Introduction

In the context of the evolving COVID-19 pandemic and recurrent outbreaks, it has been estimated that more than 10% of individuals may experience potential long-term effects of the virus1,2. The World Health Organization (WHO) defines long-term COVID-19 as the occurrence of symptoms within three months after initial infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which persist for a minimum of two months3. Numerous studies have shown that long-term COVID-19 can impact multiple organ systems in the human body4. However, the underlying pathophysiology remains unclear and could be associated with immune dysregulation, autoimmunity, persistent virus presence, and microvascular dysfunction5. COVID-19 poses a heightened risk for pregnant individuals, with the active phase of the infection potentially increasing the chances of various maternal and fetal complications, including preeclampsia, intrauterine growth restriction, preterm birth, and stillbirth6,7,8. However, the existing scientific literature on the potential negative consequences of Long-COVID-19 on pregnant women and their unborn babies remains insufficient. Therefore, the main objective of this research is to1 examine the effect of long-term COVID-19 on blood clotting functionality and maternal and fetal complications.

Through extensive investigation into SARS-CoV-2 during the COVID-19 outbreak, researchers have discovered that the primary mechanism by which SARS-CoV-2 causes harm is through its ability to infect and damage type II alveolar epithelial cells. SARS-CoV-2 accomplishes this process by attaching itself to the angiotensin-converting enzyme 2 (ACE2) receptor, resulting in the development of respiratory symptoms9.

Severe COVID-19 patients often exhibit coagulopathy and thrombosis, which are closely associated with the risk of death. Autopsy findings have consistently revealed significant thrombosis in the lungs of individuals with COVID-19, accompanied by microvascular lesions such as endothelial damage and vascular growth10,11,12. Mild COVID-19 cases typically present a hypercoagulable state of blood10. Although research suggests that pregnant individuals infected with COVID-19 do not encounter a substantial hypercoagulable condition concerning to their coagulation functionality, it is crucial to acknowledge that those experiencing long-term COVID-19 may endure enduring endothelial impairment, persistent hypoxia, and a continuing inflammatory reaction. These factors can lead to long-term activation of coagulation, microvascular damage, and persistent thrombosis in damaged blood vessels13,14,15. Therefore, it is worth investigating whether maternal Long-COVID-19 during pregnancy can also have adverse effects on maternal coagulation function.

Therefore, we conducted a retrospective cohort study to investigate the influence of maternal coagulation function and maternal and fetal complications among pregnant women with acute COVID-19, pregnant women with long-term COVID-19, and pregnant women without COVID-19. This article aimed to raise people's attention to the impact of long-term COVID-19 on maternal and fetal during pregnancy, and provide important guidance value for pregnancy counseling, clinical treatment and basic research.

Method

Study approval and registration

In order to conduct this study, a retrospective cohort investigation was carried out at the Department of Obstetrics within the First People’s Hospital of Kunming. This particular hospital, categorized as a tertiary healthcare facility in Yunnan Province, China, handles a significant number of deliveries each year, surpassing 4,000 cases. Adhering to the ethical guidelines outlined in the Declaration of Helsinki and in compliance with the relevant laws of China, the study was executed. Furthermore, ethical approval (No. YLS2023-72) was obtained from the Ethics Committee of the First People’s Hospital of Kunming in October 2023. All patients were informed about the study of their biological samples and signed written informed consent to participate.

For participants below the age of 16 informed consent was obtained from the parents of all the participants. For participants below the age of 18 informed consent was obtained from all the participants.

We committed to being able to determine information about individual participants either during data collection or after the visit.

Inclusion and exclusion criteria

A total of 3198 pregnant women who delivered in the Department of Obstetrics of Kunming First People's Hospital from November 1, 2022 to July 31, 2023 were included in this study. Among them, 623 subjects met the inclusion and exclusion criteria and completed blood sample collection and follow-up.

Criteria for inclusion: Singleton pregnant women, were routinely tested for SARS-CoV-2 by RT-PCR using nasopharyngeal swabs 72 h before delivery and during the first trimester; Serologic studies were performed within 72 h before delivery.

Exclusion criteria were: Human immunodeficiency virus infection, Toxoplasma gondii infection during pregnancy, rubella, cytomegalovirus, or syphilis. Patients with severe chronic comorbidities including chronic hypertension, type 2 and type 1 diabetes mellitus, gestational thrombocytopenic purpura, thrombophilia, history of thromboembolism, use of low molecular weight heparin and aspirin during pregnancy, and patients lost to follow-up by telephone were excluded.

Grouping for this study

Based on the different stages of SARV-CoV-2 infection, we divided the participants into three different groups (Fig. 1):

Figure 1
figure 1

Retrospective study outline for maternal.

Full size image

Croup A (Acute group): consists of individuals who were diagnosed with acute SARS-CoV-2 infection within 72 h prior to their delivery.

Croup L (The long-term COVID-19 group): includes pregnant individuals who contracted their initial SARS-CoV-2 infection during the first trimester (specifically, within the first 12 weeks of gestation). Additionally, these individuals subsequently received a diagnosis of Long-COVID-19. Importantly, they tested negative for SARS-CoV-2 through PCR within a 72-h timeframe before giving birth.

Croup N (Never group): encompassing control individuals who have never experienced an infection with SARS-CoV-2(In Yunnan Province, China, the prevention and control of COVID-19 began to be completely abandoned from January 1, 2023. Kunming First People's Hospital prohibited pregnant women with COVID-19 from visiting and giving birth before December 1, 2022. Therefore, for the patients in group N who had never been infected with COVID-19, we selected pregnant women who delivered in our hospital from November 1, 2022 to November 30, 2022 to ensure that there was no confusion about the cases with previous infections with COVID-19).

Enrollment procedure for pregnant women with long-term COVID-19

①In accordance with the diagnostic criteria for long-term COVID-19 established by the World Health Organization3. ② During early pregnancy, the subjects were infected with SARS-CoV-2 and did not experience reinfection within three days before delivery. ③We conducted telephone follow-up to gather information on symptoms experienced during the first infection of SARS-CoV-2, including a description of symptom persistence and duration. We included symptoms lasting more than 5 months after the initial infection in the statistical analysis, adhering to the defined criteria. ④Pregnant women with persistent symptoms caused by other diseases were excluded through consultation of the electronic medical record system. ⑤To ensure data accuracy in telephone follow-up, we implemented a dual-response approach where two individuals simultaneously answered calls from each subject, supervised together, and recorded data. In case of discrepancies between their responses, a third person would conduct additional follow-up calls to resolve any differences through consensus.

Demographic and anthropometric characteristics, and adverse pregnancy outcomes

We recorded all subjects: Age, body mass index (BMI) before delivery, number of pregnancies, pre-pregnancy complications (such as chronic hypertension, diabetes, polycystic ovary syndrome, etc.), family history of hypertension and diabetes, blood pressure threshold in early pregnancy, blood glucose ≥ 5.6mmol/L in early pregnancy, urinary protein in early pregnancy, gestational week of first SARS-CoV-2 infection, gestational diabetes mellitus(GDM), Hypertension in Pregnancy(HIP), and cesarean section Intrauterine delivery, postpartum hemorrhage(PH), neonatal asphyxia, neonatal transfer to neonatal intensive care unit(NICU), premature delivery(PD), fetal growth restriction(FGR), premature rupture of membranes(PROM), stillbirth.

Diagnostic criteria

①. Family history of hypertension and diabetes: parents and siblings with hypertension or diabetes16.

②. HIP, a condition characterized by high blood pressure occurring after the 20th week of pregnancy, was considered when systolic blood pressure reached or exceeded 140 mmHg or diastolic blood pressure reached or exceeded 90%. This condition usually resolved within 12 weeks after childbirth and was challenging to diagnose until the postpartum period16.

③. GDM, also known as pregnancy-related diabetes, pertains to expectant mothers who have not previously been diagnosed with pregestational diabetes mellitus (PGDM)16.

④. Definition of PH: postpartum hemorrhage, also known as excessive bleeding after childbirth, is recognized as a critical complication. It is characterized by the discharge of blood equal to or exceeding 500 mL after a vaginal delivery and 1,000 mL after a cesarean delivery within a 24-h period following the birth of the baby16.

⑤. Neonatal asphyxia is defined as the inability of newborns to breathe Line normally after birth, which is one of the main causes of neonatal death and mental retardation. Apgar score ≤ 7 is diagnosed as neonatal asphyxia16.

⑥. Neonatal transfer to NICU: neonatal transfer to NICU within 24 h after birth16.

⑦. PD: babies born at 28 but less than 37 weeks of gestation are called premature16.

⑧. FGR refers to the condition wherein the estimated fetal weight or abdominal circumference, as determined by ultrasound, falls below the 10th percentile for the corresponding gestational age16.

⑨. PROM: rupture of membranes occurs before labor16.

⑩. Stillbirth, also known as fetal demise, occurs when a fetus weighing 350 g or more and at a gestational age of 20 weeks or beyond is delivered without any noticeable signs of breathing, heartbeat, umbilical cord pulsation, or voluntary muscle movement16.

Biochemical indicators

All participants had blood samples obtained within 72 h before delivery for various laboratory tests, including platelet count(PLT), coagulation function including fibrinogen(FIB), prothrombin time(PT), and activated partial thromboplatin time(APTT)), and SARS-CoV-2 RT-PCR assays. In addition, women in group L were included to undergo RT-PCR testing for SARS-CoV-2 in the first trimester. SARS-CoV-2 nucleic acid test kit information: 2019-nCov Fluorescent PCR, Mike Biotech Co., Ltd. Registration No. : 20,203,400,184.

Content of the study

Revised sentence: Statistical techniques were utilized to compare the correlation between coagulation function (PLT, FIB, APTT, PT) and maternal and fetal complications (HIP, GDM, cesarean delivery, HP, PD, PROM, neonatal oxygen, Neonatal transfer to NICU, FRG, stillbirth) across the three groups.

Statistical analysis

In order to examine clinical and biological information, we made use of Microsoft Excel software for the purpose of data storage, while we utilized Statistical Package for the Social Sciences(SPSS) v27.0 software(developed by International Business Machines Corporation) for conducting data analysis. The presentation of quantitative data involved the use of median values [Interquartile range], while the categorial variables were defined by frequencies and percentages. To compare different groups in terms of quantitative variables, we made use of non-parametric Kruskal–Wallis tests. We used Chi-square tests to compare groups for qualitative variables, unless the expected values were less than 5, in which case we employed Fisher tests. We applied the non-parametric Spearman rank-order test to determine the correlation between variables. The significance level was established as p values < 0.05. We performed post hoc tests on the coagulation parameters and pregnancy outcome indicators with statistical differences among the three groups. For indicators with statistical differences between L group and N group in pairwise post hoc tests, if they were binary variables, multivariate binary logistic regression analysis was performed, followed by subgroup analysis and forest plot was drawn. Linear regression analysis was performed if it was a numerical variable.

Results

Flowchart for patient selection

This retrospective study comprised 3198 pregnant women who gave birth at the First People's Hospital of Kunming between November 1, 2022, and July 31, 2023. Exclusions comprised: ①. Pregnant individuals with a first SARS-CoV-2 infection during the first trimester or other than 3 days before delivery, excluding n = 1692;②. Pregnant individuals acquiring SARS-CoV-2 in early pregnancy without being long-term COVID-19 patients or lost to follow-up, excluding n = 583;③. Pregnant individuals who had never been infected with SARS-CoV-2 outside the period from November 1 through November 30, 2022, excluding n = 106;④. Pregnant individuals with comorbidities including chronic hypertension, n = 35, and diabetes, n = 58; ⑤. Pregnant individuals affected by viral infections such as HIV/toxoplasmosis/rubella, excluding n = 7; ⑥. Pregnant individuals presenting thrombocytopenia, n = 3; ⑦. Pregnant individuals using aspirin or low-molecular-weight heparin during gestation, n = 68; and ⑧. pregnancy cases lacking available data, n = 23. Figure 2 illustrates the elimination process, 623 patients were included in this study. Group A: n = 209, group L: n = 72, group N: n = 342.

Figure 2
figure 2

Flow chart of patient selection.

Full size image

Demographic and clinical characteristics of the patients

The study included 623 women with a median age of 30 years, a median pre-delivery BMI of 24.7kg/m2, and a median number of pregnancies of 3. Patients with long-term COVID-19 during pregnancy accounted for 11% of patients with first SARS-CoV-2 infection during early pregnancy. Table 1 shows the demographic and clinical characteristics of all subjects. There were no significant differences in age, BMI before delivery, number of pregnancies, history of hypertension, history of diabetes, history of polycystic ovary, blood pressure threshold, blood glucose ≥ 5.6mmol/L and urine protein in early pregnancy among the three groups. However, the gestational age at SARS-CoV-2 infection differed significantly among the three groups (p < 0.001). During pregnancy, the most prevalent indications of Long-COVID-19 were cough (n = 24, 33.3%) and dyspnea (n = 20, 27.8%).

Table 1 Demographic and clinical characteristics of the patients.
Full size table

Coagulation parameters and maternal and fetal complications were compared among the three groups

Table 2 shows the characteristics of coagulation parameters and maternal and fetal complications among the three groups. There were significant differences in PLT among the three groups (p = 0.044). The maternal and fetal complications of HIP (p = 0.015), GDM (p = 0.013), cesarean section (p < 0.001), FGR (p = 0.01), and PD (0.015) were statistically different among the three groups. There were no significant differences in other indicators among the three groups. The results of pairwise post-hoc comparisons are as follows: Cesarean section: Group A exhibited significantly higher rates compared to Groups L and N; PD: Group A showed significantly higher rates than Group L; HIP: Group L exhibited significantly higher rates compared to Groups A and N; GDM: Group L showed significantly higher rates compared to Groups A and N; FGR: Group L showed significantly higher rates than Group N; PLT: Group L demonstrated lower levels than Group N. (Refer to Fig. 3 for details).

Table 2 The coagulation indexes and pregnancy outcomes of the three groups were descriptively analyzed.
Full size table
Figure 3
figure 3

The pregnancy outcomes and coagulation parameters of the three groups were compared between each other. Data are shown for healthy controls (n = 342; Group N: Light Gray), Long-COVID-19 patients (n = 72; Group L: gray) and patients with ongoing infection (n = 209; Group A: Dark grey). Black lines represent the median. *p < 0.05; **p < 0.01;***p < 0.001.

Full size image

Linear regression analysis of PLT in long-term COVID-19 pregnant women and non-COVID-19 pregnant women

Table 3 presents the results of the linear regression analysis comparing PLT between Group L and Group N. Compared with group N, for every 1% increase in the proportion of pregnant women with long-term COVID-19 during pregnancy, the PLT value in group L decreased by 0.127 × 109/L on average. After adjusting for age, number of pregnancies and BMI before delivery (β:  − 0.133, 95% CI − 0.002–0.000), the relationship between the two groups did not change even after adjusting for confounding factors.

Table 3 Linear regression analysis of platelet count between long-term COVID-19 pregnant women and non-COVID-19 pregnant women.
Full size table

Multivariate binary logistic regression model was used to analyze the relationship between COVID-19 pregnant women and non-COVID-19 pregnant women in GDM, HIP and FGR

Table 4 depicts the multivariate binary logistic regression models for the L group and N group with respect to GDM, HIP, and FGR. To verify the stability of the results, both non-adjusted and multi-variable adjusted models were employed. The selection of adjustment variables was based on three criteria(meeting one of these criteria was selected as an adjustment variable): (1) Inclusion of a variable that would alter the matched OR by a minimum of 10% when added to the model; (2) Variables exhibiting a correlation analysis p-value < 0.05; or (3) Variables deemed confounders according to existing literature and clinical assessment. The exclusion of variables displaying strong multicollinearity occurred. Through scrutiny of tolerance values and variance inflation factor tests, the variables in the ultimate model were determined to be free of multicollinearity. The adjustment for confounding factors failed to alter the association between Long-COVID-19 and linked illnesses (GDM (OR 2.735, 95% CI 1.431–5.229), HIP (OR 3.601, 95% CI 1.611–8.650), FGR (OR 2.593, 95% CI 1.190–5.648). This suggests that Long-term COVID-19 during pregnancy may independently contribute to the elevated risk of GDM, HIP, and FGR.

Table 4 Multivariate binary logistic regression model of gestational diabetes mellitus, gestational hypertension and fetal growth restriction in long-term COVID-19 pregnant women and non-COVID-19 pregnant women.
Full size table

Subgroup analysis of HIP, GDM and FGR in pregnant women with long-term COVID-19 and non-COVID-19

Figure 4 presents the forest plot depicting the results of subgroup analysis. The analysis of subgroups uncovered similarity in the relationship between long-term COVID-19 during pregnancy and HIP, GDM, and FGR across patient subgroups distinguished by age, pre-delivery BMI, and pregnancy count.

Figure 4
figure 4

Subgroup analysis of gestational hypertension, gestational diabetes mellitus and fetal intrauterine growth restriction between pregnant women with long-term COVID-19 and non-COVID-19 pregnant women.

Full size image

Discussion

In this retrospective cohort study, the proportion of pregnant women with long-term COVID-19 following their initial infection was 11%. Among this group, 15.3% experienced HIP, 30.6% had GDM, and 20.8% faced FGR. In comparison to non-COVID-19 pregnant women, long-term COVID-19 pregnant women exhibited a significant correlation with HIP, GDM, and FGR. Following adjustment for confounding factors using multivariate binary logistic regression analysis, the association remained robust, and the results were consistent even after conducting subgroup analysis of the three maternal and fetal complications. Our findings indicate that pregnant women with long-term COVID-19 exhibited a slightly decreased PLT in comparison to non-COVID-19 pregnant women. This correlation remained significant even after controlling for potential confounding factors through linear regression analysis. Nevertheless, no statistically significant variances were observed in other coagulation parameters. Furthermore, in pregnant women with acute COVID-19 infection, a higher incidence of cesarean section and PD was observed, while no statistically significant differences were found in other maternal and fetal complications such as PROM, neonatal asphyxia, and neonate transfer to NICU.

There have been limited studies on the long-term effects of COVID-19 during pregnancy, and the incidence rate of long-term COVID-19 during pregnancy in this study was 11%, which is consistent with the 10%-20% incidence rate observed in the adult population2. Persistent symptoms included cough and dyspnea as the most prevalent, followed by post-activity palpitations and fatigue, in line with the findings from several studies conducted on adult populations17,18. At present, the pathophysiological mechanisms and effective therapeutic interventions for long-term COVID-19 remain elusive. Nevertheless, several hypotheses have recently emerged. The main theories underlying the mechanisms include the presence of a persistent viral reservoir19, sustained inflammation20, host microbiome factors21,22, enduring autoimmune responses23, endothelial dysfunction, and subsequent microvascular dysfunction24. Multiple research studies have demonstrated the effects of Long-COVID-19 on various body systems1. An observational cohort study revealed that 70% of long-term COVID-19 patients exhibited at least one form of organ dysfunction, with mild damage observed in the lungs (11%), heart (26%), kidneys (4%), pancreas (40%), liver (28%), and spleen (4%). Additionally, 23% of patients experienced multi-organ damage25. These findings underscore the significant implications of long-term COVID-19 on individuals' quality of life and potential underlying health conditions. This study represents the first comprehensive statistical analysis of maternal and fetal complications as well as coagulation function in long-term COVID-19 pregnant women. The findings indicate a significant increase in the incidence of HIP, GDM, and FGR among long-term COVID-19 pregnant women. Among these, the increased incidence of HIP may be attributed to the similarity in pathogenesis between HIP and SARS-CoV-28,16, resulting in sustained endothelial injury and consequently an elevated occurrence of HIP13,15. The rising prevalence of GDM may elevate the risk of enduring complications for both maternal and fetal, such as obesity, impaired glucose metabolism, and cardiovascular disease26,27, which could be attributed to the pancreatic exocrine function impairment in pregnant women induced by SARS-CoV-2. Studies have indicated that SARS-CoV-2 might directly impact the pancreas through binding to the ACE2 receptor, leading to pancreatitis, diabetes, and impairment of exocrine pancreas function28,29. Furthermore, the continual expansion of viral databases and the presence of anxiety-induced negative emotions may also exert an influence on the metabolic processes of expectant mothers, potentially contributing to a heightened prevalence of GDM. To date, there have been no definitive reports of vertical transmission of COVID-19 from pregnant women to their fetuses30,31. Nevertheless, a substantial body of research indicates that acute SARS-CoV-2 infection in pregnant women heightens the risk of various adverse maternal and fetal outcomes, including PD, miscarriage, and FGR32,33,34,35. Moreover, research conducted by Shanes and Menter et al. has identified evident Poor placental perfusion in pregnant individuals with acute SARS-CoV-2 infection36,37. Hence, the heightened prevalence of FGR in expectant mothers with long-term COVID-19 may also be associated with impaired placental perfusion and persistent inflammation20, necessitating further investigation for validation. Furthermore, we observed a higher incidence of cesarean section and PD in women with acute COVID-19 compared to non-COVID-19 infected pregnant women. Similar to recent research on acute COVID-19 during pregnancy, a 2020 study demonstrated that PD accounted for approximately 20% of total deliveries among women with acute COVID-19 during pregnancy38. A 2021 meta-analysis involving 42,754 pregnant women revealed a high caesarean section rate of 53.2% among women with acute COVID-1939. Fortunately, similar to numerous studies on acute SARS-CoV-2 infection during pregnancy17,40, this study revealed that long-term COVID-19 during pregnancy did not lead to a more pronounced hypercoagulable state but did indicate a slight decrease in PLT. Numerous studies on adult COVID-19 patients have demonstrated that PLT levels are typically within the normal range or slightly elevated in most cases. However, the incidence of thrombocytopenia increases among severe patients and is correlated with disease severity and mortality rate41. Nevertheless, in this study, long-term COVID-19 pregnant women exhibited only a marginal decrease in PLT, which does not preclude the possibility of interaction with other factors such as HIP. Further research is warranted to investigate this.

The highlights of this study, to the best of our knowledge, this study represents the first comprehensive statistical analysis of maternal and fetal complications as well as coagulation parameters in long-term COVID-19 pregnant women. Furthermore, our study revealed that pregnant women with long-term COVID-19 are at an increased risk of experiencing pregnancy complications and unfavorable pregnancy outcomes.

Nevertheless, this study is subject to certain limitations. Firstly, being a single-center retrospective cohort study, it precludes the establishment of a potential causal relationship. Secondly, as a single-center retrospective study, our data collection was exclusively derived from the hospital's electronic medical record system and could be traced back to the original medical records. Additionally, three individuals conducted rigorous quality control on the data and follow-up to ensure its authenticity and reliability. However, there is still a certain lack of diversity in the samples and generality in the results. Finally, when assessing the influence of potential confounding variables on maternal and fetal complications, there may be unaccounted-for confounding factors. Nevertheless, our sensitivity analysis demonstrated the robustness of the conclusions across diverse subgroups.

In future investigations, we aim to conduct more comprehensive and extensive research across multiple hospitals in the region to enhance the generalizability and diversity of the study sample. Furthermore, we intend to prolong the follow-up period to assess the long-term impact on newborns and pregnant women. Lastly, we plan to incorporate pregnant women from diverse regions and countries to strengthen external validation, thereby affirming the consistency and reliability of our findings.

In this study, we observed that pregnant women with long-term COVID-19 are at an increased risk of experiencing pregnancy complications and adverse pregnancy outcomes. This necessitates obstetricians to proactively adjust their treatment plans in anticipation of potential risks. These findings provide valuable insights for healthcare professionals to comprehensively understand the impact of long-term COVID-19 on both maternal and fetal, enabling them to take proactive measures to prevent maternal and fetal complications, mitigate adverse pregnancy outcomes and related risks, as well as offer essential guidance for prenatal consultation and clinical management.

Conclusions

In conclusion, this study demonstrates that long-term COVID-19 infection in pregnant women does not result in further hypercoagulability; however, it does elevate the incidence of HIP, GDM, and FGR. Persistent vascular endothelial damage, multiple organ system damage, persistent inflammation, and placental malperfusion may be related factors. These findings are essential for informing appropriate prenatal counseling, facilitating comprehension of the risks associated with COVID-19 infection during pregnancy, averting pregnancy-related complications and adverse outcomes, and mitigating maternal and fetal risks.