| Background and objective:Infertility is one of the diseases of the reproductive system.The World Health Organization(WHO)defines it as a person who has not successfully conceived after a normal unprotected sex life for more than 1 year.With the development of human society,the incidence of infertility caused by environmental,physical,psychological and other factors is increasing year by year,among which fallopian tube factors is the main cause.Although assisted reproductive technology has been widely used to improve the clinical pregnancy rate of infertility patients,there are still many unexplained reasons that make the clinical pregnancy rate unsatisfactory.Therefore,how to improve the clinical pregnancy rate of assisted reproduction is a hot research topic in the field of reproductive medicine.The quality of oocytes and the maternal micro-ecological environment are essential for a successful pregnancy,and the vaginal microbiota plays an important role in maintaining the health of the female reproductive tract.In recent years,there have been many studies on the relationship between vaginal microbiota and infertility,premature delivery,recurrent miscarriage and other diseases at home and abroad.However,there has been little work exploring the potential connection between the vaginal microbiota and FET.Therefore,this study included a total of 555 infertility patients who received FET.By comparing the age,endometrial preparation programs,hormone levels during endometrial transformation(luteinizing hormone,estrogen,progesterone),endometrial thickness before transplantation,embryo number and other clinical indicators of patients with different pregnancy outcomes,and use highthroughput sequencing technology to compare the vaginal microbiota of patients with successful and failed FET,explore the correlation between various factors and the outcome of FET,to further identify the potential harm of reproductive tract diseases,and to provide guidance and support for assisted reproductive technology in the diagnosis and treatment of infertility.Methods:(1)Collect 555 infertility patients who received FET infertility treatment at the Reproductive Assistance Center of Jiujiang Maternal and Child Health Hospital in Jiangxi Province from June 2017 to December 2018.According to the inclusion and exclusion criteria of this experiment,a total of 475 patients were collected Included in this study.According to the pregnancy outcome of FET fertility treatment,it was divided into two groups,namely pregnancy group(P group),with a total of 238 cases,and none pregnancy group(NP group),with a total of 237 cases.(2)Collect clinical data of the enrolled patients,including age,endometrial preparation programs,hormone levels during endometrial transformation(luteinizing hormone,estrogen,progesterone),endometrial thickness before transplantation,embryo number.The SPSS17.0 software and Graph Pad8.0 software were used for statistical analysis to explore the correlation between various factors and the outcome of FET.(3)Fifty-eight patients with tubal infertility were randomly selected to collect the vaginal secretions on the day of endometrial transformation and put them in a refrigerator at 80℃ for later use.According to pregnancy outcome,it is divided into pregnancy group(P group,N=37)and non-pregnant group(NP group,N=21).The total bacterial DNA was extracted separately,and the V3-V4 regions of the bacterial16 S rRNA gene were amplified by PCR,combined with high-throughput sequencing technology to analyze the difference in vaginal microbial composition between the two groups of patients and its correlation with FET.Results:(1)The characteristics of the clinical data between the P group and the NP group showed that age and endometrial thickness before transplantation had a significant impact on the outcome of FET(P<0.05).However,there was no statistical difference in the endometrial preparation programs,the hormone levels during endometrial transformation(luteinizing hormone,estrogen,progesterone)and embryo number.(P>0.05).(2)Alpha diversity and Beta diversity analysis showed that the Shannon index(P=0.01)and Simpson index(P=0.02)of the NP group were significantly higher than those of the P group.The Venn diagram shows that there are 452 OUTs and 465 OUTs in the P group and NP group respectively,of which there are 423 OUTs in common,accounting for 93.58%(423/452)and 90.97%(423/465)of the P and NP groups,respectively.In addition,the Partial Least Squares Discriminant Analysis(PLS-DA)showed that most of the samples in the P group were far away from the NP group suggesting that the microbial community structure between the two groups is quite different.(3)Differences in vaginal microbial communities at the phylum level: there were significant differences in Actinobacteria,Fusobacteria,and Bacteroidetes between the P and NP groups at the phylum level(P<0.05),while the differences between Firmicutes and Proteobacteria were not statistically significant(P>0.05).(4)Differences in vaginal microbial communities at the genus level: there were significant differences between the P group and the NP group at the genus level in Gardnerella,Atopobium,Sneathia,and Prevotella(P<0.05),while the difference in Lactobacillus was not statistically significant(P>0.05).(5)Analyze the differences in the metabolic pathways of the functional genes of the microbial community based on the KEGG database: there are differences in the pathways of gene replication and repair,cell growth and death,lipid metabolism and enzyme metabolism between the P and NP groups,but they are not cell movement and carbohydrate metabolism have no obvious effect.Conclusion:(1)Age and endometrial thickness before transplantation are important factors that affect the outcome of FET,while the endometrial preparation programs,the hormone levels during endometrial transformation(luteinizing hormone,estrogen,progesterone)and embryo number have no significant effect on the outcome of FET.(2)High-throughput sequencing revealed a close relationship between the vaginal microbiota and the outcome of FET.The vaginal microbiota of non-pregnant patients is characterized by reduced probiotics and excessive growth of pathogens,which may be potential indicators for a higher rate of in vitro fertilization failure. |
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