Suy et al. BMC Pregnancy and Childbirth (2021) 21:78 https://doi.org/10.1186/s12884-021-03572-4 STUDY PROTOCOL Open Access Gestation and COVID-19: clinical and microbiological observational study (Gesta- COVID19) Anna Suy1, Itziar Garcia-Ruiz2, Melchor Carbonell1, Pablo Garcia-Manau1, Carlota Rodo1, Nerea Maiz1,3* , Elena Sulleiro4, Andres Anton4, Juliana Esperalba4, Nuria Fernández-Hidalgo5,6, Marie Antoinette Frick7, Fatima Camba8, Tomas Pumarola4, Elena Carreras1 On behalf of the Gesta-COVID19 Collaboration Group Abstract Background: The Coronavirus Disease 2019 (COVID-19) is a novel disease which has been having a worldwide affect since December 2019. Evidence regarding the effects of SARS-CoV-2 during pregnancy is conflicting. The presence of SARS-CoV-2 has been demonstrated in biological samples during pregnancy (placenta, umbilical cord or amniotic fluid); however, maternal and fetal effects of the virus are not well known. Methods: Descriptive, multicentre, longitudinal, observational study in eight tertiary care hospitals throughout Spain, that are referral centres for pregnant women with COVID-19. All pregnant women with positive SARS-CoV-2 real-time reverse transcriptase polymerase chain reaction during their pregnancy or 14 days preconception and newborns born to mothers infected with SARS-CoV-2 will be included. They will continue to be followed up until 4 weeks after delivery. The aim of the study is to investigate both the effect of COVID-19 on the pregnancy, and the effect of the pregnancy status with the evolution of the SARS-CoV-2 disease. Other samples (faeces, urine, serum, amniotic fluid, cord and peripheral blood, placenta and breastmilk) will be collected in order to analyse whether or not there is a risk of vertical transmission and to describe the behaviour of the virus in other fluids. Neonates will be followed until 6 months after delivery to establish the rate of neonatal transmission. We aim to include 150 pregnant women and their babies. Ethics approval will be obtained from all the participating centres. Discussion: There is little information known about COVID-19 and its unknown effects on pregnancy. This study will collect a large number of samples in pregnant women which will allow us to demonstrate the behaviour of the virus in pregnancy and postpartum in a representative cohort of the Spanish population. Keywords: SARS-CoV-2, COVID-19, Pregnancy, Vertical transmission, Amniotic fluid, Placenta, Cord blood, Breastmilk * Correspondence: nmaiz@vhebron.net 1 Department of Obstetrics, Hospital Universitari Vall d’Hebron, Barcelona, Spain 3 Maternal-Fetal Medicine Unit, Department of Obstetrics, Vall d’Hebron University Hospital, Ps Vall d’Hebron 119-129, 08035 Barcelona, Spain Full list of author information is available at the end of the article © The Author(s). 2021 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. 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BMC Pregnancy and Childbirth (2021) 21:78 Page 2 of 6 Background Given that COVID-19 is an emerging disease and our Severe Acute Respiratory Syndrome Coronavirus 2 knowledge of it evolves from day to day, authors will be (SARS-Cov-2) was first reported in the city of Wuhan, encouraged to add other variables to the protocol as China, in December 2019, with a rapid spread to the rest new clinical evidence comes forward, with the commit- of the planet within a few weeks. Spain reported its first ment to notify any changes in protocol to the Ethics case of the Coronavirus Disease 2019 (COVID-19) in Committee. February, 2020 and as of December 29th 2020, almost 2 million cases have been reported [1]. Objective Evidence regarding the effects of SARS-CoV-2 during To investigate both the effect of COVID-19 on pregnancy pregnancy is conflicting: A study on pregnant women in and the effect of pregnancy status with the evolution of the United States of America reported that percentages SARS-CoV-2 disease. of mild, moderate and severe disease for pregnant women are similar to that of the general population (80, Outcomes (Table 1) 15 and 5%, respectively) [2] whilst others suggest higher Primary outcome: rates of Intensive Care Unit (ICU) admission and oxygen supplementation in pregnant women above 20 weeks’  Rate of perinatal morbidity in pregnant women with gestation compared to non-pregnant population with COVID-19 (preterm delivery, preeclampsia, COVID-19 [3]. Many publications regarding the maternal- hospitalization during pregnancy, and admission to fetal effects of the virus have raised: a preeclampsia-like the ICU) syndrome in women affected by COVID-19 has been described [4] and transplacental transmission and possible Secondary outcomes: fetal effects of the virus have been demonstrated [5–7]. The presence of the virus in amniotic fluid and breast milk has  Description of COVID-19 maternal symptoms also been reported [6, 8]. However, evidence on the effects  Rate of maternal mortality of the SARS-CoV-2 infection during the first trimester of pregnancy is scarce, with only few publications reporting Table 1 Definition of primary and secondary outcomes low rates of miscarriage in the first trimester [9] In our Primary outcome centre we have identified patients with these characteristics, Preeclampsia, preterm delivery, Frequency of each complication / the obstetric and neonatal results of which will be of utmost need of hospitalization, ICU total number of pregnant women importance for the correct care and treatment of this admittance disease in pregnant women. Given the little evidence Secondary outcomes published so far regarding the effects that COVID-19 can Description of COVID-19 Frequency of each symptom / have on pregnancy and vice versa, it is essential to gather as maternal symptoms Total number of pregnant women much information as possible. Maternal mortality Number of maternal deaths due to We designed an observational study in 8 tertiary care COVID-19 or any other complication hospitals in Spain, that are referral centres for pregnant during the pregnancy / Total number of pregnant women women with COVID-19 during the pandemic. The study intends to investigate both the effect of COVID-19 on Rate of fetal morbidity Number of miscarriage, stillbirth, and mortality fetal malformation, intrauterine pregnancy, and the effect of pregnancy status with the growth restriction / Total number evolution of SARS-CoV-2 disease. of foetuses Description of behaviour of Number of positive tests / Number Methods/design the virus in biological fluids of tests performed (per each fluid (urine, faeces, blood cord, or tissue) Design and setting of the study placenta and breastmilk) This is a descriptive, multicentre, longitudinal, observa- Neonatal infection at 24, 48 h Number of newborns with positive tional study in eight tertiary care hospitals throughout and 7 days RT-PCR in pharyngeal aspirate at Spain, that are referral centres for pregnant women with 24, 48 h and at 7 days / Total COVID-19: Vall d’Hebron University Hospital (HUVH), number of newborns Barcelona (coordinating centre); La Paz University Neonatal mortality Number of neonatal deaths within Hospital (HULP), Madrid; Hospital Virgen de la Arrixaca the first 7 (early) and 28 days of life / Total number of foetuses/ (HUVA), Murcia; Hospital Universitario Cruces (HUC), newborns Bizkaia; Hospital Clínico Universitario Lozano Blesa Neonatal morbidity Number of each complication / (HCULB), Zaragoza; Hospital de Torrejón, Madrid; (pneumonia, NICU Total number of neonates Hospital Universitario San Cecilio, Granada; and Hospital admission, sepsis,…) Universitario La Fe, Valencia. ICU Intensive Care Unit, NICU Neonatal Intensive Care Unit Suy et al. BMC Pregnancy and Childbirth (2021) 21:78 Page 3 of 6  Rate of fetal mortality Samples to be collected (Fig. 1):  Rate of fetal morbidity (miscarriage, stillbirth, fetal malformation, and intrauterine growth restriction)  Pregnant women: SARS-CoV-2 RT-PCR in  Description of the behaviour of the virus in nasopharyngeal and oropharyngeal swab at first biological fluids (peripheral blood, urine, faeces, visit, weekly until SARS-CoV-2 RT-PCR negativization, blood cord, placenta, and breastmilk) and serological delivery and postpartum; SARS-CoV-2 RT-PCR in response amniotic fluid, urine, faeces, peripheral blood and  Rate of neonatal infection at 24, 48 h and 7 days serum for serologic tests, according to clinical criteria; after birth SARS-CoV-2 RT-PCR in amniotic fluid, placenta, cord  Rate of neonatal mortality blood, and breast milk at delivery and postpartum.  Rate of neonatal morbidity (infection, pneumonia, Serologic tests within 4–6 weeks after negative SARS- and admission to the ICU) CoV-2 RT-PCR results.  Newborns born to COVID-19 women: SARS-CoV-2 RT-PCR in nasopharyngeal aspirate, urine, and Participants and processes faeces after delivery. Serologic tests at delivery and All consecutive pregnant women with a confirmed after 30 days and 6 months postpartum. COVID-19 diagnosis by real-time reverse transcriptase  Newborns born to COVID-19 women requiring polymerase chain reaction (RT-PCR) will be included. admission to the Neonatal Unit: SARS-CoV-2 RT- Women attended in a labour ward or outpatient clinic PCR in nasopharyngeal aspirate and tracheal aspirate with SARS-CoV-2 symptoms (fever, cough, dyspnoea, (if intubated) after delivery, 24 h, 5 days, and 14 days anosmia, ageusia, diarrhoea, fatigue, myalgias) [10] or after birth. SARS-CoV-2 RT-PCR in urine, faeces those in close contact with a COVID-19 confirmed case, and serologic tests after delivery. Serologic tests after will undergo a SARS-CoV-2 RT-PCR in nasopharyngeal 30 days and 6 months postpartum. and oropharyngeal smears. They will be managed according the presence of mild The detection of SARS-CoV-2 RNA for any sample (fever, cough, anosmia, diarrhoea), moderate (tachyp- type will be performed by means of a commercial RT- noea >30xmin, hypoxia (saturation < 93% in room air at PCR-based assay Allplex™ 2019-nCoV (Seegene, Korea). sea level or PAO2/FiO2 < 300 mmHg) or abnormal chest The serological study consists in the determination of imaging (> 50% affected lung) or severe symptoms specific IgG antibodies for pregnant women and specific (respiratory failure, shock) [10, 11] Women with mild IgG and IgM antibodies in newborns. These tests will be symptoms will be managed as outpatients and called determined from serum samples using Liaison SARS- in daily for a symptoms’ evaluation until a SARS-CoV-2 CoV-2 S1/S2 IgG (DiaSorin, Italy) and Liaison SARS- RT-PCR negativization. Women requiring hospitalization CoV-2 S1-RBD IgM. Both of them will be performed on (with moderate or severe symptoms) will be managed ac- the LIAISON® XL Analyzer (DiaSorin, Italy). cording to gestational age and symptoms until discharged Data will be collected and encoded on an electronic from the hospital. data collection sheet (REDCap®). Each patient will have All women with a previous history of COVID-19 and an identification number for the study. The correlation negative RT-PCR SARS-CoV-2 will be followed monthly between the medical record number and the study iden- in the outpatient clinic until delivery and at 4 weeks tification number will be collected in a separate and postpartum. anonymized database. Delivery of positive SARS-CoV-2 women will be per- formed according the Spanish Guidelines for COVID-19 pregnancy and delivery [12, 13]. Neonates will not be Statistics separated from their mothers but women will be instructed A descriptive analysis for the primary and secondary regarding measures to prevent contagion of their sons. outcomes will be carried out, calculating absolute and Inclusion and exclusion criteria: relative frequencies. For the maternal characteristics’ description, median and interquartile range will be used  Inclusion criteria: women with SARS-CoV-2 RT- for continuous variables and absolute and relative PCR during pregnancy or 14 days preconception frequencies for categorical variables. and newborns born to mothers infected with Logistic regression analysis will be used to explore the SARS-CoV-2. association of adverse maternal, fetal and neonatal out-  Exclusion criteria: refusal to participate in the study, comes with each specific drug used to treat COVID-19. pregnant women under 18 years of age, and All analysis will be carried out with R software. Statis- difficulty to understand informed consent. tical significance will be set at p < 0.05. Suy et al. BMC Pregnancy and Childbirth (2021) 21:78 Page 4 of 6 Fig. 1 Gesta-COVID19 Study protocol. Pregnant women timeline (SPIRIT figure) Sample size Discussion Given the short evolution of this infection, this study aims The mother-foetus binomial is unique in medicine, to be exploratory and to make a quick characterization of morbidity and mortality can therefore affect both. This the described outcomes. To this end, it has been estimated prospective longitudinal study aims to collect all con- that within a period of 4 to 6 months (peak of the epi- secutive cases of COVID-19 in pregnant women in eight demic and subsequent months), we will be able to achieve referral centres throughout Spain, with the objective of a sample size of 150 pregnancies. defining whether the pregnancy implies a change in the prognosis of the infection, and vice versa, whether the Ethics infection impacts the pregnancy. Furthermore, we intend This study was approved by the Ethics Committee of the to assess vertical transmission. coordinating centre (PR(AMI)181/2020) and by that of The electronic data collection sheet, accessible via the all the participating centres. website from eight of the main reference hospitals in Patients will be informed about the study. Given the Spain, will allow us to obtain a representative sample of current situation of the pandemic and public health the pregnant Spanish population. concerns (in order to reduce the risk of contagion), the The major strength of this study is that weekly con- Ethics Committee accepted to obtain oral consent from secutive samples of the naso / oropharyngeal smears will the patient and to record it in the medical history. be obtained, allowing to infer virus clearance. Collecting Suy et al. BMC Pregnancy and Childbirth (2021) 21:78 Page 5 of 6 other samples apart from the naso / oropharyngeal Silvia Arevalo. Department of Obstetrics. Hospital Universitari Vall d’Hebron. smears (peripheral and cord blood, serum, faeces, urine, Barcelona. Berta Serrano. Department of Obstetrics. Hospital Universitari Vall d’Hebron. amniotic fluid, placenta, breast milk and serologic Barcelona. follow-up), will grant more information on the behav- Marina Catalan. Department of Obstetrics. Hospital Universitari Vall d’Hebron. iour of the virus in other biological fluids and investigate Barcelona. Helena Tur. Department of Obstetrics. Hospital Universitari Vall d’Hebron. whether or not there is risk of perinatal transmission. Barcelona. Due to the limited information on COVID-19 in preg- Maria de la Calle. Department of Obstetrics. Hospital Universitario La Paz. nancy to date, the limitation in the management and Madrid. Juan Luis Delgado. Department of Obstetrics. Hospital Virgen de la Arrixaca. treatment of pregnant women, and the possibility of a Murcia. new outbreak when confinement measures will relax, it Leire Rodriguez. Department of Obstetrics. Hospital Universitario Cruces. is necessary to collect all possible evidence to establish Barakaldo. Sara Ruiz-Martínez. Department of Obstetrics. Hospital Clínico Universitario management protocols and working dynamics to effi- Lozano Blesa. Zaragoza. ciently react when it happens. It is the clinicians’ respon- Francisca Sonia Molina. Department of Obstetrics. San Cecilio University sibility to be updated on COVID-19 in order to be able Hospital. Instituto de Investigación Biosanitaria (IBS). Granada. Vicente Diago. Department of Obstetrics. Hospital Universitario La Fe. Valencia. to offer the best care to their patients. Silvia Mateos. Department of Obstetrics. Hospital de Torrejón. Madrid. This work has been done within the framework of the Autonomous Abbreviations University of Barcelona. SARS-CoV-2: Severe acute respiratory syndrome coronavirus 2; COVID- 19: Coronavirus Disease 2019; SARS-CoV: Severe Acute Respiratory Syndrome Authors’ contributions Coronavirus; MERS-CoV: Middle East Respiratory Syndrome Coronavirus; AS, IGR, MC, PGM, MAF, FC and NFH designed the study and collected HUVH: Vall d’Hebron University Hospital; HULP: La Paz University Hospital; samples. ES, AA, JE and TP conserved and analysed samples. NM described HUVA: Hospital Virgen de la Arrixaca; HUC: Hospital Universitario Cruces; statistical analysis. CR, NM and EC wrote the manuscript. All authors read and HCULB: Hospital Clínico Universitario Lozano Blesa; RT-PCR: Real-time reverse approved the final manuscript. transcriptase polymerase chain reaction; EC: Ethics Committee Funding The study has been awarded with a Spanish Government grant (Instituto de Acknowledgements Salud Carlos III: COV20/00188). It covers the expenses for transporting, Gesta-COVID19 Collaboration Group: processing and analyzing samples, database creation and maintenance, and Anna Suy. Department of Obstetrics. Hospital Universitari Vall d’Hebron. hiring support staff. Itziar García-Ruiz. Department of Obstetrics. Hospital Universitari Vall d’Hebron. Universitat Autònoma de Barcelona. Melchor Carbonell. Department of Obstetrics. Hospital Universitari Vall Availability of data and materials d’Hebron. Barcelona. The datasets used and/or analysed during the current study are available Pablo Garcia-Manau. Department of Obstetrics. Hospital Universitari Vall from the corresponding author on reasonable request. d’Hebron. Barcelona. Carlota Rodo. Department of Obstetrics. Hospital Universitari Vall d’Hebron. Ethics approval and consent to participate Barcelona. The Vall d’Hebron University Hospital Ethics Committee (EC), as the reference Nerea Maiz. Department of Obstetrics. Hospital Universitari Vall d’Hebron. Ethics Committee, first approved this study. The reference number is PR(AMI)181– Barcelona. 2020. All of the participating centres also obtained their own EC approval. Elena Sulleiro. Department of Microbiology. Hospital Universitari Vall The protocol, informed consent form, participant information sheet and any d’Hebron. Barcelona. applicable documents were submitted to the respective Ethics Committees Andres Anton. Department of Microbiology. Hospital Universitari Vall and regulatory authorities, and written approval was obtained. All substantial d’Hebron. Barcelona. amendments to the originally approved documents will be sent to the Juliana Esperalba. Department of Microbiology. Hospital Universitari Vall respective authorities for approval. The study did not begin until the d’Hebron. Barcelona. approval of the EC and Director’s consent was obtained. Given the current Nuria Fernández-Hidalgo. Infectious Diseases Department. Hospital pandemic and public health concerns, it was accepted to obtain oral Universitari Vall d’Hebron. Universitat Autònoma de Barcelona, Barcelona, consent from the patients and to record it in their medical history. Spain. Red Española de Investigación en Patología Infecciosa (REIPI). Instituto de Salud Carlos III, Madrid, Spain. Consent for publication Marie Antoinette Frick. Department of Pediatric Infectious Diseases and Not applicable. Immunodeficiencies Unit. Hospital Universitari Vall d’Hebron. Barcelona. Fatima Camba. Department of Neonatology. Hospital Universitari Vall Competing interests d’Hebron. Barcelona. The authors declare that they have no competing interests. Tomas Pumarola. Department of Microbiology. Hospital Universitari Vall d’Hebron. Barcelona. Author details 1 Elena Carreras. Department of Obstetrics. Hospital Universitari Vall d’Hebron. Department of Obstetrics, Hospital Universitari Vall d’Hebron, Barcelona, Barcelona. Spain. 2Department of Obstetrics, Hospital Universitari Vall d’Hebron, Manel Mendoza. Department of Obstetrics. Hospital Universitari Vall Universitat Autònoma de Barcelona, Barcelona, Spain. 3Maternal-Fetal d’Hebron. Barcelona. Medicine Unit, Department of Obstetrics, Vall d’Hebron University Hospital, Lidia Illan. Department of Obstetrics. Hospital Universitari Vall d’Hebron. Ps Vall d’Hebron 119-129, 08035 Barcelona, Spain. 4Department of Barcelona. Microbiology, Hospital Universitari Vall d’Hebron, Barcelona, Spain. 5Infectious Lorena Fernandez-Blanco. Department of Obstetrics. Hospital Universitari Vall Diseases Department, Hospital Universitari Vall d’Hebron, Universitat d’Hebron. Barcelona. Autònoma de Barcelona, Barcelona, Spain. 6Red Española de Investigación en Francisco José Barranco. Department of Obstetrics. Hospital Universitari Vall Patología Infecciosa (REIPI), Instituto de Salud Carlos III, Madrid, Spain. 7 d’Hebron. Barcelona. 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