Y2 eels did not respond to the treatment with undeveloped gonad (gonad-somatic index [GSI]: 0.3–0.9), and all these females died by 5 weeks, probably due to an abnormal physiological condition. Most S1 (81%) and S2 eels (100%) matured completely (GSI: 17.8–51.4), and finally spawned successfully (69% for S1, 89% for S2). S2 eels fully matured with oocytes of over 750 μm in diameter by significantly smaller number of injections of SPE (5–6 times) than the case of S1 eels (6–8 times). The amount of eggs released by S2 eels (0.65 ± 0.11 g/fish per body weight [BW]) was significantly larger than those by S1 eels (0.54 ± 0.09 g/fish per BW). There was no difference in fertilization and hatching rates between eggs released by S1 eels and those of S2 eels. These results indicate that the success of induced maturation and spawning in wild female Japanese eels depends on their silvering state, and matured eggs can be obtained efficiently through the use of S2 eels rather than other stages. [ABSTRACT FROM AUTHOR]

To improve stock enhancement strategies, the influences of repeated cross-breeding on several traits were studied in hatchery-reared juveniles of the Japanese flounder. Ten families (seven caught in the wild and three from cross-bred brood fish) were produced by artificial fertilization. Ninety-four days after hatching, we investigated settlement, burying in the sand and escape behavioral traits, and then measured tolerance of high salinity and six morphological traits. Juveniles from the repeatedly cross-bred brood fish showed significantly higher growth and tolerance than those from the wild-caught brood fish. Among the behavioral traits, a significantly lower settling speed and incidence of burying in the sand were observed in the juveniles from the repeatedly cross-bred families. Heritability ( h2) of growth traits was more than 0.2 in both origins, while that of other traits was close to zero. These results suggest that repeated cross-breeding of Japanese flounder under hatchery conditions improves growth performance and leads to the loss of certain behavioral traits that are considered essential to survival in wild environments. [ABSTRACT FROM AUTHOR]

Ophiodaphne formata is a conspicuously dimorphic ophiuroid; the disk diameters are approximately 1 mm for males and 5 mm for females. The dwarf male clings to the larger female, with the oral surfaces and bursae of the paired ophiuroids closely appressed. Moreover, the female of each pair adheres aborally to the oral surface of a host sand dollar, Astriclypeus manni. Spawning and external fertilization occur in August, at Tsuruga Bay, Sea of Japan. Development of the dimorphic brittle star O. formata is described for the first time, from spawning through metamorphosis, with special attention to the formation of the skeletal system and the external morphology of early juveniles. Fertilized eggs are about 90 µm in diameter, pale pink, and negatively buoyant. The embryos undergo equal, total, and radial cleavage, and the larval skeleton first forms as a pair of tetraradiate spicules. Larval development proceeds to an 8-armed planktotrophic ophiopluteus, with skeletal elements that consist of a body rod and two recurrent rods. Three weeks after fertilization, all the pluteal arms, except for the postero-lateral arms, are absorbed, and the metamorphosing larvae sink to the bottom. Metamorphosis is completed 21.5 days after fertilization, and the resulting juvenile is pentagonal and approximately 270 µm in diameter. The smallest specimen (480 µm in disk diameter) collected by field sampling exhibited male features on the skeletal plates of the jaw and disk. Sexual dimorphism, the peculiar pairing behavior, and the close relationship with the host sand dollar may have evolved as distinct reproductive characteristics in this ophiuroid with its typical ophiopluteus larvae. [ABSTRACT FROM AUTHOR]

The article reports on a study of the breeding of hermatypic coral Galaxea fascicularis and their partial reproductive isolation between colonies of different nematocyst types and enhancement of fertilization success through parental colonies in Okinawa, Japan. It mixed eggs of hermatypic corals with sperm at an appropriate dilution after successful fertilization. It made 12 standard crosses using gametes from 13 females and nine males wherein very low fertilization were observed in two of the crosses. It also founds out that fertilization experiments of Galaxea fascicularis could be performed in the presence of colony fragments.

The article focuses on the effect of low salinity on the existence and development of squid Todarodes pacificus (T. pacificus) hatchling. This ommastrephid squid is short-lived whose reproductive and recruitment success depend largely on the physical and biological environments at the spawning grounds. Temperature affects the reproductive success of T. pacificus. A recent study reported that the low salinity water distributes in Tsushima Strait. It reports on having fertilized eggs by artificial fertilization. Live hatchlings were classified as either normal of deformed. Survival rates 108 hours after the start of treatments were high (>90%) in the three highest salinities, but dropped to 77.5%. It shows that the peak of low salinity water distribution occurs in summer.

Abstract: Tetraploid loaches have been discovered amongst specimens recovered from fish markets in Japan. These tetraploids can be used as a source of diploid gametes to assist the further expansion of ploidy manipulation. Here, we produced a first generation neo-tetraploid strain by fertilizing eggs from a normal diploid female with diploid spermatozoa from a natural tetraploid male, followed 5min later by heat shock (42°C, 2min duration) to inhibit release of the second polar body. Diploid spermatozoa from the neo-tetraploid males produced were then used to create androgenetic diploid progeny by fertilizing UV-irradiated eggs from a normal diploid female. Triploid progeny were produced by crossing a normal diploid female with a neo-tetraploid male. Tetraploid progeny were produced by cold shock (1°C, 40min duration), beginning 5min after fertilizing normal eggs with diploid spermatozoa from first generation neo-tetraploid males. Reproductive performance of second generation progeny was also examined. Androgenetic diploid males generated fertile haploid spermatozoa. Triploid males were sterile, but a triploid female laid fertile haploid eggs. Second generation neo-tetraploid males were considered sterile. [ABSTRACT FROM AUTHOR]