BACKGROUND: Time-lapse imaging systems for embryo incubation and selection might improve outcomes of in-vitro fertilisation (IVF) and intracytoplasmic sperm injection (ICSI) treatment due to undisturbed embryo culture conditions, improved embryo selection, or both. However, the benefit remains uncertain. We aimed to evaluate the effectiveness of time-lapse imaging systems providing undisturbed culture and embryo selection, and time-lapse imaging systems providing only undisturbed culture, and compared each with standard care without time-lapse imaging.
METHODS: We conducted a multicentre, three-parallel-group, double-blind, randomised controlled trial in participants undergoing IVF or ICSI at seven IVF centres in the UK and Hong Kong. Embryologists randomly assigned participants using a web-based system, stratified by clinic in a 1:1:1 ratio to the time-lapse imaging system for undisturbed culture and embryo selection (time-lapse imaging group), time-lapse imaging system for undisturbed culture alone (undisturbed culture group), and standard care without time-lapse imaging (control group). Women were required to be aged 18-42 years and men (ie, their partners) 18 years or older. Couples had to be receiving their first, second, or third IVF or ICSI treatment and could not participate if using donor gametes. Participants and trial staff were masked to group assignment, embryologists were not. The primary outcome was live birth. We performed analyses using the intention-to-treat principle and reported the main analysis in participants with primary outcome data available (full analysis set). The trial is registered on the International Trials Registry (ISRCTN17792989) and is now closed.
RESULTS: 1575 participants were randomly assigned to treatment groups (525 participants per group) between June 21, 2018, and Sept 30, 2022. The live birth rates were 33·7% (175/520) in the time-lapse imaging group, 36·6% (189/516) in the undisturbed culture group, and 33·0% (172/522) in the standard care group. The adjusted odds ratio was 1·04 (97·5% CI 0·73 to 1·47) for time-lapse imaging arm versus control and 1·20 (0·85 to 1·70) for undisturbed culture versus control. The risk reduction for the absolute difference was 0·7 percentage points (97·5% CI -5·85 to 7·25) between the time-lapse imaging and standard care groups and 3·6 percentage points (-3·02 to 10·22) between the undisturbed culture and standard care groups. 79 serious adverse events unrelated to the trial were reported (n=28 in time-lapse imaging, n=27 in undisturbed culture, and n=24 in standard care).
CONCLUSIONS: In women undergoing IVF or ICSI treatment, the use of time-lapse imaging systems for embryo culture and selection does not significantly increase the odds of live birth compared with standard care without time-lapse imaging.
BACKGROUND: Barts Charity, Pharmasure Pharmaceuticals, Hong Kong OG Trust Fund, Hong Kong Health and Medical Research Fund, Hong Kong Matching Fund.

BACKGROUND: The occurrence of blastocyst collapse may become an indicator of preimplantation embryo quality assessment. It has been reported that collapsing blastocysts can lead to higher rates of aneuploidy and poorer clinical outcomes, but more large-scale studies are needed to explore this relationship. This study explored the characteristics of blastocyst collapse identified and quantified by artificial intelligence and explored the associations between blastocyst collapse and embryo ploidy, morphological quality, and clinical outcomes.
METHODS: This observational study included data from 3288 biopsied blastocysts in 1071 time-lapse preimplantation genetic testing cycles performed between January 2019 and February 2023 at a single academic fertility center. All transferred blastocysts are euploid blastocysts. The artificial intelligence recognized blastocyst collapse in time-lapse microscopy videos and then registered the collapsing times, and the start time, the recovery duration, the shrinkage percentage of each collapse. The effects of blastocyst collapse and embryo ploidy, pregnancy, live birth, miscarriage, and embryo quality were studied using available data from 1196 euploid embryos and 1300 aneuploid embryos.
RESULTS: 5.6% of blastocysts collapsed at least once only before the full blastocyst formation (tB), 19.4% collapsed at least once only after tB, and 3.1% collapsed both before and after tB. Multiple collapses of blastocysts after tB (times ≥ 2) are associated with higher aneuploid rates (54.6%, P > 0.05; 70.5%, P < 0.001; 72.5%, P = 0.004; and 71.4%, P = 0.049 in blastocysts collapsed 1, 2, 3 or ≥ 4 times), which remained significant after adjustment for confounders (OR = 2.597, 95% CI 1.464-4.607, P = 0.001). Analysis of the aneuploid embryos showed a higher ratio of collapses and multiple collapses after tB in monosomies and embryos with subchromosomal deletion of segmental nature (P < 0.001). Blastocyst collapse was associated with delayed embryonic development and declined blastocyst quality. There is no significant difference in pregnancy and live birth rates between collapsing and non-collapsing blastocysts.
CONCLUSIONS: Blastocyst collapse is common during blastocyst development. This study underlined that multiple blastocyst collapses after tB may be an independent risk factor for aneuploidy which should be taken into account by clinicians and embryologists when selecting blastocysts for transfer.

OBJECTIVE: Before blastocyst development, embryos undergo morphological and metabolic changes crucial for their subsequent growth. This study aimed to investigate the relationship between morula compaction and blastocyst formation and the subsequent chromosomal status of the embryos.
METHODS: This retrospective cohort study evaluated embryo development (n = 371) using time-lapse imaging; 94 blastocysts underwent preimplantation genetic testing for aneuploidy (PGT-A).
RESULTS: The embryos were classified as fully (Group 1, n = 194) or partially (Group 2, n = 177) compacted. Group 1 had significantly higher proportions of good- and average-quality blastocysts than Group 2 (21.6% vs. 3.4%, p = 0.001; 47.9% vs. 26.6%, p = 0.001, respectively). The time from the morula stage to the beginning and completion of compaction and blastocyst formation was significantly shorter in Group 1 than in Group 2 (78.6 vs. 82.4 h, p = 0.001; 87.0 vs. 92.2 h, p = 0.001; 100.2 vs. 103.7 h, p = 0.017, respectively). Group 1 embryos had larger surface areas than Group 2 embryos at various time points following blastocyst formation. Group 1 blastocysts had significantly higher average expansion rates than Group 2 blastocysts (653.6 vs. 499.2 μm2/h, p = 0.001). PGT-A revealed a higher proportion of euploid embryos in Group 1 than in Group 2 (47.2% vs. 36.6%, p = 0.303).
CONCLUSIONS: Time-lapse microscopy uncovered a positive relationship between compaction and blastocyst quality and its association with embryo ploidy. Hence, compaction evaluation should be prioritized before blastocyst selection for transfer or cryopreservation.

The cariogenicity of Streptococcus mutans relates to its ability to form biofilms on dental surfaces. The aim of this work was to develop a flowcell system compatible with time-lapse confocal microscopy to compare the adhesion and accumulation of S. mutans cells on surfaces in unsupplemented media against media containing sucrose or sucralose (a non-metabolized sweetener) over a short period of time. Fluorescent S. mutans 3209/pVMCherry was suspended in unsupplemented media or media supplemented with 1% sucrose or 1% sucralose and passed through a 3D-printed flowcell system. Flowcells were imaged over 60 minutes using a confocal microscope. Image analysis was performed, including a newly developed object-movement-based method to measure biomass adhesion. Streptococcus mutans 3209/pVMCherry grown in 1% sucrose-supplemented media formed small, dense, relatively immobile clumps in the flowcell system measured by biovolume, surface area, and median object centroid movement. Sucralose-supplemented and un-supplemented media yielded large, loose, mobile aggregates. Architectural metrics and per-object movement were significantly different (P < 0.05) when comparing sucrose-supplemented media to either unsupplemented or sucralose-supplemented media. These results demonstrate the utility of a flowcell system compatible with time-lapse confocal microscopy and image analysis when studying initial biofilm formation and adhesion under different nutritional conditions.

Delays in influenza vaccine delivery and administration can hinder vaccine coverage and protection. This study examines the differentials in distributing and administering adjuvanted trivalent (aTIV) and quadrivalent influenza vaccines (aQIV) to older adults in Italy\'s primary care setting and its potential impact on hospitalization risk over 5 epidemic seasons.
Using a primary care database, individuals aged ≥ 65 years were selected. The proportion of vaccine distribution to regional authorities and subsequent administration by GPs was estimated using census data. Using quantile (median) regression, we examined the relationship between velocities of vaccine distribution and administration (doses/week) and the incidence of hospitalizations.
Over the 5 influenza seasons, the velocity of distribution and administration of aTIV/aQIV ranged 341-833 and 152-270 median doses/week; no trend was yielded for the difference between these velocities (p = 0.189) or vaccine coverage (p = 0.142). An association was observed for each differential dose/week between distributed and administered vaccines and all-cause hospitalizations with a 10% increase in 2017-2018, 54% in 2018-2019, and 12% in 2020-2021 season.
These findings highlight the importance of minimizing the time lapse between vaccine distribution and administration to mitigate the impact of influenza and address factors that contribute to vaccination barriers.

The Association for the Study of Reproductive Biology (ASEBIR) Interest Group in Embryology (in Spanish \'Grupo de Interés de Embriología\') reviewed key morphokinetic parameters to assess the contribution of time-lapse technology (TLT) to the ASEBIR grading system. Embryo grading based on morphological characteristics is the most widely used method in human assisted reproduction laboratories. The introduction and implementation of TLT has provided a large amount of information that can be used as a complementary tool for morphological embryo evaluation and selection. As part of IVF treatments, embryologists grade embryos to decide which embryos to transfer or freeze. At the present, the embryo grading system developed by ASEBIR does not consider dynamic events observed through TLT. Laboratories that are using TLT consider those parameters as complementary data for embryo selection. The aim of this review was to evaluate review time-specific morphological changes during embryo development that are not included in the ASEBIR scoring system, and to consider them as candidates to add to the scoring system.

Does short-interval second ejaculation improve sperm quality, embryo development and clinical outcomes for oligoasthenozoospermia males received intracytoplasmic sperm injection (ICSI) treatment?
All enrolled male patients underwent short-interval secondary ejaculation on the day of oocyte retrieval, and 786 sibling MII oocytes from 67 cycles were equally divided into two groups based on whether the injected spermatozoons originated from the first or second ejaculation. Semen parameters, embryo development efficiency, morphokinetic parameters and clinical outcomes were compared between the two groups to assess the efficiency and clinical value of short-interval second ejaculation in ICSI cycles.
Short-interval second ejaculation significantly improved sperm motility, normal morphological rate, and sperm DNA integrity both before and after sperm swim-up. The high-quality blastocyst rate (24.79% versus 14.67%), available blastocyst rate (57.56% versus 48.44%), and oocyte utilization rate (52.93% versus 45.29%) were significantly higher in the second ejaculation group (P<0.05). The clinical pregnancy rate (59.09% versus 47.37%), implantation rate (42.11% versus 32.35%) and live birth rate (40.91% versus 31.58%) were higher in the second ejaculation group, but the differences were not significant (P>0.05). Time-lapse analysis showed that morphokinetic time points after the 7-cell stage were earlier in the second ejaculation group but without a significant difference (P>0.05), and abnormal embryo cleavage patterns between the two groups were not significantly different (P>0.05).
Short-interval second ejaculation significantly improves sperm quality in oligoasthenozoospermic males, and is beneficial for blastocyst formation efficiency in ICSI cycles. This study suggested a non-invasive and simple but effective strategy for improving ICSI treatment outcomes.

Cell culturing methods are increasingly used to reduce and replace the use of live animals in biomedical research and chemical toxicity testing. Although live animals are avoided when using cell culturing methods, they often contain animal-derived components of which one of the most commonly used is foetal bovine serum (FBS). FBS is added to cell culture media among other supplements to support cell attachment/spreading and cell proliferation. The safety, batch-to-batch variation, and ethical problems with FBS are acknowledged and therefore world-wide efforts are ongoing to produce FBS free media. Here, we present the composition of a new defined medium with only human proteins either recombinant or derived from human tissues. This defined medium supports long-term culturing/routine culturing of normal cells and of cancer cells, and can be used for freezing and thawing of cells, i.e. for cell banking. Here, we show for our defined medium, growth curves and dose response curves of cells grown in two and three dimensions, and applications such as cell migration. Cell morphology was studied in real time by phase contrast and phase holographic microscopy time-lapse imaging. The cell lines used are human cancer-associated fibroblasts, keratinocytes, breast cancer JIMT-1 and MDA-MB-231 cells, colon cancer CaCo-2 cells, and pancreatic cancer MiaPaCa-2 cells as well as the mouse L929 cell line. In conclusion, we present the composition of a defined medium without animal-derived products which can be used for routine culturing and in experimental settings for normal cells and for cancer cells, i.e. our defined medium provides a leap towards a universal animal product free cell culture medium.

Does complete resection of endometriosis improve embryo quality as assessed by morphokinetic parameters using time-lapse microscopy?
For this retrospective study we analysed 237 fertilised, cultured and transferred embryos from 128 fresh IVF and/ or ICSI transfer cycles. Endometriosis was confirmed or excluded by laparoscopy. Patients were stimulated with recombinant FSH using GnRH agonist and antagonist protocols. After fertilisation, a time-lapse incubation system was used for observation. Embryo quality was assessed using the KIDScore™ D3 and D5 implantation data algorithm.
The analysis showed a median KIDScore™ D5 of 2.6 (on a scale of 1 to 9.9) for embryos from patients with endometriosis without complete resection. The control group without endometriosis achieved a score of 6.8 (p = 0.003). The median score for embryos from endometriosis patients with complete resection was 7.2, which was a significant increase compared to embryos from patients without complete resection (p = 0.002). We observed an effect size of r = 0.4 for complete resection versus no resection of endometriosis using the KIDScore™ D5. There were no differences in KIDScore™ D3 between the three patient groups. Pregnancy and miscarriage rates showed the same clinical trends. In three of our four case series of patients who underwent IVF/ ICSI cycles before and after complete resection, we found a marked improvement in embryo quality after complete resection.
Complete resection of endometriosis could significantly improve the otherwise poor embryo quality of patients undergoing IVF-procedures. The data, therefore, strongly support recommending surgery to patients with endometriosis prior to assisted reproduction.

OBJECTIVE: To investigate whether high relative humidity conditions (HC), when using a time-lapse system (TLS) with sequential culture media, are beneficial to embryo culture, improving ongoing pregnancy rates.
METHODS: We included patients undergoing their first ICSI cycle treatment from April 2021 to May 2022. Patients assigned to dry conditions (DC) or HC were 278 and 218, respectively. We used a GERI TLS, three chambers configured in humidity conditions and three in dry conditions. The effect of HC on ongoing pregnancy rate was assessed by the propensity matched sample, to reduce potential differences between women undergoing either HC or DC and reduce biased estimation of treatment effect.
RESULTS: After adjusting for several confounding variables and applying the propensity score (PS), no significant differences were observed in the rates of normal (2PN) and abnormal (1PN and 3PN) fertilization, blastulation, top-quality blastocysts, frozen blastocysts, ongoing pregnancies, and miscarriages. The 2-cell (t2) and 4-cell (t4) stages and cell divisions between such stages occurred earlier and were more synchronous in the in DC.
CONCLUSIONS: These results suggest that HC conditions do not improve the rate of ongoing pregnancy and several embryological outcomes, under the conditions used in this study based on a time-lapse system and sequential culture with day 3 medium change-over.