Successive stamen movement in Saxifraga candelabrum is responsive to weather and pollinator visits.

BACKGROUND: Successive stamen movement is a complex plant behavior involving successive uplift of stamens and pollen release, which plays a role in reducing sexual interference, increasing pollen deposition and promoting pollen export. Although reported from several taxa, studies on whether the movement can be influenced by abiotic and biotic factors are scarce. METHODS: In this study, we here for the first time described a pattern of successive stamen movement in Saxifraga candelabrum (Saxifragaceae). We then compared the rates of stamen movement in S. candelabrum under different weather and varying pollinator visits. Pollen packaging and presentation schedule of S. candelabrum were also investigated. RESULTS: The results showed that the number of stamens bent per day in sunny days was significantly higher than overcast and rain. Flowers that receive more pollinator visits (control treatment) had significantly higher number of stamen movement than those that received fewer (removal treatment) and none (bagging treatment). Throughout the staminate phase of a flower, there was a progressive increase in both pollen quantity of individual stamens and pollen presentation during each day. CONCLUSION: Our research demonstrates that successive stamen movement in S. candelabrum was accelerated by favorable weather and increased pollinator visits, which may promote pollen export. Moreover, incremental pollen packaging is likely an adaptation to seasonal regularity in variations of sex ratio resulting from protandry.

The rise of baobab trees in Madagascar.

The baobab trees (genus Adansonia) have attracted tremendous attention because of their striking shape and distinctive relationships with fauna1. These spectacular trees have also influenced human culture, inspiring innumerable arts, folklore and traditions. Here we sequenced genomes of all eight extant baobab species and argue that Madagascar should be considered the centre of origin for the extant lineages, a key issue in their evolutionary history2,3. Integrated genomic and ecological analyses revealed the reticulate evolution of baobabs, which eventually led to the species diversity seen today. Past population dynamics of Malagasy baobabs may have been influenced by both interspecific competition and the geological history of the island, especially changes in local sea levels. We propose that further attention should be paid to the conservation status of Malagasy baobabs, especially of Adansonia suarezensis and Adansonia grandidieri, and that intensive monitoring of populations of Adansonia za is required, given its propensity for negatively impacting the critically endangered Adansonia perrieri.

Evaluation of a concise fall risk stratification among older adults with cataracts in day surgery settings: A historically controlled study.

AIM: This study aimed to evaluate the use of a concise fall risk stratification in assessing and predicting falls compared with the Morse Falls Scale among older adults with cataracts in day surgery settings. METHODS: A historically controlled study conducted from July 2020 to June 2022 was used in a municipal ophthalmic hospital in China. The concise fall risk stratification which directly graded fall risk by multifactorial judgment was used during the intervention period, while the Morse Falls Scale which graded fall risk by scale scores was used during the control period. The fall risk levels, fall assessment time, fall rates, fall-related injuries, predictive validity, and patient satisfaction with day surgery care were extracted. Propensity score matching was performed to balance baselines. RESULTS: After matching, 4132 patients were included in the final analysis. Compared with the control group, the intervention group had significantly higher assessment results for fall risk level, a significantly shorter (by 48.15%) fall assessment time, and higher patient satisfaction. There were no differences in fall rates and fall-related injuries. Compared with the Morse Falls Scale, the concise fall risk stratification had higher sensitivity and negative predictive validity, and lower specificity and positive predictive validity, while the area under curve did not differ significantly. CONCLUSION: The use of the concise fall risk stratification reduced fall assessment time, improved patient satisfaction, and is unlikely to impact falls with an overall predictive performance comparable to that of the Morse Falls Scale for older cataract adults in day surgery settings.

Defects in phospholipase C zeta cause polyspermy and low fertilization after conventional IVF: not just ICSI failure.

ABSTRACT: Phospholipase C zeta (PLCζ) is a key sperm-borne oocyte-activating factor that triggers Ca2+ oscillations and the subsequent block to polyspermy following gamete fusion. Mutations in PLCZ1, the gene encoding PLCζ, cause male infertility and intracytoplasmic sperm injection (ICSI) fertilization failure; and PLCζ expression and localization patterns are significantly correlated with ICSI fertilization rate (FR). However, in conventional in vitro fertilization (cIVF), whether and how sperm PLCζ affects fertilization remain unclear. Herein, we identified one previously reported and two novel PLCZ1 mutations associated with polyspermy in vitro that are characterized by excessive sperm-zona binding and a delay in pronuclei (PN) formation. Immunofluorescence staining and oocyte activation testing revealed that virtually all spermatozoa from patients lacked functional PLCζ and were thus unable to evoke Ca2+ oscillations. ICSI with an artificial oocyte activation treatment successfully rescued the polyspermic phenotype and resulted in a live birth. Furthermore, we analyzed PLCζ in an additional 58 males after cIVF treatment in the Reproductive and Genetic Hospital of CITIC-Xiangya (Changsha, China) between February 2019 and January 2022. We found that the proportion of spermatozoa that expressed PLCζ was positively correlated with both 2PN rate and total FR. The optimal cutoff value below which males were likely to experience low FR (total FR ≤30%) after cIVF was 56.7% for the proportion of spermatozoa expressing PLCζ. Our study expands the mutation and the phenotypic spectrum of PLCZ1 and further suggests that PLCζ constitutes a promising biomarker for identifying low FRs cases in cIVF due to sperm-related oocyte activation deficiency and that sperm PLCζ analysis may benefit the wider male population and not only men with ICSI failure.

Novel biallelic ASTL variants are associated with polyspermy and female infertility: A successful live birth following ICSI treatment.

Fertilization of the egg by the sperm is the first vital stage of embryogenesis. In mammals, only one sperm is incorporated into the oocyte. Polyspermy is a key anomaly of fertilization that is generally lethal to the embryo. To date, only a few causative genes for polyspermy have been reported. In a recent study, a homozygous variant in astacin-like metalloendopeptidase (ASTL), which encodes the ovastacin enzyme that cleaves ZP2 to prevent polyspermy, was found to be associated with female infertility characterized by polyspermy in vitro. Herein, we identified two ASTL variants in a Chinese woman likely responsible for her primary infertility and polyspermy in in vitro fertilization. Both variants were located within the key catalytic domain and predicted to alter hydrogen bonds, potentially impairing protein stability. Moreover, expression and immunoblot analyses in CHO-K1 cells indicated abnormal ovastacin zymogen activation or decreased enzyme stability. Intracytoplasmic sperm injection treatment successfully bypassed the defect in polyspermy blocking and resulted in a live birth. Our study associates ASTL variants with human infertility and further supports the contribution of this gene to blocking polyspermy in humans. Our findings expand the spectrum of ASTL mutations and should facilitate the diagnosis of oocyte-borne polyspermy.

HES1 deficiency impairs development of human intestinal mesenchyme by suppressing WNT5A expression.

Serious intestinal side-effects that target the NOTCH-HES1 pathway in human cancer differentiation therapy make it necessary to understand the pathway at the human organ level. Herein, we endogenously introduced HES1-/- mutations into human embryonic stem cells (hESCs) and differentiated them into human intestinal organoids (HIO). The HES1-/- hESCs retained ES cell properties and showed gene expression patterns similar to those of wild-type hESCs when they differentiated into definitive endoderm and hindgut. During the formation of the HES1-/- lumen we noted an impaired development of mesenchymal cells in addition to the increased differentiation of secretory epithelium. RNA-Seq revealed that inhibited development of the mesenchymal cells may have been due to a downregulation of WNT5A signaling. Overexpression of HES1 and silencing of WNT5A in the intestinal fibroblast cell line CCD-18Co indicated that HES1 was involved in the activation of WNT5A-induced fibroblast growth and migration, suggesting the likelihood of the Notch pathway in epithelial-mesenchymal crosstalk. Our results facilitated the identification of more precise underlying molecular mechanisms displaying distinct roles in HES1 signaling in stromal and epithelial development in human intestinal mucosa.

A 1-kb human CDCA8 promoter directs the spermatogonia-specific luciferase expression in adult testis.

Cell division cycle associated 8 (CDCA8) is a component of the chromosomal passenger complex and plays an essential role in mitosis, meiosis, cancer growth, and undifferentiated state of embryonic stem cells. However, its expression and role in adult tissues remain largely uncharacterized. Here, we studied the CDCA8 transcription in adult tissues by generating a transgenic mouse model, in which the luciferase was driven by a 1-kb human CDCA8 promoter. Our previous study showed that this 1-kb promoter was active enough to dictate reporter expression faithfully reflecting endogenous CDCA8 expression. Two founder mice carrying the transgene were identified. In vivo imaging and luciferase assays in tissue lysates revealed that CDCA8 promoter was highly activated and drove robust luciferase expression in testes. Subsequently, immunohistochemical and immunofluorescent staining showed that in adult transgenic testes, the expression of luciferase was restricted to a subset of spermatogonia that were located along the basement membrane and positive for the expression of GFRA1, a consensus marker for early undifferentiated spermatogonia. These findings for the first time indicate that the CDCA8 was transcriptionally activated in testis and thus may play a role in adult spermatogenesis. Moreover, the 1-kb CDCA8 promoter could be used for spermatogonia-specific gene expression in vivo and the transgenic lines constructed here could also be used for recovery of spermatogonia from adult testes.

Low-dose telomerase is required for the expansion and migration of placental mesenchymal stem cells.

Telomerase is activated in pluripotent stem cells and the majority of tumors and is postulated to be necessary for the acquisition of self-renewal and long-term proliferation. Placental mesenchymal stem cells (PMSCs) are very promising in regenerative medicine owing to their great capacity for self-renewal and differentiation potential. Although telomerase activity in the placenta is naturally low, it remains unclear whether telomerase activity is required for the properties of PMSCs. We herein isolated and identified a PMSC line carrying compound heterozygote variations in hTERT (DC-PMSCs) that lost telomerase activity, showed a typical surface phenotype of MSCs, and was able to differentiate into multiple cell lineages. DC-PMSCs showed accelerated telomere erosion, advanced senescence, and diminished migratory and invasive capabilities. RNA-seq identified 361 differentially expressed genes between DC-PMSCs and control groups, most of which were enriched in extracellular matrix, ECM, and related pathways. Knockdown of telomerase subunit genes in PMSCs confirmed the phenotype and attenuated the expression of extracellular matrix components and matrix metalloproteases. Our results suggest that low telomerase activity is not essential for the properties of MSCs, but that it is required for community maintenance and for the migration of PMSCs.

De novo transcriptome assembly using Illumina sequencing and development of EST-SSR markers in a monoecious herb Sagittaria trifolia Linn.

Background: Sagittaria trifolia Linn. is a widespread macrophyte in Asia and southeast Europe and cultivated in parts of Asia. Although a few genomic studies have been conducted for S. trifolia var. sinensis, a crop breed, there is limited genomic information on the wild species of S. trifolia. Effective microsatellite markers are also lacking. Objective: To assemble transcriptome sequence and develop effective EST-SSR markers for S. trifolia. Methods: Here we developed microsatellite markers based on tri-, tetra-, penta-, and hexa-nucleotide repeat sequences by comparatively screening multiple transcriptome sequences of eleven individuals from ten natural populations of S. trifolia. Results: A total of 107,022 unigenes were de novo assembled, with a mean length of 730 bp and an N50 length of 1,378 bp. The main repeat types were mononucleotide, trinucleotide, and dinucleotide, accounting for 55.83%, 23.51%, and 17.56% of the total repeats, respectively. A total of 86 microsatellite loci were identified with repeats of tri-, tetra-, penta-, and hexa-nucleotide. For SSR verification, 28 polymorphic loci from 41 randomly picked markers were found to produce stable and polymorphic bands, with the number of alleles per locus ranging from 2 to 11 and a mean of 5.2. The range of polymorphic information content (PIC) of each SSR locus varied from 0.25 to 0.80, with an average of 0.58. The expected heterozygosity ranged from 0.29 to 0.82, whereas the observed heterozygosity ranged from 0.25 to 0.90. Conclusion: The assembled transcriptome and annotated unigenes of S. trifolia provide a basis for future studies on gene functions, pathways, and molecular mechanisms associated with this species and other related. The newly developed EST-SSR markers could be effective in examining population genetic structure, differentiation, and parentage analyses in ecological and evolutionary studies of S. trifolia.

Evolution of complex genome architecture in gymnosperms.

Gymnosperms represent an ancient lineage that diverged from early spermatophytes during the Devonian. The long fossil records and low diversity in living species prove their complex evolutionary history, which included ancient radiations and massive extinctions. Due to their ultra-large genome size, the whole-genome assembly of gymnosperms has only generated in the past 10 years and is now being further expanded into more taxonomic representations. Here, we provide an overview of the publicly available gymnosperm genome resources and discuss their assembly quality and recent findings in large genome architectures. In particular, we describe the genomic features most related to changes affecting the whole genome. We also highlight new realizations relative to repetitive sequence dynamics, paleopolyploidy, and long introns. Based on the results of relevant genomic studies of gymnosperms, we suggest additional efforts should be made toward exploring the genomes of medium-sized (5-15 gigabases) species. Lastly, more comparative analyses among high-quality assemblies are needed to understand the genomic shifts and the early species diversification of seed plants.

Identification of a heterozygous variant of ZP2 as a novel cause of empty follicle syndrome in humans and mice.

STUDY QUESTION: Is a recurrent heterozygous mutation in ZP2, c.1925G>A (p.R642Q), associated with the Empty follicle syndrome (EFS)? SUMMARY ANSWER: ZP2, c.1925G>A (p.R642Q), led to female infertility related to EFS in humans and mice and resulted in ZP2 accumulation in the cytoplasm of oocytes. WHAT IS KNOWN ALREADY: EFS is a complex disease defined as a complete failure of oocyte retrieval after ovarian stimulation and after repeated aspirations and flushing of mature ovarian follicles. Furin-mediated cleavage is a post-translational modification (PTM) involved in various physiological processes, but the clear role of PTM mediated by furin cleavage of ZP2 protein on female fertility needs to be further explored. PTM is required for proteins to function in physiological conditions, and its perturbation has been linked to a growing number of human pathologies. Zona pellucida (ZP) proteins, which are important for oocyte development, are regulated post-translationally by well-characterized glycosylation events, as well as by furin-mediated cleavage. However, knowledge of the relevance of the consensus furin cleavage site of ZP proteins in female reproduction remains lacking. STUDY DESIGN, SIZE, DURATION: This was a basic medical research project to assess the pathogenicity of a heterozygous mutation in the ZP2 gene in EFS. PARTICIPANTS/MATERIALS, SETTING, METHODS: We studied 3 families with EFS and a control group 2213 women with proven fertility. Whole-exome sequencing detected a heterozygous mutation in the ZP2 gene in all EFS patients. The mouse strain Zp2Arg635Gln/+ (ZP2R642Q) was generated by CRISPR-Cas9-mediated genome editing. RNA-sequencing was applied to investigate transcriptional changes in the ovaries of heterozygous ZP2R642Q knock-in (KI) mice compared to WT mice. MAIN RESULTS AND THE ROLE OF CHANCE: We found a heterozygous mutation of ZP2, c.1925G>A (p.R642Q), in unrelated females with EFS, which was inherited in an autosomal-dominant manner. We used CRISPR-Cas9 to generate a mouse model encoding the orthologous variant of ZP2R642Q detected in humans, and the female ZP2R642Q KI mice recapitulated the human EFS phenotype. We further found the decreased expression of key genes involved in oocyte maturation in ZP2R642Q KI mice compared to WT mice by RNA-sequencing analysis. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: Only three families affected by EFS with the mutation were available because of its rare incidence. Although we have found different expressions of the several indispensable genes related to oocyte development between WT mice and ZP2R642Q KI mice through RNA-sequencing analysis, the specific regulatory mechanisms of the oocyte apoptosis in ZP2R642Q KI mice need to be studied further. WIDER IMPLICATIONS OF THE FINDINGS: These results are expected to open new avenues for researchers in the exploration of potential therapeutic strategies in treating EFS. STUDY FUNDING/COMPETING INTEREST(S): This project is funded by the National Key Research and Development Program of China (2018YFC1002804, 2017YFC1001500 and 2016YFC1000200). All authors declared no competing interests. TRIAL REGISTRATION NUMBER: N/A.

To roll or not to roll: nyctinastic behavior in young leaves of sacred lotus, Nelumbo nucifera.

Foliar nyctinasty refers to the daily rhythm in leaf orientation that occurs in evolutionarily diverse taxa. Traditionally, two mechanisms have been identified for the sleep movement of leaves, namely pulvinar and non-pulvinar. Here, we report upon some novel aspects of the nyctinastic behavior of leaves of the sacred lotus (Nelumbo nucifera Gaertn., Nelumbonaceae). We discovered that expanding leaves underwent daily oscillations in leaf orientation with a more vertical position at night, which is similar to many nyctinastic species. Additionally, however, the immature leaves were observed to exhibit a second type of nyctinasty that involved nocturnal rolling of leaf blades, which has not previously been reported. These two distinct mechanisms acted synergistically to make the young leaves more compact at night during a brief period (6 days) of leaf ontogenesis, during which petiole and blade showed substantial growth. We propose that the nyctinastic behavior of immature N. nucifera leaves is an adaptation that facilitates leaf growth at night by reducing mechanical interference with other leaves and plants.

Literary runaway: Increasingly more references cited per academic research article from 1980 to 2019.

References are employed in most academic research papers to give credits and to reflect scholarliness. With the upsurge in academic publications in recent decades, we are curious to know how the number of references cited per research article has changed across different disciplines over that time. The results of our study showed significant linear growth in reference density in eight disciplinary categories between 1980 and 2019 indexed in Web of Science. It appears that reference saturation is not yet in sight. Overall, the general increase in the number of publications and the advanced accessibility of the Internet and digitized documents may have promoted the growth in references in certain fields. However, the seemingly runaway tendency should be well appreciated and objectively assessed. We suggest that authors focus on their research itself rather than on political considerations during the process of writing, especially the selection of important references to cite.

The Welwitschia genome reveals a unique biology underpinning extreme longevity in deserts.

The gymnosperm Welwitschia mirabilis belongs to the ancient, enigmatic gnetophyte lineage. It is a unique desert plant with extreme longevity and two ever-elongating leaves. We present a chromosome-level assembly of its genome (6.8 Gb/1 C) together with methylome and transcriptome data to explore its astonishing biology. We also present a refined, high-quality assembly of Gnetum montanum to enhance our understanding of gnetophyte genome evolution. The Welwitschia genome has been shaped by a lineage-specific ancient, whole genome duplication (~86 million years ago) and more recently (1-2 million years) by bursts of retrotransposon activity. High levels of cytosine methylation (particularly at CHH motifs) are associated with retrotransposons, whilst long-term deamination has resulted in an exceptionally GC-poor genome. Changes in copy number and/or expression of gene families and transcription factors (e.g. R2R3MYB, SAUR) controlling cell growth, differentiation and metabolism underpin the plant's longevity and tolerance to temperature, nutrient and water stress.

Case Report: A Novel Heterozygous ZP3 Deletion Associated With Empty Follicle Syndrome and Abnormal Follicular Development.

Background: Empty follicle syndrome (EFS) is defined as the complete failure to retrieve oocytes after ovarian stimulation. Although several mutations in ZP1, ZP2, ZP3, and LHCGR have been identified as genetic causes of EFS, its pathogenesis is still not well-understood. Methods: Whole-exome sequencing (WES) was employed to identify the candidate pathogenic mutations, which were then verified by Sanger sequencing. A study in CHO-K1 cells was performed to analyze the effect of the mutation on protein expression. Additionally, immunohistochemistry (IHC) staining was used to examine follicular development and zona pellucida (ZP) assembly in the ovary of an EFS patient. Results: A novel heterozygous deletion in ZP3 (c.565_579del[p.Thr189_Gly193del]) was identified in the EFS patient. It was inherited dominantly and resulted in significant degradation of the ZP3 protein. Oocytes with degenerated cytoplasm and abnormal ZP assembly were observed in follicles up to the secondary stage, and many empty follicle-like structures were present. Conclusion: We identified a novel ZP3 mutation that expands the mutational spectrum associated with human EFS. We also showed the abnormal follicular development and ZP assembly of the EFS patient with the heterozygous ZP3 mutation, which provides new insights into the pathogenesis of EFS.

Subcutaneous tunneling technique to improve outcomes for patients undergoing chemotherapy with peripherally inserted central catheters: a randomized controlled trial.

OBJECTIVE: To compare the value of the subcutaneous tunneling technique versus the normal technique in improving the outcomes of patients undergoing chemotherapy with peripherally inserted central catheters (PICCs). METHODS: One hundred thirty patients were randomly divided into an experimental group (subcutaneous tunneling technique) and control group (normal technique) according to the PICC placement technique, and clinical data were compared between the groups. RESULTS: In total, 129 PICCs were successfully inserted. Compared with the control group, the experimental group had a lower occurrence of complications after placement (especially catheter dislodgement: 3.1% vs. 15.4%, venous thrombosis: 3.1% vs. 15.4%, and wound oozing: 14.1% vs. 27.7%), lower occurrence of unscheduled PICC removal (3.1% vs. 13.8%), greater comfort during placement (14.16 ± 2.21 vs. 15.09 ± 2.49 on a scale ranging from 6 to 30 points, with higher scores indicating lower degrees of comfort), and lower costs of PICC maintenance (median (interquartile range) per-day maintenance cost: 13.90 (10.99-32.83) vs. 15.69 (10.51-57.46) Yuan). The occurrence of complications and amount of bleeding during placement were not significantly different between the two groups. CONCLUSIONS: The subcutaneous tunneling technique can improve PICC placement by reducing complications and costs of maintenance with better patient comfort during placement.

Homozygous pathogenic variants in ACTL9 cause fertilization failure and male infertility in humans and mice.

Total fertilization failure (TFF) can occur during in vitro fertilization (IVF) treatments, even following intracytoplasmic sperm injection (ICSI). Various male or female factors could contribute to TFF. Increasing evidence suggested that genetic variations in PLCZ1, which encodes 1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase zeta-1 (PLCζ), is involved in oocyte activation and is a key male factor in TFF. In the present study, we explored the genetic variants in male individuals that led to TFF. A total of 54 couples with TFF or poor fertilization (fertilization rate < 20%) were screened, and 21 couples were determined to have a male infertility factor by the mouse oocyte activation test. Whole-exome sequencing of these 21 male individuals identified three homozygous pathogenic variants in ACTL9 (actin like 9) in three individuals. ACTL9 variations led to abnormal ultrastructure of the perinuclear theca (PT), and PLCζ was absent in the head and present in the neck of the mutant sperm, which contributed to failed normal calcium oscillations in oocytes and subsequent TFF. The key roles of ACTL9 in the PT structure and TFF after ICSI were further confirmed in an Actl9-mutated mouse model. Furthermore, assisted oocyte activation by calcium ionophore exposure successfully overcame TFF and achieved live births in a couple with an ACTL9 variant. These findings identified the role of ACTL9 in the PT structure and the correct localization of PLCζ. The results also provide a genetic marker and a therapeutic option for individuals who have undergone ICSI without successful fertilization.